SEMINARIO EL CO2 EN EL CALENTAMIENTO GLOBAL
INTRODUCCION
Antes de la Era Industrial, cerca 1750, la concentración de dióxido de carbono en la
atmósfera (CO2) era de 280 + 10 ppm; pero para 1999 se tenían aproximadamente 367 ppm. Las
concentraciones de CO2 en la atmósfera han sido medidas directamente con presición desde
1957 (IPCC, 2001).
El incremento de CO2 en la atmósfera en la actualidad es causado por emisiones
antropogénicas. Cerca de tres cuartas partes de estas emisiones son debidas a la quema de
combustibles fósiles. La quema de combustibles fósiles (más una pequeña contribución de
producción de cemento) emitió durante 1980 a 1989 en promedio 5.4 + 0.3 toneladas metricas
por año, y 6.3 + 0.4 toneladas metricas por año durante 1990 a 1999. El cambio de uso de la tierra
es responsable de el resto de las emisiones ((IPCC, 2001).
La concentración de CO2 en la atmósfera es regulada por procesos naturales que
intercambian carbono entre la atmósfera, el océano y la biota del suelo. El estudio del ciclo del
carbono permite entender y cuantificar los mecanismos, mediante los cuales se transfieren
carbono entre y dentro de estos reservorios. Un buen entendimiento del ciclo del carbono es
necesario para predecir los niveles de de CO2, y por consiguiente el cambio climático futuro. Los
modelos climáticos auguran un importante aumento de la temperatura (1.5 a 4.5°C globalmente)
con el doble de CO2. El océano con el tiempo absorverá más del CO2 que se encuentra en exceso
en la atmósfera, y el CO2 atmosférico lentamente se estabilizará hacia una concentración
asintótica por arriba de los valores pre-industriales. Pero, el alcance de este nuevo equilibrio
tomará siglos después de que las emisiones de combustibles fósiles sean detenidas. Las próximas
décadas, sin embargo, tanto las plantas como los suelos jugarán un papel muy importante para
ayudar al océano a controlar el CO2. Si los árboles en primera instancia toman ventaja en el
aumento de CO2 para crecer, la biosfera del suelo ganará carbono y disminuirán los incrementos
de CO2 (Rodó y Comín, 2003).
El exceso de CO2 atmosférico, debido principalmente a la combustión de combustibles fósiles y
un Cambio en el Uso de la Tierra, es un hecho indiscutible. Este continúo aumento de CO2 ha
tenido importantes consecuencias para la vegetación. El crecimiento de las plantas es
incrementado por excesos de CO2. Un incremento de CO2 puede resultar en un incremento en la
producción primaria( Schimel, 1995) y la subsecuente asignación y utilización.
Para mitigar las concentraciones de CO2 en la atmósfera se han desarrollado nuevas
estratégias y políticas apropiadas para el manejo de la agricultura y los bosques. Una opción se
basa en la captura de carbono en los suelos o biomasas terrestres, sobre todo en las tierras usadas
para la agricultura o la forestación. A partir del Protocolo de Kyoto esto se conoce como Uso de la
Tierra (LU), Cambio en el Uso de la Tierra y Forestación (LULUCF) y concierne a los artículos
1.3 y 1.4 del Protocolo de Kyoto (IPPC, 2000).
La toma de acción para la captura de carbono bajo el Protocolo de Kyoto u otro tratado postKyoto no solo estimulará cambios importantes en el manejo del suelo sino que también, por
medio de un incremento en el contenido de materia orgánica tendrá efectos significativos directos
en sus propiedades y un impacto positivo sobre las cualidades ambientales o agrícolas y sobre la
biodiversidad. Las consecuencias incluirán una mayor fertilidad del suelo y productividad de la
tierra para la producción de alimentos y para la seguridad alimentaria. Esta herramienta
económica también hará que las practicas agrícolas sean más sostenibles y ayudará a prevenir o
mitigar la degradación de los recursos de la tierra (FAO, 2001).
Este es un trabajo para conocer a grandes rasgos el efecto del CO2 en el calentamiento
global, y como afecta este a los microorganismos
CARBONO Y DIOXIDO DE CARBONO
El carbono es un no metal que se presenta en dos formas naturales: cristalina (diamante y
grafito) y amorfa (Hulla, antracita, lignito y turba) y estas variedades alotrópicas se han podido
obtener de manera artificial (Alcantara, 1992).
El carbono se encuentra libre en la naturaleza y combinado forma varios millones de
compuestos. Las sustancias como el petróleo y el gas natural contienen carbono. Todas las plantas
y los animales, incluyendo al hombre, contienen carbono en infinita variedad de compuestos
(Alcantara, 1992).
El carbono es el elemento básico de carbohidratos, grasas, proteínas, ácidos nucleicos
(como el DNA y RNA) y otros compuestos orgánicos necesarios para la vida (Miller, 1994).
Los principales compuestos del carbono que se convierten en agentes contaminantes son el
monóxido de carbono, el dioxido de carbono, el dioxido de azufre, los hidrocarburos, los
aldehídos y su interrelaciones quimicas con óxidos de nitrógeno, y otras muchas partículas , por
ejemplo, las de fertilizantes y sustancias radiactivas (Alcantara, 1992).
Monóxido de Carbono: Es el producto de combustiones incompletas de sustancias orgánicas, las
cuales ocurren cuando la cantidad de oxígeno es insuficiente:
Compuestos orgánicos + O2 → CO + H2 O
2CO + O2 → CO2
El CO se produce en los escapes de los automóviles, autobuses y camiones, en las emanaciones de
combustibles de plantas eléctricas, calentadores, enfriadores, etc.
Dioxido de Carbono: Es el producto normal de todas las combustiones de los compuestos
orgánicos.
CICLO DEL CARBONO
El ciclo del carbono está estrechamente ligado al flujo de energía, debido a que las
principales reservas de energía de los organismos son compuestos de carbono reducidos que han
derivado de la fijación del bióxido de carbono atmosérico ya sea por medio de la fotosíntesis o,
con mucho menor frecuencia, de la quimiosíntesis (Miller, 1994). En el Cuadro 1 se se pueden
apreciar las cantidades de carbono localizadas en la Tierra:
Cuadro 1: Cantidad de carbono presente en varias formas en la tierra
Forma en que se halla el carbono
Masa de carbono (1018)
Carbonato de Ca (La mayor parte en las rocas sedimentarias)
35000
Carbonato de Ca+Mg (la mayor parte en las rocas sedimentarias)
25000
Materia orgánica sedimentaria (como kerógeno)
15000
Carbonato y bicarbonato, disuelto en el océano
42
Combustibles fósiles, recuperables (carbón y petróleo)
4.0
Carbón (humus, caliche, etc)
3.0
Bióxido de carbono atmosférico
0.72
En todos los seres vivos (plantas y animales)
0.56
El ciclo del carbono global: storages
El ciclo del carbono en el suelo
Los productores primarios en el suelo son principalmente las plantas superirores, aunque
las cianobacterias y a un grado menor las algas eucariótas, pueden ser importantes en algunas
situaciones como los arrozales o los suelos erosionados y durante la formación del suelo. Por lo
general, el bióxido de carbono en la litosfera está a concentraciones mayores que en el aire, pero
hay intercambio entre los dos por difusión a través de los espacios del aire del suelo y por medio
del equilibrio entre el agua del suelo y el bicarbonato (Campbell, 1987).
La materia orgánica llega al suelo a partir de los productores, ya sea de sus hojas, raíces y
tallos muertos (conocidos como hojarasca) o de los exudados de las raíces. Se puede considerar
que hay tres fuentes principales de materia orgánica en el suelo: insolubles, solubles y
microbianas. El carbono insoluble incluye la celulosa, y lignina de las paredes celulares vegetales,
quitina de los exoesqueletos de artrópodos y las paredes de algunos hongos, así como otras
sustancias que requieren desdoblamiento enzimático antes de que produzcan metabolitos
utilizables. El humus es parte del material insoluble. El carbono soluble es aquel que se encuentra
en una forma inmediatamente disponible para otros organismos y puede ser liberado por
organismos vivos, como las raíces de una planta o después de la descomposición de los
productores primarios, consumidores y los mismos microbios degradadores. Una parte
considerable del carbón soluble es temporalmente inmovilizado en las células de los
microorganismos desintegradotes y su concentración por lo general es baja (Campbell, 1987).
El ciclo del carbono en el océano
El dióxido de carbono es ràpidamente soluble en agua. Algo de este CO2 disuelto,
permanece en el mar, y parte es removido por lo productores fotosintetizantes. Cuanto más
caliente esté el agua, mayor es la cantidad de los gases de dióxido de carbono y oxígeno disueltos
que regresan a la atmósfera.
En los ecosistemas marinos, algunos organismos toman moléculas disueltas de CO2 o
iones carbonato (CO3-2) del agua del océano y forman carbonato de calcio ligeramente soluble
(CACO3) para elaborar conchas, rocas y esqueleto de los organismos marinos, desde minúsculos
protozoarios hasta corales. Cuando los organismos aconchados mueren, partículas finas de sus
conchas y huesos caen lentamente a las profundidades del océano, y son enterradas durante eones
(o tiempos muy largos) en los sedimentos del fondo.
El carbono en estos sedimentos profundos del océano reingresa al ciclo muy lentamente,
cuando parte de los sedimentos se disuelvan y formen el dióxido de carbono disuelto que puede
entrar a la atmósfera
El ciclo del Carbono antes de la Revolución Industrial
El océano es con mucho la reserva de carbono más grande cerca de 40000 ton de C.
pools
over
El ciclo del Carbono Después de la Era Industrial
Especialmente desde 1950, cuando la población del mundo y el uso de recursos ha
aumentado rápidamente, hemos intervenido en el ciclo del carbono principalmente de dos
maneras:


Eliminación de bosques y otras vegetaciones sin replantación suficiente, lo que deja menos
vegetación para absorber CO2.
Utilización de combustibles fósiles que contienen carbono y combustión de madera más
ràpido de lo que puede volver a reproducirse. Esto produce CO2 que fluye a la atmósfera.
Algunos cientificos proyectan que este dióxido de carbono, junto con otras sust
Impactos futuros del cambio climático sobre el Ciclo del Carbono
En el Océano
La cantidad total de carbono en el océano es 50 veces más que la que se encuentra en la
atmósfera, y es intercambiada con la atmósfera a gran escala desde hace cientos de años.
La respuesta a largo plazo del ciclo del carbono en el océano involucra un cambio
climático en las temperaturas de la superficie del mar, en la biología del océano y en la circulación
del carbono y nutrientes disueltos. Estos procesos no son independientes uno del otro. Por
ejemplo, disminuye la oceanografía de las aguas profundas, rica en nutrientes, y esto puede tener
dos consecuncias puestas. Primeramente, se reducirá la presión parcial de CO2 por la de
oceanografía de aguas supersaturadas a la superficie, el proceso mediante el cual el agua del mar
profundo sube a la superficie upwelling For instance involves changing term response deeper
Ecosistemas terrestres
EL DIOXIDO DE CARBONO Y EL EFECTO INVERNADERO
En un invernadero, el aire se encuentra caliente comparado con el exterior. Este efecto se
debe a la energía solar, que en forma de radiaciones ultravioleta visibles, pasa a través deñ vidrio
y es absorbida por las plantas y objetos que hay dentro del invernadero. Cuando el interior
absorbe la radiación, ésta se convierte en calor; este calor no puede salir a través del vidrio y se
concentra en el interior. Esto se conoce como efecto inverndero.
Eldióxido de carbono, CO2,actúa de la misma forma que el vidrio de los invernaderos. La energía
luminosa y ultravioleta que alcanza la tierra evidentemente es capaz de atravesar la atmósfera,
pero después que es absorbida por la tierra y desprendida como energía calorífica, es absorbida
por el dióxido de carbono CO2, calentando la atmósfera. Sin este efecto de invernadero, la
atmósfera estaría mucho mas fría de que se encuentra. La temperatura promedio de la superficie
de la tierra sería mucho menor si no existiera el efecto invernadero. El vapor de agua y las nubes,
junto con el dióxido de carbono, contribuyen al efecto invernadero.
Sin embargo, como el CO2 es producto de las combustiones, existe la posibilidad de que este
efecto se vuelva pernicioso. En efecto, la tala de árboles, el constante aumento de la quema de
combustibles fósiles, entre otros factores, alteran la cantidad de dióxido de carbono en l
atmósfera, haciendo a ésta más caliente, lo cual, a la larga, tendría efectos nocivos.
El sobrecalentamientode la Tierra es una posibilidad real. Y sus efectos serían tan reales como
dramaticos. Normalmente, el calor absorbido por el suelo durante el verano se disipa a lo largo del
otoño e invierno; po reso las ultimas semanas de invierno y las primeras de primavera suelen ser
bastante frías.
Sin embargo, el aumento del CO2 atmósferico, al intensificar el efecto invernadero, impide que se
disipe todo el calor y que, al verano siguiente, la gente sienta que “hace mas calor que el año
pasado”. De este modo va aumentando la cantidad de calor atmosférico, lo cual podría provocar
grandes deshielos en las zonas árticas y antárticas, aumentar el nivel de las aguas océanicas e
inundar las zonas costeras continentales.
Calentamiento Global
Adjunto al cambio en las propiedades del litter, los efectos de las concentraciones de los gases de
invernadero han sido pronosticadas en los modelos de circulación atmosférica global y hay un
consenso general que el efecto primario de un calentamiento global será de 4°C. Por
consiguiente ambos aspectos no pueden ser disociados. Agreement
Los experimentos de calentamiento de suelo en ecosistemas naturales han servido para
generar datos de algunas preguntas importantes incluyendo los efectos del calentamiento del
suelo sobre los procesos biquímicos (por ejemplo descomposición, intemperismo del material
parental, ciclo del nitrogeno y traza de gas de emisión), la reacción de aquellos cambios en estos
procesos puede tener un cambio climático sobre la atmósfera. buried in addition dirigir ardes
litter
APORTE DE CO2 EN LA ATMOSFERA POR MICROORGANISMOS
La función más importante de la flora microbiana es la degradación de materiales
orgánicos. El número y diversidad de compuestos disponibles para degradación microbiológica es
enorme. El conjunto de ácidos orgánicos, polisacáridos. Ligninas, hidrocarburos alifáticos y
aromáticos, azúcares, alcoholes, aminoácidos, purinas, pirimidina, proteínas, lípidos y ácidos
nucleicos es atacada por una u otra población.
El CO2 que emite el suelo es generado por la respiración de las raíces de las plantas y los
organismos, tales como bacterias, hongos, gusanos e insectos (Maddock, 2004).
La tasa a la cual el CO2 es liberado durante la mineralización del humus varía
ampliamente con el tipo de suelo. Bajo condiciones de laboratorio controladas y a temperaturas en
el intervalo mesofílico, 20-30°C, la tasa de producción de CO2 generalmente es de 5 a 50 mg de
CO2 por Kg de suelo por día, pero se pueden encontrar en ocasiones 300 mg o más. En el campo,
la tasa de formación de CO2 puede ser tan baja como 0.5 o mayor de 10 g de CO2 por metro
cuadrado por día y algunas veces se encuentran valores tan altos como 25 (Alexander, 1980).
Interacciones entre el enriquecimiento de CO2 atmósférico y la fauna del suelo
Es ampliamente aceptado que los incrementos de CO2 en la atmósfera, y otros gases de
invernadero, modificaran la fisiología de los ecosistemas. En los ecosistemas terrestres, los
efectos de este incremento afectaran al ciclo del carbono, y por ende la traslocación del C dentro
de la planta y su transformación por los microorganismos (Madeleine, Bolger, 2000)
Montealegre et al., 2002, investigaron la influencia del incremento de CO2 en la atmósfera
sobre el número y actividad de las bacterias y las comunidades microbianas del suelo en
paztizales bajo atmosferas enriquecidas con dioxido de carbono (FACE). La composición de las
comunidades de los microorganismos, en la rizosfera y un volumen de suelo, bajo dos niveles de
CO2 bulk free-air Los cambios en la actividad microbiana, número y composición de la
comunidad probablemente ocurren bajo elevadas concentraciones de CO2, pero la magnitud de
esos cambios va ha depender de la especie de planta y la distancia a la cual los microorganismos
esten de las inmediaciones de la las raices de las plantas (Montealegre, 2002).
Hay algunas evidencias que los hongos son estimulados más que las bacterias cuando son
sometidas a elevadas concentraciones de CO2 (Grayston et al, 1998; Rillig et al., 1999) y
diferentes integrantes de las comunidades bacterianas pueden ser afectadas en forma distinta
(Marilley et al., 1999; ELhottova et al., 1997; Phillips et al., 2002). Sin embargo, otros estudios no
indicaron algún efecto significativo en la estructura de las comunidades microbianas (Zak et al;
2000b) .
Muchos reportes presentados, demuestran que a elevadas concentraciones de CO2 se
incrementa el porcentaje de infección de las micorrizas vesiculo arbuscular (VAM) como se
puede observar en el siguiente cuadro:
Cuadro 2: Efecto de elevadas concentraciones de CO2 sobre
raices.
la Colonización micorrizas de
Especie
Micorriza
Condiciones
crecimiento
de Porcentaje
de infección
Referencia
Pascopyrum smithii
VAM
Cárama de crecimiento
I
Monz et al
(1994)
Trifolium repens
VAM
Camara de crecimiento
I
Jongen et al.
(1996)
Liriodendron tulipifera
VAM
Camara abierta de arriba
SE
O.Neill et al
(1991)
Beilschmiedia pendula
VAM
Camara abierta de arriba
I
Lovelock
al. (1996)
Pinus taeda
EM
Invernadero
SE
Lewis et al
(1994)
Pinus echinata
EM
Cárama de crecimiento
I
O’Neill et al
(1987)
Pinus radiata
EM
Cárama de crecimiento
SE
Conroy et al
(1990)
Pinus caribaea
EM
Cárama de crecimiento
SE
Conroy et al
(1990)
Pinus silvestres
EM
Cárama de crecimiento
I
Ineiche et al
(1995)
Pinus silvestres
EM
Cárama de crecimiento
SE
Perez-Soba et
al (1995)
Quercus alba
EM
Cárama de crecimiento
I
O’Neill et al
(1987)
Fuente: Luo, Y., y Mooney, H.A. (1999)
VAM: Micorriza vesiculoarbuscular ; EM: Ectomicorriza SE:Sin efecto I: Incrementó
et
El cuadro anterior es una síntesis elaborada por Luo, Y., y Mooney, H.A. (1999) donde
mencionan que se tiene que tener cuidado en la interpretación de los datos de porcentaje de
infección, ya que solo se usaron datos de porcentajes de infección para evaluar el grado de
coliniaciòn micorrizal, pero no la función total del hongo simbiote, la cual es más importante en
términos de interacción entre plantas y el componente fúngico. Altos porcentajes de infección no
significa una alta cantidad de hifa microrrizal, y viscevesa. Medir la logitud hifal de las micorrizas
o biomasa es necesario para resolver este problema.
Ronn et al., 2003 midieron el efecto de elevadas concentraciones de CO2 durante el
desarrollo del trigo (Triticum aestivum cv. Minaret), y sobre las poblaciones de bacterias y
protozoarios del suelo, encontrando que no hubo efecto de la concentración de CO2 en el número
de bacterias, pero el número de protozoarios fue alto en los tratamientos con elevadas
concentraciones de CO2.
Ebersberger et al., 2004 investigaron sobre la estructura de las comunidades microbianas
en pastizales de suelos calcareos que han sido expuestos a elevadas concentraciones de CO2 (600
microlitros.L-1) por seis estaciones de crecimiento. Encontrando que en conjunto, solo se
detectaron efectos pequeños sobre la estructura de las comunidades microbianas, corroborando
previamente que la entrada de carbono al suelo probablemente cambia mucho menos que en la
respuesta fotosintética de la planta.
La tendencia general en la fauna del suelo respecto a los cambios en el clima son difíciles
de predecir porque hay pocos datos y los que existen son generados en experimentos a corto
plazo, en areas restringidas y condiciones artificiales. Extrapolaciones a largo plazo y escalas
espaciales deben por lo tanto hacerse con precaución porque la fauna del suelo abarca un rango
amplio de especies en términos de talla, ciclo de vida, escala de acción, adaptación o potencial de
migración que no es bien conocida actualmente, debido a que están interaccionando intimamente
con la vegetación y la microflora del suelo.
La mayoría de los aspectos del efecto del cambio global sobre la fauna del suelo es
pobremente explorada bajo condiciones experimentales y hay un gran rango de prioridades para
desarrollar y que puede sobrepasar las especulaciones actuales.
Respuesta de las plantas a elevadas concentraciones atmosféricas de CO2.
En numerosos estudios se ha reportado que las plantas a elevadas concentraciones de CO2
incrementan su producción de materia seca y algunas veces hacen un uso eficiente del agua
(Tubiello et al, 1999). Kang et al, 2002, trabajaron con tres cultivos: trigo, maíz y algodón,
dichos cultivos se sometieron a tres contenidos de humedad: alto (85 – 100% de Capacidad de
Campo, θF), medio ( 65 – 85% θ) y bajo ( 45 – 65% ); y a dos concentraciones de CO2, baja (
350 microlitros. L-1) y alta (700 microlitros.l-1), al final de este estudio se llegó a la conclusión
que al menos a corto plazo, la plantas C3 tales como el algodón y el trigo pueden beneficiarse de
las concentraciones de CO2 especialemente bajo condiciones de escazes de agua
•
Las elevadas concentraciones de CO2 en la atmósfera probablemente aumentarán la
fotosíntesis y crecimiento de las plantas, lo que provocará un incremento en los indices de
respiración Sin embargo, en algunos casos los indices de respiración de los tejidos de las plantas
se reducen cuando son expuestas a altas concentraciones de CO2, debido a a los efectos directos
sobre las enzimas y efectos indirectos derivados de los cambios en la composición química de la
planta (Gonzales, 2004) .
Gonzales, et al ( 2004) realizaron un revisión bibliográfica sobre la respiración de la
planta y elevadas concentraciones de CO2 en la atmósfera y contrario a lo que pensaban, los
indices de respiración generalmente no se reducen cuando las plantas se someten a altas
concentraciones de CO2. Sin embargo todos los ecosistemas estudiados muestran que la
respiración del dosel no se incrementa proporcionalmente al incrementarse la biomasa en
respuesta al aumento de la concentración de CO2, aunque una gran proporción de la respiración se
lleva a cabo en las raíces. El conocimiento fundamental de cómo la respiración y el soporte de los
procesos son controlados fisiológicamente aún es insuficiente, por esta razón se toman medidas
acertadas para interpretar como ciertas plantas responden al proceso de respiración cuando se
eleva la concentración de CO2. Por lo tanto el papel que juega la respiración de la planta en el
aumento de la capacidad de fijacion de los ecosistemas terrestres es aún incierto.
La respiración es esencial para el crecimiento y mantenimiento de todos los tejidos de
las plantas y juega un papel importante en el balance de carbono en las células, así como también
en el ciclo del carbono global.
Los ecosistemas terrestres intercambian cerca de 120 toneladas de carbono por año con la
atmósfera a través de los procesos de fotosíntesis y respiración (Schlesinger, 1997).
Aproximadamente la mitad del CO2 es asimilado anualmente a través de la fotosíntesis y el
demás es liberado a la atmósfera por la respiración de las plantas (Gifford, 1994 ; Amthor, 1995
Efecto del CO2 sobre la descomposición de la materia orgánica y el ciclo del
nitrógeno
Hay una incertidumbre acerca de como los ciclos del carbono y nitrógeno cambiaran con
una acumulación de CO2 en la atmósfera. Zak et al., 2000 resumieron datos de 47 publicaciones
de los ciclos del carbono y nitrógeno en el suelo bajo elevadas concentraciones de CO2 en un
intento por generalizar si los índices disminuirán, incrementaran o no cambiaran. Esta síntesis se
centra sobre cambios en la respiración del suelo, respiración microbiana, biomasa microbiana,
mineralización total del N, imbolización microbiana y mineralización neta de nitrógeno, porque
estas reservas y procesos representan puntos de control importante para el flujo de C y N interno.
Determinar si las diferencias en la asignación de C entre las formas de vida de la planta afectan
el ciclo del carbono y nitrógeno del suelo de manera predecible, los autores sintetizaron las
respuestas en gramineas, herbáceas y árboles que crecen en ambientes con elevadas
concentraciones de CO2. Las reservas internas y los procesos que sintetizaron se caracterizaron
por un alto grado de variabilidad (coeficiente de variación de 80-800%), hacer generalizaciones
dentro y entre formas de vida de la planta es difícil. En pocas excepciones, los indices de
respiración microbiana fuero más rápidos bajo concentraciones elevadas de CO2, indicando que
1) generalmente el crecimiento de planta bajo elevadas concentraciones de CO2 incrementa la
cantidad de C en el suelo, y 2) un sustrato adicional fue metabolizado por los microorganismos
del suelo. Sin embargo, la biomasa microbiana, la mineralización total del N, inmobilización
microbiana y la mineralización neta de N son caracterizadas por un gran incremento y
disminución bajo concentraciones elevadas de CO2, contribuyendo a un alto grado de varibilidad
dentro y entre las formas de vida de planta. De este estudio ellos concluyeron que hay datos
insuficientes para predecir como la actividad microbiana y los indices del ciclo del nitrogeno y
carbono cambiaran al aumentar las concentraciones de CO2 en la atmósfera.Argumentando que
en la actualidad hay lagunas para entender la biología de las raíces finas que limita su habilidad
para predecir la respuesta de los microorganismos al incrementar el CO2 en la atmosfera, y
entender las diferencias de longevidad de las raíces finas y su bioquímica entre especies de plantas
es necesario para desarrollar un modelo que pronostique los cambios en el ciclo del N y C bajo
elevadas concentraciones de CO2 atmosférico.
El cambio climático afectará directamente la mineralización del carbono y nitrogeno a
través de los cambios de temperatura y humedad del suelo, pero indirectamente puede afectar los
indices de mineralización a través de los cambios en la calidad del suelo. Keller et al (2004)
realizaron un experimento en un mesocosmo para examinar los efectos de seis años de rayos
infrarrojos (calentamiento) y láminas de agua sobre el potencial anaerobico del nitrogeno y
carbono water-table donde in bog fen peat frared loading pathways
La predicción de que la calidad del matillo, y por lo tanto los indices de descomposición
del mantillo, pueden reducir cuando las plantas crecen en atmósferas enriquecidas con CO2 ha
sido basada en la observación de la concentración de N foliar que generalmente es baja en
elevadas concentraciones de CO2. Norby, et al (2000) evaluaron el supuesto que el proceso de
resorción de nutriente estacional es diferente en plantas eriquecidas con CO2. La resorción del N
fue estudiada en dos especies de arboles de maple (Acer rubrum L y A. saccharum Marsh). Estos
encontraron que los efectos de elevadas concentraciones de CO2 sobre el N del litter son
intrínsecamente más difíciles de detectar que las diferencias en hojas verdes porque los factores
que afectan la senescencia y resorción son muy variables. Los resultados de este experimento
soportan la aproximación usada en ecosistemas modelo en los cuales la efectividad de resorción es
constante a ambiente y elevadas concentraciones de CO2, pero los resultados tambien indican
que otros factores pueden alterar la eficiencia de resorción. La descomposición del mantillo en
bosques Mediterraneos disminuye bajo elevadas concentraciones de CO2, pero si este efecto es
combinado a un aumento de producción primaria , aumentará el almacenamiento de C en suelos
de ecosistemas forestales. Entonces los suelos forestales, por consiguiente, representan un
sumidero potencial para los excesos de carbono (Angelis et al., 2000).
Torbert et al., (2000) utilizaron cultivos de algodón, trigo, sorgo y soya para analizar los
impactos de elevadas concentraciones de CO2 sobre la descomposición de los residuos de estos
cultivos dentro de agroecosistemas, Las observaciones de campo y laboratorio de este estudio
indican que con elevadas atmosferas de CO2, el indice de descomposición de los residuos de la
planta puede ser limitado por el N y la liberación de N de la descomposición de los residuos de la
planta pueden ser bajos.
ALTERNATIVAS PARA MITIFAR CO2 EN LA ATMOSFERA
CAPTURA DE CARBONO
La fotosíntesis remueve alrededor de 105 Pg C (1 Pg=1*109) de la atmósfera cada año,
cerca de la mitad es tomado por la plantas, releases with just over order removes. Las plantas de
la biosfera pueden tener un enorme impacto sobre los niveles de CO2 atmosféricos Revisar
artículo .
Actualmente, la producción de cultivos produce una modesta cantidad de CO2 a la atmósfera
(AAFC, 1999). Con un manejo correcto y un cambio de cultivo que incremente la cantidad de
materia orgánica y reduzca la los indices de descomposición del material orgánico del suelo, los
suelos agrícolas pueden llegar a ser el mejor sumidero de CO2 sink crop change Currently.
Captura de Carbono en tierras áridas
Las tierras áridas se definen por en índice de aridez que representa la realación de la
precipitación con la evapotranspiración potencial (P/PET) con valores <0.05 para tierras
hiperáridas, <0.20 para tierras áridas y de 0.20 a 0.50 para tierras semiáridas. Estas son las tierras
secas más características, pero a menudo la zona árida subhúmeda (0.5-0.65) tambien se incluye
en la misma (Middleton y Thomas, 1997) citado por FAO ( 2001). Las tierras áridas representan
cerca del 40% de las tierras del globo. Las zonas hiperáridas naturales cubren un área estimada en
1000 millones de hectáreas mientras que las tierras áridas, semiáridas y áridas subhumedas cubren
un área de 5100 millones de hectáreas.
Si bien el contenido de carbono y la capacidad de fijar CO2 por unidad de superficie en las
tierras áridas son bajos, pueden de cualquier manera hacer una contribución importante a la
captura global de carbono y al mismo tiempo prevenir o diminuir la tasa de desertificación. Con
esta amplia definición, una gran parte de las tierras áridas se incluyen en el área tropical definida
como la parte intertropical del mundo, la que representa el 37.2 por ciento de la superficie
terrestre (4900 millones de hectáreas)
En un estudio realizado en el desierto de Negev en Yatir, USA con el pino Aleppo (Pinus
halepensis Mill) durante 35 años encontraron que el aumento de CO2 en la atmósfera puede
conducir a un aumento en la disponibilidad del agua, debido a un uso eficiente del agua por las
plantas al incrementar el CO2. Esto permite extender las actividades forestales en regiones aridas.
Estas 2800 ha de tierras forestales aridas contiene 6.5+ 1.2 kg Cm-2 y continúa acumulandose en
0.13-0.24 kg Cm-2 año-1. El CO2 es alto durante el invierno, Los esfuerzos de reforestación
dentro de regiones áridas puede ser significativo para la captura de C y los beneficios
(restauración de la tierra degradada, reducción de ecorrentía, erosión y compactación del suelo,
beneficio a la vida salvaje) debido a la gran extensión de las regiones potencialmente
involucradas ( aproximadamente 2*109 de las tierras con arbustos y paztizales C4). Información
cuantitativa de las actividades forestales bajo condiciones áridas es relevante para regiones que
preveen un aumento en la aridez (Grunzweig et al., 2003 ).
Captura de Carbono en áreas tropicales
Captura de carbono en tierras de pastoreo
Las tierras de pastoreo juegan un papel muy importante en la captura de carbono. En
primer lugar, las tierras de pastoreo, según FAO, ocupan 3200 millones de hectáreas y almacenan
entre 200 y 420 Pg en el ecosistema total, una gran parte del mismo debajo de la superficie y, por
lo tanto, en un estado relativamente estable. El carbono del suelo en las tierras de pastoreo es
estimado en 70 t/ha, cifra similar a las cantidades alamacenadas en los suelos forestales
(Trumbmore et al., 1995; Balesdent y Arrouays, 1999) citado por (FAO, 2001). Muchas áreas de
tierras de pastoreo en la zonas tropicales y àridas son mal manejadas y están degradadas; por lo
tanto ofrecen variadas posibilidades de suecuestro de carbono.
Captura de carbono en ecosistemas forestales
Los bosques cubren el 29% de las tierras y contienen el 60 % del carbono de la vegetación
terrestre. El carbono almacenado en los suelos forestales representa el 36% del total del carbono
del suelo a un metro de profundidad (1500 Pg).
Los ecosistemas forestale contienen más carbono por unidad de superficie que cualquier
otro tipo de uso de la tierra y sus suelos, que contienen cerca del 40% del total del carbono son de
importancia primaria cuando se considera el manejo de los bosques.
Por lo general, en los bosques naturales el carbono del suelo está en equilibrio, pero tan
pronto como ocurre la deforestación o la reforestación, ese equilibrio es afectado. En los lugares
donde no se puede ser detenida la deforestación, es necesario un manejo correcto para minimizar
las pérdidas por carbono. La reforestación, sobre todo en los suelos degradados con bajo
contenido de materia orgánica, será una forma importante de secuestro de carbono a largo plazo,
tanto en la biomasa como en el suelo. Comparado con los demás ecosistemas, los suelos turbosos
son un pequeño sumidero de dioxido de carbono (Turunen et al., 2002).
Captura de carbono en tierras cultivadas
En los sistemas agrícolas, una gran parte de carbono es almacenado en el suelo. La
entrada de carbono en los suelos es determinada por la producción primaria neta y la fracción
que permanece sobre el campo. La pérdida de carbono está determinada por la descomposición y
por la erosión de la capa arable. La descomposición es controlada por la temperatura ambiente y
las propiedades fisicas y químicas del suelo. En general, los rendimientos de los cultivos son
bajos, los contenidos de carbono en el suelo son altos y los indices de descomposición de la
materia orgánica en el suelo son altos lo cual incrementa la pérdida de carbono (Freibauer, 2004).
outside Enhance focus overall sink baseline
El desarrollo de la agricultura ha implicado una gran pérdida de materia orgánica del
suelo. Hay varias formas de las diferentes prácticas de manejo de tierras que pueden ser usadas
para aumentar el contenido de materia orgánica del suelo, tales como el incremento de la
productividad y de la biomasa –variedades, fertilización e irrigación. El cambio climático global
puede tener un efecto similar. Las principales formas de obtener un incremento de la materia
orgánica en el suelo están asociadas a la agricultura de conservación y conforman la labranza
mínima o cero y el uso de cobertura vegetal continúa y protectora formada por materiales
vegetales vivos o muertos sobre la superficie del suelo. Baseline whether depends sink lead
PRINCIPALES CONSECUENCIAS E IMPACTO DE LA CAPTURA DE
CARBONO
La captura de carbono y el aumento de la materia orgánica del suelo tendrán un impacto directo
sobre la calidad y la fertilidad de los suelos. Habrá también efectos positivos importantes sobre el
ambiente y la resiliencia y la sostenibilidad de la agricultura.
Calidad y fertilidad del suelo
La materia orgánica del suelo tiene funciones esenciales desde el punto de vista biológico, físico y
químico del suelo. El contenido de materia orgánica es generalmente considerado como uno de
los indicadores primarios de la calidad del suelo, tanto en sus funciones agrícolas como
ambientales.
La materia orgánica es de especial interés en el caso de los suelos tropicales -excepto en los
vertisoles- con arcillas de baja actividad que tienen una pobre capacidad de intercambio de
cationes. La capacidad de intercambio de cationes aumenta en función del incremento de la
materia orgánica . La biodisponibilidad de otros elementos importantes tales como el fósforo
podrá mejorar y la toxicidad de otros elementos podrá ser inhibida por la formación de quelatos u
otras uniones, por ejemplo, aluminio y materia orgánica (Robert, 1996a).
En una agricultura con bajo uso de insumos, el reciclaje de los nutrimentos -N, P, K, Ca- por
medio de la descomposición gradual de las plantas y los residuos de los cultivos es de importancia
fundamental para la sostenibilidad (Sánchez y Salinas, 1982; Poss, 1991).
En el caso de la erosión, se ha establecido una correlación entre la disminución histórica de la
materia orgánica del suelo y el desarrollo de la erosión. Todos los tipos de manejo de los cultivos
que capturan carbono favorecen la cobertura del suelo y limitan la labranza y de este modo
previenen la erosión.
Impactos ambientales
La captura de carbono en los suelos agrícolas se contrapone al proceso de desertificación por
medio del papel que juega el incremento de la materia orgánica sobre la estabilidad de la
estructura -resistencia a la erosión hídrica y eólica- y a la retención de agua, y al aspecto esencial
de la cobertura de la superficie del suelo directamente por las plantas o por los residuos de las
plantas -o cobertura muerta- para prevenir la erosión e incrementar la conservación del agua
(FAO, 2001).
La materia orgánica, al incrementar la calidad del suelo, también tiene una función protectiva al
fijar los contaminantes -ya sean orgánicos como los pesticidas o minerales como los metales
pesados o el aluminio- los cuales, en general, disminuyen en su toxicidad (FAO, 2001).
La calidad del aire está principalmente relacionada con la disminución de la concentración del
CO2 atmosférico, pero considerando también los otros gases de invernadero, en particular metano
y óxido nitroso (CH4 y N2O). El principal factor que controla su génesis es la anaerobiosis proceso de reducción del suelo- la cual está generalmente ligada a las condiciones hidromórficas.
Cuando aumentan las pasturas o las tierras para pastoreo, la emisión de metano por el ganado
debe también ser tomada en consideración (FAO, 2001).
En algunos ambientes y dependiendo de las condiciones climáticas -áreas húmedas- o propiedades
del suelo -alto contenido de arcilla- puede ser formado N2O. Por lo tanto, se debe hacer un
cuidadoso balance de los distintas emisiones de gases.
El cultivo del arroz en tierras húmedas es el sistema de cultivos más complejo en relación a la
captura de carbono. Si la materia orgánica se acumula en un suelo húmedo, también se forma
CH4. El efecto de invernadero del metano es mucho mayor que el del CO2. La estrategia más
común para prevenir la formación de metano es disminuir el período de inundación, de modo que
la materia orgánica esté menos protegida de la mineralización y puedan ser emitidos CO2 y N2O o
NH4. Por estas razones, parecería muy difícil, por el momento, manejar simultáneamente la
producción de arroz en tierras húmedas y la captura de carbono.
Los últimos hallazgos en la agricultura de conservación respecto a los sistemas arroz-trigo son
positivos; por ejemplo, los rendimientos del arroz pueden ser mantenidos o mejorados sin
saturación de agua, encharcamiento o reducción del suelo y con grandes ahorros de agua en el
período de crecimiento del arroz. Este nuevo enfoque ha sido convalidado por los agricultores en
varios miles de hectáreas en países como India y Brasil.
Las tierras húmedas tienen condiciones anaeróbicas similares con menor emisión de CH4 que los
campos de arroz húmedos y un mayor potencial de secuestro de carbono que puede llevar a la
formación de turba. Esto tiene también otras ventajas ambientales importantes que deben ser
protegidas; no es realista, sin embargo, esperar rápidos incrementos.
La calidad del agua también es mejorada por una disminución de la escorrentía, de los
contaminantes y de la erosión. En el caso específico de la labranza de conservación, también se
evita o minimiza una fuerte mineralización de la materia orgánica con la subsecuente formación
de nitratos.
Los cambios en el uso de la tierra y en su manejo también tienen un efecto importante sobre la
distribución de la precipitación pluvial entre escorrentía y almacenamiento o infiltración, con un
aumento de la última en el caso de las tierras de pastoreo, bosques y labranza de conservación con
cobertura de suelo. La cobertura del suelo previene la erosión; por lo tanto, si hubiera alguna
escorrentía, el agua estaría libre de partículas asociadas con contaminantes -elementos traza, PO4.
La contaminación a distancia por productos solubles también disminuirá en relación directa con la
menor escorrentía. Esta es una de las bases de la ecocondicionalidad en la US Farm Bill desde
1996. Con tales cambios en las prácticas agrícolas puede ser enfrentado el desafío de la calidad
del agua. Una vez que los cambios hayan tenido lugar en grandes áreas, también decrecerá la
severidad y frecuencia de las inundaciones.
El efecto general del incremento de la materia orgánica del suelo es un mejoramiento de la
capacidad amortiguadora y de la resiliencia del suelo a diferentes tipos de degradación o estrés.
Biodiversidad y función biológica del suelo
Los cambios en la biodiversidad son evidentes cuando ocurre la deforestación. En el caso de la
reforestación, dependerán del tipo de bosque establecido. Los sistemas agro-forestales bien
manejados involucran una amplia gama de biodiversidad. Por lo general, la biodiversidad de los
mamíferos es preservada en el caso de los bosques, el número de especies de aves se reduce a la
mitad y las especies vegetales disminuyen en un tercio (de 420 a 300), (IPCC, 2000). Likey,
(ICRAF) se refiere a un mosaico de manchas, cada una de ellas compuesta de muchos nichos, o
sea un sistema favorable para la biodiversidad.
En el pasado, los sistemas agrícolas más intensivos llevaron a una sensible disminución de la
biodiversidad, junto a una paralela reducción de la materia orgánica debida sobre todo a la
labranza y al uso de pesticidas (Rovira, 1994).
En el caso de las tierras de cultivo, el aumento de la biodiversidad en relación con el incremento
de la materia orgánica se basa, sobre todo, en el aumento de la biodiversidad del suelo (Copley,
2000). La Figura 13 presenta una organización jerárquica de la biodiversidad del suelo, la cual
depende directamente del abastecimiento de materia orgánica fresca y de las prácticas
agronómicas. Esta biodiversidad varía desde los genes hasta los microorganismos, la fauna y la
biodiversidad encima de la tierra. La cantidad de bacterias puede aumentar en forma exponencial,
de 103 a 1012, tan pronto como la materia orgánica sea abundante. La labranza cero favorecerá el
desarrollo de hongos los cuales son sumamente activos en la agregación del suelo. Sin embargo,
solo 5 a 10 por ciento de las especies de la microflora del suelo son conocidas y en la actualidad
sería posible investigar, gracias a las nuevas técnicas moleculares, la evaluación de la
biodiversidad específica o interespecífica de los microorganismos.
Figura 13. Organización jerárquica de la biodiversidad del suelo
Cuando la materia orgánica fresca -residuos de las plantas o plantas de cobertura- está presente en
la superficie del suelo, habrá un incremento de las distintas categorías de la fauna, sobre todo de
los descomponedores. Las cadenas alimenticias asociadas a los detritos serán estimuladas
(Hendricks et al., 1986) -bacterias, hongos, microartró-podos, nematodos, enquitreidosmacroartrópodos. Las lombrices de tierra, las termites y las hormigas, que son los principales
grupos que componen la macrofauna (>1 cm) a menudo son llamados ingenieros del suelo en
razón de la función que tienen sobre la porosidad -bioporos- y estructura del suelo; su número se
incrementa paralelamente al aumento de la materia orgánica con una disminución del disturbio del
suelo, o sea la no labranza (Figura 14). Son buenos indicadores de la calidad del suelo (Lavelle,
2000; Lobry de Bruyn, 1997) y tienen un papel fundamental en la agricultura de conservación.
Son, por ejemplo, indispensables para asegurar la distribución a través del suelo -incluso a más de
un metro de profundidad- de la materia orgánica acumulada en la superficie.
Figura 14. Efecto del sistema anterior de labranza sobre el número de lombrices de tierra en
varias fincas. Canterbury, Nueva Zelandia (de Fraser, en Soil biota, 1994)
Un aumento en la captura de carbono causa un incremento en la biodiversidad activa y un
funcionamiento más efectivo de los elementos biológicos del suelo, lo cual es un proceso
relativamente lento en la mayoría de los suelos agrícolas. La biodiversidad de todo el agro-sistema
(vegetación, aves, etc.) también depende del tipo de manejo.
Todas las consecuencias y los beneficios de este enfoque también deberían ser apreciados en
relación con la sostenibilidad de la agricultura, incluso con respecto a los depósitos de genes y el
control biológico de las plagas.
Efectos del cambio climático
Mientras que un aumento del contenido atmosférico de gases de invernadero está llevando a un
cambio climático, también ocurrirán numerosos efectos complejos, contrastantes y opuestos
(Brinkman y Sombroek, 1996).
Todos los resultados experimentales demuestran que un aumento de la concentración de CO2 en la
atmósfera induce un incremento de la biomasa o de la Red Primaria de Producción por medio de
la fertilización con carbono, con un papel muy importante sobre la fotosíntesis y el crecimiento de
las plantas. La ganancia en la fijación de CO2 podría ser importante. El incremento en la
productividad medido a causa de la duplicación de la concentración del CO2 -predicha para el año
2100- es de cerca del 30 por ciento para las plantas C3. Otro efecto importante del aumento del
CO2 es la disminución de la transpiración de las plantas a través de los estomas lo cual redunda en
una mayor eficiencia en el uso del agua (WUE), sobre todo en las plantas C4. En lo que se refiere
al agua, hay un efecto neto favorable del CO2 sobre la reducción de la transpiración de las plantas
(Gregory et al., 1998). Evidentemente, para llegar a un aumento de rendimiento en el campo,
también deben ser satisfechos otros requerimientos de las plantas como el agua y los nutrientes
disponibles.
En lo que se refiere al ciclo del carbono, habrá una mayor captura de carbono por la biomasa
aérea y un correlativo ingreso de carbono en el suelo a partir de los residuos de las plantas y del
crecimiento y la muerte de las raíces más finas. Los compuestos de las raíces tienen una mayor
relación C/N y son más estables.
Otro factor que juega un papel importante en la captura de carbono es la temperatura, la que
podría aumentar en algunas partes del globo terráqueo. Tal incremento podría provocar una mayor
tasa de mineralización de la materia orgánica por los microorganismos y una mayor tasa de
respiración de las raíces. Este efecto de la temperatura sobre la mineralización podría ser
significativo en los países fríos, donde la temperatura es un factor limitante y donde puede ser
esperado un incremento de las emisiones de CO2. Sin embargo, en la mayor parte del mundo es de
esperar un aumento de la captura de carbono (van Ginkel et al., 1999).
Para estimar el efecto del cambio climático sobre la captura de carbono pueden ser usados
modelos. Los resultados de muchos estudios recientes confirman el incremento de la tasa de
crecimiento de los bosques en las zonas templadas y en los países nórdicos. En lo que se refiere a
los bosques tropicales, existen algunas medidas hechas en la Amazonía donde se ha encontrado un
aumento de la biomasa (Phillips et al., 1998) de 0,62 t C/ha/año, lo cual para un área de 7 000
millones de hectáreas significa una captura de carbono de Gt 0,44 C/año. La causa de esto no es
simple, ya que la influencia de El Niño está probablemente involucrada en el aumento de la
humedad del área.
USO DE COMBUSTIBLES O ENERGIA ALTERNATIVA
CONCLUSIONES
BIBLIOGRAFIA
Angelis, P., Chigwerewe, K.S. and Scarascia Mugnozza, G.E. 2000. Litter quality and
decomposition in a CO2-enriched Mediterranean forest ecosystem. Plant and Soil 224: 31-41
Luo, Y. and Mooney, H.A. 1999. Carbon Dioxide and Enviromental Stress. Ed. Academic Press,
San Diego California, USA. . 418 pp.
Norby, R.J., Long, T.M., Hartz-Rubin, J.S and O’Neill, E.G. 2000. Nitrogen resorption in
senescing tree leaves in a warmer, CO2-enriched atmosphere. Plant and Soil 224: 15-29.
Torbert, H.A., Prior, S.A., Rogers, H.H. and Wood, C.W. 2000. Review of elevated atmospheric
CO2 effects on agro-ecosystems:residue decomposition processes and soil C storage. Plant and
Soil 224: 59-73.
DIAGRAMA DEL PROCESO DE PRODUCCION DE SETAS Y RECICLAJE DE LA
PAJA DE CEBADA, PROVENIENTE DE LA PRODUCCION DEL HONGO SETA
(Pleurotos ostreatus)
Remojado de pacas de cebada
Pasteurización (45 min.)
Escurrimiento y encalado (40 min)
Siembra
Incubación (20-40 días)
Cosecha (60- 90 días)
Residuo de paja de cebada de tres oleadas
Residuo de paja de cebada de cinco oleadas
oleadas
Residuo de paja de cebada contaminada
Mezcla de pajas
Elaboración de pilas y llenado de las pilas
Inoculación de lombriz de tierra
Control de temperatura, aireación y humedad de la
vermicomposta
Caracterización del compost
ANALISIS DE GRUPOS MICROBIANOS EN EL PROCESO DE PRODUCCION DE
COMPOSTA Y VERMICOMPOSTA
Erica Morales Hernández1
1
Alumna de la Especialidad de Edafología, IRENAT-Colegio de Postgraduados, Montecillos,
Estado de México
La cinética de los grupos microbianos es muy importante en la producción de compost y
vermicompost, ya que éstos junto con los factores climáticos y la humedad pueden disminuir el
tiempo para degradar la materia orgánica a compuestos más estables y asimilables por las plantas.
Por otra parte la incorporación de lombrices al proceso de composteo ayuda y favorece en gran
medida a la transformación de los residuos orgánicos sin valor comercial a productos con valor
comercial. Es recomendable realizar un análisis fisicoquímico y microbiológico cuando un
residuo ha sido previamente sometido a algún tratamiento, ya sea físico o químico, antes de
empezar con el proceso de composteo y vermicomposteo y aun más cuando los residuos han
recibido un precomposteo, como es el caso del sustrato que es utilizado para el crecimiento del
hongo Pleurotos ostreatus y residuos hortofrutícolas , ya que en el primero, la paja de avena ha
recibido una pasteurización, el hongo ha consumido lignina y celulosa principalmente, y además
se quedan residuos de micelio y cuerpos fructíferos que tienen un alto contenido de proteínas, lo
que da lugar a que el residuo sea más rápidamente degradado, mientras que en el segundo, los
microorganismos ya han degrado moléculas como almidón y ha aumentado la población de
microorganismos, lo que favorece el proceso de vermicomposteo y composteo. Además es
recomendable realizar análisis fisicoquímicos y microbiológicos del compost y vermicompost;
dentro del químico recomendaría iones solubles (calcio, magnesio, sodio, potasio, cloruros,
carbonatos, bicarbonatos, sulfatos, nitratos, amonio, fosfatos, boro, molibdeno), ya que estos son
fácilmente disponibles para las plantas, y con estos datos podría hacer una balance nutrimental de
acuerdo al cultivo que se desee manejar, ya que los análisis que le hicieron de Nitrógeno y
Fósforo total al compost y vermicompost en este trabajo, no se sabe si están disponible o como
macromoléculas todavía no asimilables por las plantas, y la conductividad eléctrica solo es un
indicador de salinidad, pero no nutrimental, y lo que realmente nos interesa cuando se realiza una
composta es su calidad nutrimental, ya que de eso depende si se utiliza o no con fines agrícolas.
En cuanto a los análisis físicos recomendaría humedad, curva de tensión de humedad, porosidad,
infiltración, ya que esto es necesario cuando se usa como sustrato para la producción de algún
cultivo en particular.
BIBLIOGRAFIA
Corlay, C., L., Ferrera, C., R., Etcheverts, B., J., Echegaray, A., Alfredo y Santizo, R., J. 1999.
Cinetica de grupos microbianos en el proceso de producción de composta y vermicomposta.
Agrociencia 33, 375-380..
New
Volume
147 Issue
doi:10.1046/j.1469-8137.2000.00687.x
1 Page
201
Elevated atmospheric CO2, fine roots
microorganisms: a review and hypothesis
-
and
the
July
response
Phytologist
2000
of
DONALD R. ZAK, KURT S. PREGITZER, JOHN S. KING & WILLIAM E. HOLMES
There is considerable uncertainty about how rates of soil carbon © and nitrogen (N) cycling will
change as CO2 accumulates in the Earth’s atmosphere. We summarized data from 47 published
reports on soil C and N cycling under elevated CO2 in an attempt to generalize whether rates will
increase, decrease, or not change. Our synthesis centres on changes in soil respiration, microbial
respiration, microbial biomass, gross N mineralization, microbial immobilization and net N
mineralization, because these pools and processes represent important control points for the belowground flow of C and N. To determine whether differences in C allocation between plant life forms
influence soil C and N cycling in a predictable manner, we summarized responses beneath graminoid,
herbaceous and woody plants grown under ambient and elevated atmospheric CO 2. The belowground pools and processes that we summarized are characterized by a high degree of variability
(coefficient of variation 80 800%), making generalizations within and between plant life forms difficult.
With few exceptions, rates of soil and microbial respiration were more rapid under elevated CO 2,
indicating that (1) greater plant growth under elevated CO 2 enhanced the amount of C entering the
soil, and (2) additional substrate was being metabolized by soil microorganisms. However, microbial
biomass, gross N mineralization, microbial immobilization and net N mineralization are characterized
by large increases and declines under elevated CO2, contributing to a high degree of variability within
and between plant life forms. From this analysis we conclude that there are insufficient data to predict
how microbial activity and rates of soil C and N cycling will change as the atmospheric CO 2
concentration continues to rise. We argue that current gaps in our understanding of fine-root biology
limit our ability to predict the response of soil microorganisms to rising atmospheric CO 2, and that
understanding differences in fine-root longevity and biochemistry between plant species are
necessary for developing a predictive model of soil C and N cycling under elevated CO 2.
soil
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Kurt S. Pregitzer. (2002) Fine roots of trees a new perspective. New Phytologist 154:2, 267-270
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G. A. Bauer, G. M. Berntson and F. A. Bazzaz. (2001) Regenerating temperate forests under elevated CO2 and
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Letters 4:4,
344-347
Evidence that decomposition rates of organic carbon in mineral soil do not vary with
temperature
CHRISTIAN P. GIARDINA* AND MICHAEL G. RYAN†
Nature 404, 858 - 861 (20 April 2000); doi:10.1038/35009076

Department of Natural Resources and Environmental Management, University of Hawaii at
Manoa,
1910
East-West
Road,
Honolulu,
Hawaii
96822
,
USA
† United States Department of Agriculture-Forest Service, Rocky Mountain Research Station,
240 West Prospect Street, Fort Collins, Colorado 80526, USA, and
Graduate Degree Program in Ecology, Colorado State University, Fort Collins, Colorado
80523, USA
Correspondence should be addressed to C.P.G. (e-mail: giardina@hawaii.edu).
It has been suggested that increases in temperature can accelerate the decomposition of
organic carbon contained in forest mineral soil (Cs ), and, therefore, that global warming
should increase the release of soil organic carbon to the atmosphere1-6. These predictions
assume, however, that decay constants can be accurately derived from short-term
laboratory incubations of soil or that in situ incubations of fresh litter accurately represent
the temperature sensitivity of C s decomposition. But our limited understanding of the
biophysical factors that control Cs decomposition rates, and observations of only minor
increases in Cs decomposition rate with temperature in longer-term forest soil heating
experiments7-12 and in latitudinal comparisons of Cs decomposition rates13-15 bring these
predictions into question. Here we have compiled Cs decomposition data from 82 sites on
five continents. We found that Cs decomposition rates were remarkably constant across a
global-scale gradient in mean annual temperature. These data suggest that Cs decomposition
rates for forest soils are not controlled by temperature limitations to microbial activity,
and that increased temperature alone will not stimulate the decomposition of forest-derived
carbon in mineral soil.
To examine the long-term influence of temperature on the decomposition of Cs in forest soils, we
assembled results from studies that used one of two standard methods for estimating Cs loss from
soil. Method 1 studies estimate Cs loss by measuring in situ changes in the 13C/12C ratio and total
Cs content of soil after existing vegetation is replaced with vegetation that uses a different
photosynthetic pathway (for example, C3 forest to C4 pasture)16. The change in vegetation alters
the 13C/12C ratio of new detritus, allowing an estimate of the loss of Cs formed before conversion.
Method 2 studies estimate Cs loss by incubating soils in the laboratory for 1 yr at temperatures
representative of field conditions, and quantifying the CO2 evolved17, 18. The 82 sites examined
here span 8 soil orders and the global range of mean annual temperature (MAT) for forests (see
Supplementary Information).
Calculated Cs turnover times for method 1 studies were unrelated to MAT (R2 = 0.01, P = 0.50;
Fig. 1); turnover times under moist tropical conditions were similar to those in cool temperate
soils. In these studies, soil clay content, which is thought to control Cs storage1, did not explain
the lack of a relationship between Cs decomposition rates and MAT. For soils with similar (15–
27%) clay content, Cs turnover was still unrelated to MAT (R2 = 0.05, P = 0.28). In method 2
studies, Cs turnover time was positively related to incubation temperature, with Cs decomposing
more slowly at higher temperatures (R2 = 0.14, P = 0.02; Fig. 2). The Cs lost from method 1 and
method 2 soils is the most active, and therefore the most temperature-sensitive, carbon in mineral
soil2, 9, 19. However, the decomposition rates of forest-derived Cs reviewed here are insensitive to
temperature, unlike the response predicted by models1-6.
Figure 1 Relationship between turnover time for mineral soil carbon and
mean annual temperature for method 1 studies.
Full legend
High resolution image and legend (11k)
Figure 2 Relationship between observed or modelled Cs turnover time
and incubation temperature for incubated forest soils. Full legend
High resolution image and legend (14k)
In our calculations of turnover time, we assumed a single-pool model for soil carbon. To test
whether this assumption affected the results, we compared Cs mass loss per year across MAT,
with method 1 sites segregated by time since conversion. For method 1 studies, C s mass loss per
year decreased with increasing MAT for sites sampled <11 yr after conversion (R2 = 0.66; P <
0.01), and was unrelated to MAT for sites sampled 11–45 yr after conversion (R2 = 0.01; P =
0.72) or >45 yr after conversion (R2 = 0.15; P = 0.35). Despite a 20 °C gradient in MAT, Cs mass
loss as a function of time since conversion was insensitive to temperature (Fig. 3). Cs mass loss
was roughly constant with time until about 30 years or about 60% mass loss. After this time, Cs
decomposition appears to slow dramatically or stop. For method 2 studies, Cs mass loss for the 1yr incubations decreased with incubation temperature (R2 = 0.19, P < 0.01). We conclude that our
choice of a single-pool model did not cause the lack of a relationship between Cs decomposition
rates and temperature.
Figure 3 Relationship between Cs mass loss and time (in years) since
conversion
of
vegetation
type.
Full legend
High resolution image and legend (14k)
Our results conflict with those of Trumbore et al.2, who used 14C-based estimates of light-fraction
Cs age (separated from generally smaller quantities of heavy-fraction C s during density
fractionation of total Cs) to model the decomposition of light-fraction Cs across a gradient in
MAT. These authors found that decomposition rates of light-fraction Cs increased exponentially
with increasing temperature. An explanation for the discrepancy with our findings may be that in
the study of Trumbore et al., other factors that alter Cs decomposition (moisture, disturbance, and
litter quality20, 21) were highest at sites with the highest decomposition rates. The warmer sites
were generally wetter, more disturbed, and supported vegetation that produced higher-quality
litter.
Although many short-term studies of Cs or litter decomposition show that decomposition rates
increase with temperature3, 6, 10, transient responses to increasing global temperature are unlikely
to represent the response of most detrital carbon in forests. First, detrital carbon in forests resides
primarily in the mineral soil (up to 70% in boreal forests13 and 95% in the lowland tropics22, 23),
and in situ Cs mass loss rates are much slower than losses of fresh litter or forest-floor material13,
23
. Second, all method 2 studies show large, rapid declines in decomposition rates in the first
weeks of incubation, during which <5% of total Cs is typically released9-12, 18, 21. These declines
indicate either the depletion of a very small, active pool of Cs, or—because soils are processed
before incubation—a return to pre-disturbance conditions. Third, long-term incubations of forest
soil9-12, and in situ comparisons of Cs content in heated and unheated soil8 or Cs turnover along
gradients in MAT13-15, show responses to increased temperature that are small, ephemeral or nonexistent (that is, Q10 values of 1.0–1.4, where Q 10 = reaction rate at T + 10 °C/reaction rate at T,
and T is temperature). Taken together, these data and the data presented in Figs 1 and 2 suggest
that sustained, temperature-related increases in the decomposition rate of forest-derived Cs should
not be expected.
A global-scale relationship between Cs decomposition rates and MAT is central to predictions that
global warming will accelerate the release of carbon stored in mineral soil1-6. However,
decomposition is performed by enzymes, and enzyme activity is limited by temperature only
when the supply rate of substrate exceeds the reaction rate for that substrate. Therefore, the most
tenable explanation for the apparent temperature insensitivity of Cs decomposition is that
heterotrophic microbes in mineral soil (those organisms responsible for decomposing C s) survive
on a supply of substrate that is sub-optimal for growth7, 21, 24. Soil clay content, available moisture,
and Cs quality are three factors that may influence substrate availability1, 21.
The binding of Cs to clay particles and physical protection within soil aggregates are thought to
lower Cs availability1, 21. At a given location, where variations in climate and biota are more
uniform, turnover times are longer for Cs associated with clay than with sand20, 25. If clay controls
substrate availability, we would expect Cs turnover times to increase as clay content increases.
However, data from method 1 and method 2 studies provide only weak support for clay
limitations to substrate availability. For both method 1 and method 2 studies, Cs turnover time was
nearly constant across sites, and variation among soils within a region was similar to global
variation. For method 1 studies, where clay varied from 7% to 84%, there was no relationship
between Cs turnover time and soil clay content (R 2 = 0.05, P = 0.14; see Supplementary
Information ). In method 2 studies, Cs mass loss was weakly related to clay content in soils with
7–30% clay (R2 = 0.15, P = 0.05; Supplementary Information). For clay >30%, Cs mass loss
decreased with increased clay (R2 = 0.41, P = 0.06; Supplementary Information). While these
results conflict with established modelling assumptions1, other studies have also found weak
relationships between clay content and Cs decomposition rates17, 26.
Available moisture exerts a large influence on soil microbial activity21, and low soil moisture
probably reduces microbial populations. However, available moisture did not affect substrate
availability in method 2 studies because soils were maintained at or near field moisture capacity
for the length of the incubation. Although we have no information on differences in soil moisture
among method 1 sites, method 1 data were taken primarily from sites that had previously
supported closed-canopy forests, which indicates moisture regimes that are at least adequate for
decomposition.
Low Cs quality may limit substrate availability for microbes, and perhaps also limit microbial
populations21. Forest-derived Cs consists of lignin-dominated remains and precipitated byproducts of plant and microbial residue decomposition. These compounds are poor carbon sources
for microbes, because energy yields are low for the energy expended to digest them 21. Evidence
that low quality of Cs limits Cs decomposition rates in mineral soil includes low C s decomposition
rates compared with rates for fresh litter23 and rapid increases in CO2 release from nonrhizosphere soils that are amended with labile substrate24. 13C nuclear magnetic resonance
analyses of forest-derived Cs show that the relative abundance of Cs functional groups (for
example, alkyls, O-alkyls, aromatics and carbonyls) varies minimally across global-scale
gradients in MAT (ref. 27). If Cs quality limits decomposition rates, then low global-scale
variation in Cs decomposition rates may reflect low variation in the chemical composition of Cs.
Whether Cs decomposition is controlled by temperature or by substrate availability will change
predictions for the effect of global warming on the large quantity of Cs stored in tropical soils1, 2, 28
and in soils that are frozen for most of the year28, 29. If temperature limits Cs decomposition, as
assumed in most current ecosystem models, then tropical soils would provide the main source of
additional carbon released in a warmer climate because Cs decomposition rates in high-latitude
soils would be constrained by perennially low soil temperatures1, 2. In contrast, if substrate
availability limits Cs decomposition rates, increased global temperatures alone would have little
influence on Cs decomposition rates in the tropics. Warming at high latitudes, however, would
expose larger amounts of Cs to microbial activity by lowering the depth of frozen soil, lowering
the water table, and extending the duration of thawed conditions29. Once these soils are thawed or
the water table lowered, Cs decomposition rates would be constrained by substrate quality and
moisture rather than by low temperatures.
The turnover of Cs in forest soils appears to be remarkably constant on a global scale, and
insensitive to differences in MAT. However, the relationship between Cs turnover and MAT
presented here serves only as a proxy for the changes that will occur in situ in response to global
warming. The influence of temperature on Cs decomposition rates needs to be directly examined
across a range of sites to better constrain predictions of the effects of warming on carbon release
from soil.
Methods
Method 1 studies primarily examined the loss of forest-derived Cs . All sites were disturbed during
conversion, but land management varied during and following conversion. Details on
methodology for a typical method 1 study are given in ref. 16. For method 2 studies, all soils were
sampled from closed-canopy forests, similarly processed, and maintained at constant moisture
levels near field capacity for the length of the incubation period. We used studies in which soils
were incubated for at least 1 yr, because long-term incubations provide information on the
mineralization potential of both the small labile Cs pool and the larger intermediate Cs pool9.
Microbial activity and Cs decomposition rates also may be less sensitive to sampling disturbance
in long-term incubations than in short-term incubations. Method 2 studies have some important
limitations. Soil macro-structure is altered during sampling and processing. Soils are incubated at
a constant temperature and high moisture, whereas field environments fluctuate and are generally
drier. Finally, incubations eliminate carbon inputs, which can shift microbial populations 18.
Although incubation-derived indices of Cs decomposition rates will probably differ from those
developed in the field, relative differences among sites are presumably real17, 18, 21, 26.
We used single-pool exponential decay models to estimate Cs turnover time in method 1 and
method 2 studies. Single-pool models are widely used to describe Cs turnover, but they may overestimate rates because the approach assumes that all Cs will behave as did the Cs released during
the study30. It is unlikely, however, that our single-pool approach masked a relationship between
the turnover of a large, temperature-sensitive Cs pool and temperature. In method 1 studies, we
would suspect masking if Cs mass loss per year increased with temperature early in the
decomposition sequence. However, for sites with <11 yr since conversion, Cs mass loss per year
was higher for cool sites (9.5% yr-1) than for warm (4.5% yr-1 ) climate soils (P < 0.01). In method
2 studies, we would suspect masking if the total quantity of Cs released per gram of soil increased
with temperature because the release of this C s is independent of total Cs pool size. However, Cs
released per kilogram of soil in one year declined with increasing incubation temperature (R2 =
0.11, P = 0.04). Further, for masking to have occurred, the proportion of total Cs that is
temperature sensitive or fast-cycling must decline steeply with increasing temperature. Using the
best techniques available, Trumbore et al.2 found no relationship between the proportion of fastcycling C s (from 50% to 80% of total Cs) and MAT. Nonetheless, we tested the potential for
masking by assuming that all Cs released in method 2 studies was fast cycling and that 50%, 65%
and 80% of total C s in tropical, temperate and subalpine soils, respectively, was fast cycling. We
then recalculated turnover times for fast-cycling Cs alone. Overall, patterns of Cs decomposition
were unchanged.
Supplementary information accompanies this paper.
Received 10 August 1999; accepted 14 March 2000
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different amounts of clay. Soil Biol. Biochem. 13, 313-321 (1981). | Article | ISI |
27. Mahieu,N., Powlson,D. H. & Randall,W. Statistical analysis of published carbon-13 CPMAS spectra of
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Acknowledgements. We thank R. Hubbard, D. Binkley, R. Waring, S. Hart, S. Trumbore, I.
Døckersmith, R. Sanford Jr and M. Bashkin for comments on earlier versions of this manuscript,
and R. King for help with statistics. This work was supported by the US National Science
Foundation and the USDA Forest Service.
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2003;
10.1104/pp.103.030569
Plant Physiology, January 2004, Vol. 134, pp. 520-527
ENVIRONMENTAL STRESS AND ADAPTATION
Respiratory Oxygen Uptake Is Not Decreased by an Instantaneous Elevation of [CO2], But Is
Increased with Long-Term Growth in the Field at Elevated [CO2]1
Phillip A. Davey2, Stephen Hunt, Graham J. Hymus3, Evan H. DeLucia, Bert G. Drake,
David F. Karnosky and Stephen P. Long*
Departments of Crop Sciences and Plant Biology, University of Illinois, Urbana, Illinois 61801
(P.A.D., S.P.L., E.H.D.); Department of Biology, Queen’s University, Kingston, Ontario, K7L
3N6, Canada (S.H.); Smithsonian Environmental Research Center, Edgewater, Maryland 21307
(G.J.H., B.R.D.); and School of Forest Resources and Environmental Science, Michigan
Technological University, Houghton, Michigan 49931 (D.F.K.)
ABSTRACT
Averaged across many previous investigations, doubling the CO2 concentration ([CO2]) has
frequently been reported to cause an instantaneous reduction of leaf dark respiration measured as
CO2 efflux. No known mechanism accounts for this effect, and four recent studies have shown
that the measurement of respiratory CO2 efflux is prone to experimental artifacts that could
account for the reported response. Here, these artifacts are avoided by use of a high-resolution
dual channel oxygen analyzer within an open gas exchange system to measure respiratory O2
uptake in normal air. Leaf O2 uptake was determined in response to instantaneous elevation of
[CO2] in nine contrasting species and to long-term elevation in seven species from four field
experiments. Over six hundred separate measurements of respiration failed to reveal any decrease
in respiratory O2 uptake with an instantaneous increase in [CO2]. Respiration was found
insensitive not only to doubling [CO2], but also to a 5-fold increase and to decrease to zero. Using
a wide range of species and conditions, we confirm earlier reports that inhibition of respiration by
instantaneous elevation of [CO2] is likely an experimental artifact. Instead of the expected
decrease in respiration per unit leaf area in response to long-term growth in the field at elevated
[CO2], there was a significant increase of 11% and 7% on an area and mass basis, respectively,
averaged across all experiments. The findings suggest that leaf dark respiration will increase not
decrease as atmospheric [CO2] rises.
A quantitative analysis of prior studies weighted for replication and experimental variation
concluded that a doubling of atmospheric CO2 concentration would decrease respiratory CO2
efflux by 18% in woody plants (Curtis and Wang, 1998 ). This decrease is considered the result of
a direct instantaneous effect of increased CO2 concentration and a longer term indirect effect due
to changes in leaf composition with long-term growth at elevated CO2 (for review, see Drake et
al., 1999 ). In an analysis of 45 species, 36 showed an average 15% instantaneous reduction in net
respiratory CO2 efflux per unit leaf area (Rd,CO2) on transfer to elevated [CO2] (for review, see
Amthor, 1997 ). Terrestrial plant respiration releases 10 times more carbon per annum than fossil
fuel combustion (Amthor, 1997 ). Therefore, a 15% to 20% decrease in foliar respiration might
increase the potential of terrestrial vegetation to sequester carbon into biomass, providing a partial
amelioration to the rate of increase of atmospheric [CO2] (for review, see Gonzalez-Meler and
Siedow, 1999 ).
However, there is considerable variation in the reported instantaneous effects of elevated [CO2] on
Rd,CO2 with some studies reporting a large decrease and others reporting no change (Bunce, 2002 ;
Bruhn et al., 2002 ). Furthermore, whereas some biochemical reactions within plant respiratory
metabolic pathways are sensitive to [CO2], none of those identified exert sufficient metabolic
control to account for the reported decreases in respiration (Gonzalez-Meler and Siedow, 1999 ).
Four developments now suggest that the decreases observed when measuring Rd,CO2 from leaves
in gas exchange systems incorporating infrared CO2 analyzers may be the result of artifacts in the
measuring system. Amthor (2000 ) showed that very thorough sealing and use of an enlarged
cuvette enclosing the leaf greatly reduced the apparent instantaneous effect of doubling [CO2] on
Rd,CO2, although a small significant effect still remained. Jahnke (2001 ) systematically analyzed
and eliminated potential artifacts within a gas exchange system and subsequently failed to detect
any response in bean (Phaesoleus vulgaris) and poplar (Populus tremula) leaves when corrections
were applied. Bouma et al. (1997 ) and Burton and Pregitzer (2002 ) similarly demonstrated that
respiratory efflux of CO2 from roots was independent of measurement CO2 concentration.
Assuming common dark respiratory metabolism between roots and leaves, this casts further doubt
on the possibility of a direct response in the leaf. Avoiding the potential artifacts of CO2 exchange
measurement by determining dark respiratory O2 uptake (Rd,O2), Amthor et al. (2001 ) showed no
significant change in respiration of five Rumex crispus leaves in sharp contrast to earlier
measurements of Rd,CO2 in the same species (Amthor et al., 1992 ).
Measurement of O2 uptake has three important advantages over measurement of CO2 efflux for an
instantaneous effect of change in [CO2] on respiration. First, the gas being measured is not the gas
being altered in concentration, avoiding any need for instrument recalibration. Second, the
concentration gradient of [O2] between the cuvette enclosing the leaf and the surrounding air is
unaltered when [CO2] is changed. Finally, O2, unlike CO2, is not easily absorbed and adsorbed by
surfaces in the gas exchange system. Yet, even when the effect of elevated [CO2] on O2 uptake has
been examined, findings have been variable. In contrast to Amthor et al. (2001 ), Reuveni et al.
(1993 ) measured a decrease in respiratory O2 uptake under elevated [CO2]. Controversy over the
presence or absence of a direct effect of [CO2] on Rd,CO2 continues. Some recent studies of Rd,CO2,
despite controlling for errors likely to arise from leaks, continue to report a significant
instantaneous decrease in Rd,CO2 on elevation of [CO2]. For example, decreases have been reported
of 19% for rice (Oryza sativa) canopies on elevation of [CO2] from 350 to 700 µmol mol-1 (Baker
et al., 2000 ); 31% to 79% for Scots pine (Pinus sylvestris) needles also on elevation from 370 to
700 µmol mol-1 (Jach and Ceulemans, 2000 ); 16% to 40% for eight species of crop and crop
weeds on increase from 60 to 1,000 µmol mol-1 (Bunce, 2001 ); an 11% average for leaves of 12
grass species on elevation from 360 to 1,300 µmol mol-1 (Tjoelker et al., 2001 ); 18% in the
second leaf of soybean (Glycine max) on elevation from 350 to 1,400 µmol mol-1 (Bunce, 2002 );
and 4.5% decrease for seedling shoots of the tropical rainforest tree Tectona grandis on elevation
from 370 to 600 µmol mol-1 (Holtum and Winter, 2003 ). Precautions taken in measurement were
sufficient for Bunce (2002 ) to rebut the idea that the direct effect of elevated [CO2] on respiration
was an artifact: “These precautions coupled with observed effects of dark [CO2] treatments on
mass accumulation and translocation, suggest that the observed changes in respiration were real.”
Although these decreases are smaller than those suggested by earlier studies, and some use larger
increases in [CO2] to elicit an effect, they continue to suggest direct instantaneous inhibition of
leaf dark respiration by increase in [CO2]. An added uncertainty is whether an instantaneous
suppression of respiration by elevated [CO2] is limited to certain species or conditions (Drake et
al., 1999 ). Although maintaining the same [CO2] outside the cuvette to that inside will minimize
diffusive leaks, it will not eliminate the substantial problem of CO2 absorption/desorption from
surfaces in the gas exchange system (Bloom et al., 1980 ; Long et al., 1996 ; Jahnke, 2001 ) or
the mass flow of CO2 between the cuvette and the outside through the intercellular air space when
only part of the leaf is enclosed (Jahnke and Krewitt, 2002 ). Thus, some of these further artifacts
may still affect even these very careful studies. Clearly, we are some way from any consensus
over the existence, or otherwise, of a direct effect of increase in [CO2] on leaf respiration in the
dark.
If the artifacts suggested by Amthor (2000 ), Jahnke (2001 ), and Jahnke and Krewitt (2002 )
apply to the effect of an instantaneous increase in [CO2] on Rd,CO2, then the same errors would at
least in part apply when these systems are used to measure the effect of long-term acclimation to
elevated [CO2] on Rd,CO2. Plants grown in elevated [CO2] in the long-term (1-10 years) typically
show decreased nitrogen content and increased photosynthesis, growth, leaf mass per unit area,
nonstructural carbohydrate concentrations (Drake et al., 1997 ), and large increases in leaf
mitochondrial numbers (Robertson et al., 1995 ; Griffin et al., 2001 ; Tissue et al., 2002 ). With
the exception of nitrogen content, these are all factors that might be expected to lead to increased
leaf respiration. However, a quantitative survey using meta-analytical analysis concluded that
long-term growth at elevated [CO2] decreased Rd,CO2 significantly and by 18% (Wang and Curtis,
2002 ).
Recently, high-sensitivity dual lead-gold (fuel cell) detectors have been developed that allow
measurement of small differences in [O2] (approximately 1-2 µmol mol-1) between the inlet and
outlet air streams of a leaf cuvette in normal air (Willms et al., 1997 , 1999 ; Hunt, 2003 ). The
objectives of this study were to use such a measurement system to answer two questions: Can the
findings of Amthor et al. (2001 ) of no instantaneous effect of doubling of [CO2] on leaf Rd,O2 be
extended to a wide range of species, growth conditions, and measurement [CO2]? Using a range of
long-term experiments in which a wide range of plants have been grown under elevated [CO2] in
the field (Table I) is Rd,O2 decreased or unaffected as a result of acclimation to elevated [CO2]?
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Table I. Species, [CO2] treatments, plant and leaf developmental stage at the
time of measurement
Locations for material growing in the field and microclimate for controlled
environment grown plants are given under “Materials and Methods”.
References providing a description of each of the long-term field elevated
[CO2] experiments used are included.
RESULTS
There was no significant instantaneous effect of [CO2] on leaf Rd,O2 in any of the nine species
studied (Fig. 1; Tables II) and III). The results for the ANOVA were, for CO2, F3,332 = 0.032, P
= 0.99; and for species x CO2, F24,332 = 0.528, P = 0.97. A complete lack of any effect of [CO2]
was evident in all species not only on doubling [CO2], but when [CO2] was increased more than
five times above the current ambient concentration or decreased to zero (Fig. 1; Tables II and III).
The extremely high probabilities for accepting the null hypothesis (P > 0.97) eliminate any
possibility of a Type II error, i.e. a difference undetected because of variability. This is also
reflected in the combined regression of response relative to ambient [CO2] (Fig. 1), where the
probability that the response is independent of [CO2] was P = 0.99 (F3,360 < 0.001). This lack of
response of respiration to elevated [CO2] was independent of treatment method (F3,360 = 0.222, P
= 0.88), developmental stage (F3,360 = 0.174, P = 0.92), beginning or end of night (F3,360 =
0.638, P = 0.59), and the [CO2] at which the plants had been grown (F3,100 = 0.080, P = 0.97).
Although there was no interaction with [CO2], absolute rates of respiration generally decreased
with time in the dark.
Figure 1. The instantaneous effect of change in [CO2],
from the current ambient concentration, on leaf respiratory
O2 uptake in the dark. Respiration is plotted as the
percentage of change (±1 SE) relative to the rate of O2
consumption at current ambient [CO2]. Each mean
illustrated is based on measurements of 274 plants and nine
species (Table I). Leaves were in darkness and their
temperature was maintained at 25°C. Results for an analysis
View larger version (13K): of variance (ANOVA) are given in the results text, and
[in
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[in
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Table II. The response of leaf respiration to changes in [CO2] for plants
grown at current [CO2] only.
The mean dark leaf O2 uptake on an area basis (micromoles per minute per
second), and its SE (n = 6), are given for each of the four species at the
beginning and end of the 10-h night.
Table III. The response of leaf respiration to changes in (CO2) for species
grown at long-term elevated (CO2) experiments
The mean respiration rate and its SEM are given on an area basis and a dry weight
basis. n indicates the number of plots or open-top chambers from which
measurements of each treatment were made. Bold values show the critical longterm comparison of leaves grown and measured at their growth concentrations.
An asterisk indicates that leaves grown and measured at elevated (CO2) have
significantly different rates (P < 0.05) to those grown and measured at current
ambient (CO2).
Despite the lack of an instantaneous effect, long-term growth at elevated [CO2] did affect
respiration. Leaves grown and measured at elevated [CO2] had the same or a significantly higher
Rd,O2 than those grown and measured at current ambient [CO2] (Table III). The largest increases in
respiration were for Quercus geminata (23%) and soybean (22%) on a leaf area basis. By contrast,
there was no increase in Acer saccharum, Betula papyrifera, and Populus tremuloides (Table
IIIa). When all values from the long-term experiments were combined, there was a significant
increase in Rd,O2 of 11% and 7% on a leaf area and mass basis, respectively, and relative to
controls (Fig. 2).
Figure 2. The effect of long-term (lifetime or >3 years)
growth at elevated [CO2] in the field on leaf respiration.
The data points show mean leaf respiratory O2 uptake
(±90% confidence intervals) on an area and mass basis for
View larger version (8K): leaves grown at elevated [CO2] relative to controls grown at
[in
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[in
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using meta-analytical statistical methods.
TOP
ABSTRACT
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE
CITED
DISCUSSION
Six hundred separate measurements of different leaves, encompassing four long-term field CO2
enrichment experiments revealed no evidence of any decrease in Rd,O2 in response to
instantaneous elevation of [CO2]. This was consistent for seven tree and two crop species, the
majority sampled from long-term elevated [CO2] experiments where plants have developed
throughout their life or for several seasons under elevated [CO2]. This lack of response was
irrespective of developmental stage, growth conditions, and growth [CO2]. Bunce (2001 )
reported that the sensitivity of Rd,CO2 to an instantaneous elevation of [CO2] increased with
duration of the dark period. However, no effect on Rd,O2 was evident here (Tables II and III),
regardless of whether measurements were made at the start or end of the natural dark period.
Jahnke (2001 ) similarly found no effect of [CO2] on Rd,CO2 even after 72 h of darkness. Tjoelker
et al. (2001 ) noted that the instantaneous decrease in Rd,CO2 with elevated [CO2], although minor
(1.8%) in response to increase from 360 to 700 µmol mol-1, was far more substantial (11%) on
increase from 360 to 1,300 µmol mol-1. However, a 5-fold increase above current ambient [CO2]
or removal of CO2 from the atmosphere around the leaf did not alter Rd,O2 (Fig. 1). These findings
support the conclusions of Amthor et al. (2001 ) and of Jahnke (2001 ) and extend them to more
species and to major long-term [CO2] field manipulation studies. Although the results show no
effect of instantaneous elevation of [CO2] on Rd,O2, this cannot rule out some effect on apparent
Rd,CO2 due to non-respiratory processes, for example, CO2 uptake into carboxylic acids. However,
in one of the very few studies to measure CO2 and O2 exchange at different [CO2], Amthor et al.
(2001 ) found no effect of elevated [CO2] on respiratory quotient.
It has been suggested that the variability of response to instantaneous increase in [CO2] apparent
in the literature may be the result of species differences (Drake et al., 1999 ; Baker et al., 2000 ;
Hamilton et al., 2001 ). Without definitive testing, it is impossible to be certain on this point.
However, this study contains a diverse range of species and functional types (including C3 and C4
photosynthetic types, herbaceous and woody forms, two major crops, and seven forest dominant
gymnosperm and angiosperm species) that individual exceptions exist now seem somewhat
improbable. Moreover, we deliberately selected species previously reported to show some of the
largest inhibitions of respiration by elevated CO2 with previous methods and technology: soybean
40% (Bunce, 1995 ), Pinus taeda 14% (Teskey, 1995 ), Quercus rubra 5.6% (Amthor, 2000 ),
and maize (Zea mays) 46% (Cornic and Jarvis, 1972 ).
Any errors associated with measuring the instantaneous response of respiratory CO2 efflux to
elevated [CO2] may be equally applicable to measurement of the long-term effects of growth at
elevated [CO2] on Rd,CO2. The significant 7% increase in Rd,O2 on a mass basis, averaged across
seven species from four long-term experiments (Fig. 2), contrasts sharply with the average 18%
decrease reported by Wang and Curtis (2002 ) in their meta-analysis of Rd,CO2 on a mass basis
measured in long-term experiments. The significant increase on a mass, as well as an area basis,
suggests that increased respiration per unit leaf area is not just the result of the increased mass per
unit area that is commonly observed in leaves grown at elevated [CO2] (Drake et al., 1999 ).
However, as noted above, this mean increase masks marked differences between species in this
long-term response (Table III).
To our knowledge, this is the first study to use this alternative method to investigate the effects of
long-term growth at elevated [CO2] on respiration. The increase in respiration shown here is
consistent with the widely reported increase in leaf-soluble carbohydrates with long-term growth
at elevated [CO2] (Moore et al., 1999 ). Azcon-Bieto and Osmond (1983 ) showed that increase
in leaf nonstructural carbohydrate content produced by manipulation of photosynthesis during the
photoperiod increased subsequent dark respiration rates. Therefore, a similar response might be
expected when elevated [CO2] increases photosynthesis and leaf nonstructural carbohydrate
content during the photoperiod.
In conclusion, instantaneous elevation of [CO2] had no effect on leaf O2 uptake in these nine
species. This further confirms the suggestion that reports of respiratory inhibition, based on
measurements of CO2 uptake, are the result of experimental artifacts and not the result of any
sensitivity of plant respiration to the [CO2] at the time of measurement. A 15% to 20% reduction
of terrestrial plant respiration with a doubling of atmospheric [CO2] concentration would represent
some amelioration, at least temporarily, in the rate of rise in the global atmospheric [CO2] (for
review, see Gonzalez-Meler and Siedow, 1999 ). It can no longer be assumed that the direct effect
of elevated [CO2] on plant respiration will reduce future ecosystem CO2 efflux. On the basis of the
long-term field manipulations of [CO2] studied here, a small increase in respiration per unit leaf
area should be expected—this will amplify the effect of any increase in leaf area due to growth at
elevated [CO2].
MATERIALS AND METHODS
Growth Conditions
The species, developmental stage, growth conditions, and experimental site are listed in Table I.
Seven out of nine species were grown at current ambient (368 µmol mol-1) and at an elevated
[CO2] (550 or 700 µmol mol-1). Two of the species were measured at more than one
developmental stage; soybean (Glycine max) at vegetative and podfill stages, and Pinus taeda at
juvenile and mature stages. Soybean was grown in controlled environment- and open-field
conditions (FACE).
Oxygen Analysis System
Leaf O2 uptake rate was determined in an open gas exchange system incorporating a dual-cell
oxygen analyzer (S-3A/DOX; AEI Technologies, Pittsburgh) described by Willms et al. (1997 ),
and incorporating a stainless steel and glass leaf gas exchange cuvette (MPH-1000; Campbell
Scientific, Logan, UT). The analyzer incorporates a novel differential oxygen sensor obtained by
modifying two electrochemical fuel cells (model KE-25; Figaro USA, Wilmette, IL), placing one
in a reference gas stream and the other in an analytical gas stream, and then ensuring that the two
gas sensors were exposed to an identical temperature and pressure (Willms et al., 1997 ). The O2
fuel cells contain a Teflon membrane through which O2 can diffuse, and a weak acid electrolyte
bathes a lead anode and a gold cathode electrode. In the environment of the sensor, the O2 oxidizes
the lead, forming PbO, which dissolves into the solution and generates a small current. The
difference in output of the two sensors is directly proportional to the [O2] between the air streams.
The selective diffusive properties of Teflon and the specific electrochemistry ensure that the
differential sensor responds directly only to O2 in air. However, changes in partial pressures of
CO2 and water vapor due to respiration and transpiration will affect that of O2 causing an indirect
effect (Willms et al., 1999 ). For this reason, these gases are removed from the air stream before
the O2 sensors (Hunt, 2003 ). The analyzer was secured to a vibrationless table top and additional
layers of insulation were wound around the O2 measurement cells and instrument casing to further
improve resolution. The differential resolution of the modified dual-cell O2 analyzer was 1 to 2
µmol mol-1 against a background [O2] of 210 mmol mol-1. The cuvette was large enough to
enclose the entire leaf, leaflet, or cohort of needles; typically, 50 to 120 cm2 of leaf were enclosed.
This avoided errors associated with gas flow between the cuvette and surrounding air via the
intercellular air space that Jahnke and Krewitt (2002 ) have shown to result from the use of
cuvettes that only enclose part of the leaf. The cuvette (MPH-1000; Campbell Scientific) has been
described in detail previously (Bingham et al., 1980 ), adapted from the earlier design of Bingham
and Coyne (1977 ). Typical boundary layer conductances to water vapor are 2.5 to 4 mol m-2 s-1
(calculated from Bingham and Coyne, 1977 ; Bingham et al., 1980 ). A gas exchange system (LI6400; LI-COR, Lincoln, NE) was used to provide a controlled [CO2] to the cuvette, and air flow
rate was measured by a pair of flow meters (F350; AEI Technologies), calibrated as described
previously (Long et al., 1996 ). The gas flow across the leaf was maintained at 2.5 cm3 s-1. This
flow rate and the enclosed leaf area resulted in a [O2] of 40 to 80 µmol mol-1. The entry air
streams to the oxygen analyzer were scrubbed of water and CO2 by magnesium perchlorate and
soda lime. Column sizes were calculated to remove all CO2 and to be sufficient to dry the air
stream to the equilibrium vapor pressure of magnesium perchlorate (approximately 0.1 Pa). The
efficacy of these columns was tested by switching dry air (<0.01 kPa water vapor) with a
humidified air stream (2.0 kPa water vapor) at the cuvette inlet; no change in [O2] from 0 µmol
mol-1 was recorded. Similar tests showed that the soda lime column was effective in preventing
interference from differences in [CO2] between the cuvette and reference air streams.
The response time of the gas exchange system was assessed using two cylinders of compressed air
found to differ in [O2] by 80 µmol mol-1. Air from one of these cylinders was fed to the empty
cuvette inlet and also the reference air stream until a constant [O2] of 0 µmol mol-1 was obtained
at the analyzer. Air from the second cylinder was then switched in at the cuvette inlet and the time
taken to obtain a constant [O2] of 80 µmol mol-1 at the analyzer was recorded. The change was
98% complete within 10 min and complete within the resolution of the analyzer at 15 min.
Therefore, it was assumed that 20 min was adequate to record any response of respiratory O2
efflux to an instantaneous change in [CO2].
Respiration Measurements
Respiration was measured in the last 2.5 h of the night in all species and additional measurements
were made in the first 2.5 h of the night for three species (Table I). Measurements were made at
each location listed in Table I. However, because of its size and stability, the gas exchange system
was housed in a field laboratory at each site and leaves were detached shortly before measurement.
Petioles were cut under water and their cut ends were kept immersed in water until measurements
were complete. The leaves remained turgid throughout. For pot-grown soybean (Table I), parallel
measurements on attached and detached leaves were made; no significant differences in
respiration or its response to [CO2] were found. Leaves were equilibrated in the dark to 25°C
before measurement. For a single replicate measurement, respiration was measured at a [CO2] of
2,000 µmol mol-1, and then again at 700, 550, 360, and 0 µmol mol-1 CO2. To avoid systematic
error, each alternate replicate measurement was started at 0 µmol mol-1 CO2 and was then
switched to 368, 700, 550, and 2,000 µmol mol-1 CO2. Species from the FACE experiments at
Duke Forest, Rhinelander, and the University of Illinois, and the open-top chamber experiment at
Merritt Island were measured at 550 µmol mol-1 [CO2] rather than at 700 µmol mol-1 [CO2]
because this was the growth concentration (Table I). After each step-wise change in [CO2], a 20min waiting period ensured adequate time for any instantaneous response in Rd,O2. Each set of
measurements took 80 min to complete, and leaves were maintained in the dark at a leaf
temperature of 25°C for the entire measurement period. After each set of measurements, leaf area
was measured by digital imaging and the leaves were then dried to a constant weight at 80°C. Leaf
O2 evolution rates were calculated on a leaf area and a dry mass basis by adapting the CO2 uptake
equations of von Caemmerer and Farquhar (1981 ) for open system gas exchange measurements.
Statistical Analysis
In the four long-term field experiments, replicate number was determined by the number of
treatment plots (Table III). This avoided pseudoreplication. A minimum of three repeat
measurements were taken in each treatment plot of each experiment, and their pooled values
provided the individual sample measure. Only these data were used to assess the effect of longterm growth at elevated [CO2] on respiratory O2 uptake. Outside of these long-term elevated [CO2]
studies, measurements were made from six randomly selected plants each of P. taeda seedlings,
Q. rubra mature trees, and maize flowering plants growing under current ambient [CO2] in the
open. Soybean was grown in two separate controlled environment studies, and here, replicates
represent the number of separate chambers used (Table I).
ANOVA was used to test the effect of immediate changes of [CO2] on leaf respiration rate for all
nine species combined (SYSTAT, Evanston, IL). This tested the effect of species, growth
conditions, developmental stage, time of measurement, and growth [CO2] on the instantaneous
response to elevated [CO2]. In addition, a regression analysis of response relative to ambient
[CO2] was tested (SYSTAT).
Because of the very different experimental designs, durations, and treatment procedures of the
experiments in which plants were grown at elevated [CO2], and the possibility that species may
respond differently to long-term growth in elevated [CO2], the pooled effect of all field studies
was assessed by meta-analytical statistical techniques (Hedges et al., 1999 ; Ainsworth et al.,
2002 ; Morgan et al., 2003 ) using a software package (MetaWin, Rosenberg et al., 2000). This
allows determination of the consensus of studies differing in design. The natural log of the
response ratio (respiratory O2 uptake for leaves grown at elevated [CO2]/respiratory O2 uptake for
leaves grown at the current ambient [CO2]) was used as the metric for analysis (Hedges et al.,
1999 ). A mixed-model analysis was used based on the assumption of random variation in effect
sizes between individual measurements. A weighted parametric analysis was used (Gurevitch and
Hedges, 1999 ), and each individual observation was weighted by the reciprocal of the mixedmodel variance (Hedges et al., 1999 ). If the response ratio was not significantly different from
unity, growth at elevated [CO2] failed to change leaf O2 uptake from that at current ambient [CO2].
However, if significantly different from 1, growth at elevated [CO2] on average altered O2 uptake
across the studies.
ACKNOWLEDGMENTS
We thank Henry Ginsberg and Joseph Veltre of AEI Technologies, and Nicholas Dowling of
Qubit Systems for technical help and modifications to the O2 analysis system; Jeff Amthor,
Miquel Gonzalez-Meler, Andrew Leakey, and Shawna Naidu for comments on draft versions of
the manuscript; Carlos Pimentel and Kate George for the supply of plant material; and Lisa
Ainsworth for advice on the meta-analysis.
Received July 20, 2003; returned for revision August 20, 2003; accepted September 25, 2003.
FOOTNOTES
1
This work was supported by the U.S. Department of Energy’s Office of Science (to B.E.R.), as
were the long-term elevated [CO2] experiments in Florida, North Carolina, and Wisconsin.
2
Present address: Department of Biological Sciences, University of Essex, Colchester, CO4 3SQ,
UK.
3
Present address: Dipartimento di Scienze dell’ Ambiente Forestale e delle Sue Risorse,
Università della Tuscia, Via S. Camillo de Lellis, 01100 Viterbo, Italy.
Article,
publication
date,
and
citation
information
www.plantphysiol.org/cgi/doi/10.1104/pp.103.030569.
•
can
be
found
at
Corresponding author; e-mail stevel@life.uiuc.edu; fax 217-244-7563.
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INVITED REVIEW
PubMed
Plant Respiration and Elevated Atmospheric CO2
Concentration:
Cellular
Responses
and
Global
Significance
PubMed Citation
Articles by GONZALEZ-MELER, M. A.
Articles by TRUEMAN, R. J.
MIQUEL
A.
GONZALEZ-MELER*,
TANEVA and REBECCA J. TRUEMAN
LINA
Agricola
Department of Biological Sciences, University of Illinois
at Chicago, 845 West Taylor St, Chicago, IL 60607, USA
•
Articles by GONZALEZ-MELER, M. A.
Articles by TRUEMAN, R. J.
For correspondence. E-mail mmeler@uic.edu
Received: 2 June 2004
September 2004
Returned for revision: 14 June 2004
Accepted: 6 July 2004
Published electronically: 8
TOP
ABSTRACT
INTRODUCTION
DIRECT AND INDIRECT EFFECTS...
INTEGRATED EFFECTS OF ELEVATED...
ACKNOWLEDGEMENTS
LITERATURE
CITED
ABSTRACT
•
Background Elevated levels of atmospheric [CO2] are likely to enhance photosynthesis and
plant growth, which, in turn, should result in increased specific and whole-plant respiration rates.
However, a large body of literature has shown that specific respiration rates of plant tissues are
often reduced when plants are exposed to, or grown at, high [CO2] due to direct effects on
enzymes and indirect effects derived from changes in the plant’s chemical composition.
•
Scope Although measurement artefacts may have affected some of the previously reported
effects of CO2 on respiration rates, the direction and magnitude for the effects of elevated [CO2]
on plant respiration may largely depend on the vertical scale (from enzymes to ecosystems) at
which measurements are taken. In this review, the effects of elevated [CO2] from cells to
ecosystems are presented within the context of the enzymatic and physiological controls of plant
respiration, the role(s) of non-phosphorylating pathways, and possible effects associated with plant
size.
•
Conclusions Contrary to what was previously thought, specific respiration rates are
generally not reduced when plants are grown at elevated [CO2]. However, whole ecosystem
studies show that canopy respiration does not increase proportionally to increases in biomass in
response to elevated [CO2], although a larger proportion of respiration takes place in the root
system. Fundamental information is still lacking on how respiration and the processes supported
by it are physiologically controlled, thereby preventing sound interpretations of what seem to be
species-specific responses of respiration to elevated [CO2]. Therefore the role of plant respiration
in augmenting the sink capacity of terrestrial ecosystems is still uncertain.
Key words: Respiration, elevated CO2, cellular processes, ecosystem respiration, oxidation
INTRODUCTION
Respiration is essential for growth and maintenance of all plant tissues, and plays an important
role in the carbon balance of individual cells, whole-plants and ecosystems, as well as in the global
carbon cycle. Through the processes of respiration, solar energy conserved during photosynthesis
and stored as chemical energy in organic molecules is released in a regulated manner for the
production of ATP, the universal currency of biological energy transformations, and reducing
power (e.g. NADH and NADPH). A quantitatively important by-product of respiration is CO2 and,
therefore, plant and ecosystem respiration play a major role in the global carbon cycle.
Terrestrial ecosystems exchange about 120 Gt of carbon per year with the atmosphere, through the
processes of photosynthesis and respiration (Schlesinger, 1997 ). Roughly, half of the CO2
assimilated annually through photosynthesis is released back to the atmosphere by plant
respiration (Gifford, 1994 ; Amthor, 1995 ). Because terrestrial biosphere-atmosphere fluxes of
CO2 far outweigh anthropogenic inputs of CO2 to the atmosphere, a small change in terrestrial
respiration could have a significant impact on the annual increment in atmospheric [CO2]
(Amthor, 1997 ). A large body of literature has indicated that plant respiration is reduced in plants
grown at high [CO2]. For example, it was estimated that the observed 15–20 % reduction in plant
tissue respiration caused by doubling current atmospheric [CO2] (Amthor, 1997 ; Drake et al.,
1997 ; Curtis and Wang, 1998 ), could increase the net sink capacity of global ecosystems by 3·4
Gt of carbon per year (Drake et al., 1999 ), and thus offset an equivalent amount of carbon from
anthropogenic CO2 emissions. Therefore, not only are gross changes in respiration important for
large-scale carbon balance issues, changes in specific rates of respiration can have significant
impact on basic plant biology such as growth, biomass partitioning or nutrient uptake (Amthor,
1991 ; Wullschleger et al., 1994 ; Drake et al., 1999 ).
Scaling the effects of an increase in atmospheric [CO2] on plant respiration at the biochemical
level to the whole ecosystem is difficult for at least two important reasons: (1) the fine and coarse
control points of respiratory pathways in tissues and whole plants are not well known; and (2) it is
unclear how respiration rates actively or passively adjust to the effects that elevated [CO2] may
have both upstream (e.g. on substrate availability) and downstream (e.g. on energy demand) of the
carbon oxidation pathways. In addition, accurate measurements of respiratory rates as CO2
evolution have been proven difficult with current techniques (Gonzalez-Meler and Siedow, 1999 ;
Hamilton et al., 2001 ; Jahnke, 2001 ), presenting yet another limitation to scaling up processbased measurements of respiration rates to organism, ecosystem or global levels. In this
document, we re-evaluate the theory on respiration responses to elevated [CO2] in view of recent
studies that have provided new insights on the effects of a short- and long-term change in
atmospheric [CO2] on plant respiration.
TOP
ABSTRACT
INTRODUCTION
DIRECT AND INDIRECT EFFECTS...
INTEGRATED EFFECTS OF ELEVATED...
ACKNOWLEDGEMENTS
LITERATURE
CITED
DIRECT AND INDIRECT EFFECTS OF [CO2] ON
PLANT-SPECIFIC RESPIRATION RATES
Amthor (1991) described two different effects of [CO2] on plant respiration rates that can be
distinguished experimentally: a direct effect, in which the rate of respiration of mitochondria or
tissues could be rapidly and reversibly reduced following a rapid increase in [CO2] (Amthor et al.,
1992 ; Amthor, 2000a ), and an indirect effect in which high [CO2] changes the rate of
respiration in plant tissues compared with the rate seen for those plants grown in normal ambient
[CO2] when both are tested at a common background [CO2] (Azcón-Bieto et al., 1994 ). Most of
the studies investigating the effects of a change in [CO2] on respiration rates have described
magnitude (from non-significant to 60 %) and direction (either stimulation, inhibition or no effect)
of these two effects, with little progress on their underlying mechanisms. Confounding
measurement artefacts are an added complication for establishing the presence and magnitude of
direct and indirect effects of [CO2] on respiration (Gonzalez-Meler and Siedow, 1999 ; Amthor,
2000a ; Jahnke, 2001 ; Davey et al., 2004 ).
Direct
effects
Previously, a rapid short-term doubling of current atmospheric CO2 levels was reported to inhibit
respiration of mitochondria and plant tissues by 15–20 % (Amthor, 1997 ; Drake et al., 1997 ;
Curtis and Wang, 1998 ). Although direct effects of [CO2] on respiratory enzymes have been
reported, the magnitude of the direct inhibition of intact tissue respiration by elevated [CO2] has
now been shown to be explained by measurement artefacts (i.e. Gonzalez-Meler and Siedow,
1999 ; Jahnke, 2001 ), diminishing the impact that such reductions in respiration rate may have
on plant growth and the global carbon cycle.
Direct
effects
on
enzymes
Doubling current levels in [CO2] (here we review the effects of only two- or three-fold increases
in atmospheric CO2) have been shown to inhibit the oxygen uptake of isolated mitochondria and
the activity of mitochondrial enzymes under some conditions (Reuveni et al., 1995 ; GonzàlezMeler et al., 1996a , b ). Twice current atmospheric [CO2] reduces the activity of cytochrome c
oxidase and succinate dehydrogenase in isolated mitochondria from cotyledon and roots of
Glycine max (Gonzàlez-Meler et al., 1996b ). Mitochondrial oxygen uptake may show either no
response or up to 15 % inhibition by a rapid increase in [CO2], depending on the electron donor
used in the assay (Gonzàlez-Meler et al., 1996b ; see also Fig. 1), indicating that the response of
mitochondrial enzymes to high [CO2] depends on the cell’s metabolism (Gonzalez-Meler and
Siedow, 1999 ; Affourtit et al., 2001 ).
FIG. 1. Effect of doubling the concentration of CO2 on the
oxygen uptake of mitochondria isolated from 5-d-old
soybean cotyledons in state 4 conditions (absence of ADP).
Oxidation of NADH and pyruvate was measured in intact
mitochondria that were incubated for 10 min with 0 (open
bars) or 0·1 (solid bars) mM dissolved inorganic carbon
(DIC) at 25 °C. Experiments were done using the oxygen
isotope technique as described in Ribas-Carbo et al.
(1995) to detect the effects of CO2 on the total activity of
mitochondrial respiration (Vt), the activity of the
View larger version (19K): cytochrome pathway (vcyt) and the activity of the
[in
this
window] alternative pathway (valt). Results are average of four
[in
a
new
window] replicates and standard errors.
In scaling the direct effects of high [CO2] on respiratory enzymes to intact tissues two factors need
to be considered: (1) cytochrome oxidase exerts up to 50 % of the total respiratory control (for
definitions, see Kacser and Burns, 1979 ) at the level of the mitochondrial electron transport chain
(reviewed by Gonzalez-Meler and Siedow, 1999 ); and (2) the competitive nature of the
cytochrome and alternative pathways of mitochondrial respiration (Affourtit et al., 2001 ). The
activity of the alternative pathway of respiration could increase upon doubling of the [CO2],
masking a direct CO2 inhibition of the cytochrome pathway (Fig. 1). The oxygen isotope
technique (Ribas-Carbo et al., 1995 ) allows for the distinction of direct effects of [CO2] on
oxygen uptake activity by the cytochrome and the alternative oxidases. Figure 1 shows that
mitochondrial electron transport activity is affected when CO2 (a mild inhibitor) restricts the
normal electron flow through one of the pathways (Cyt pathway – vcyt). Under conditions of low
alternative pathway activity (for details, see Ribas-Carbo et al., 1995 ), inhibition of the
cytochrome pathway (18 %) by doubling the ambient [CO2] is compensated for by a similar
increase in alternative pathway activity (valt), resulting in no significant reduction in overall
oxygen uptake of the isolated mitochondria. These results show that enhanced activity of the
alternative pathway upon an increase in [CO2] provides a mechanism by which the direct [CO2]
inhibition of the cytochrome oxidase may not be seen in the total oxygen uptake under conditions
of high adenylate control (low ADP). Gonzàlez-Meler and Siedow (1999) argued that direct
effects of CO2 on respiration exceeding >10 % inhibition are likely due to factors other than
inhibition of mitochondrial enzymes. Due to confounding measurement artefacts (Jahnke, 2001 ),
rates of tissue respiration have been reported to decrease >10 % when the atmospheric [CO2]
doubles.
Direct
effects
on
intact
tissues
Recent evidence indicates that, in studying direct effects of CO2 on respiratory CO2 efflux of
intact tissues, investigators should first be concerned about measurement artefacts. Amthor
(1997) and Gonzalez-Meler and Siedow (1999) showed that gas CO2 exchange measurement
errors could augment and explain the magnitude of the direct effect. New studies made in this
context have shown that respiration rates are little or not at all inhibited by a doubling of
atmospheric [CO2] (Amthor, 2000a ; Amthor et al., 2001 ; Bunce, 2001 ; Hamilton et al., 2001
; Tjoelker et al., 2001 ; Bruhn et al., 2002 ; Davey et al., 2004 ). Indeed, Jahnke (2001) and
Jahnke and Krewitt (2002) confirmed that measurement artefacts due to leakage in CO2exchange systems could be as large as the previously reported direct inhibitory effects. They also
found that leaks through the aerial spaces of homobaric leaves showed a significant apparent
inhibition of CO2 efflux that was not due to an inhibition of respiration by elevated [CO2].
Therefore, considerations of the impact direct effects of [CO2] on plant respiration rates may have
on the global carbon cycle were overstated (Gonzalez-Meler et al., 1996a ; Drake et al., 1999 ).
Corrections for instrument leaks can be applied in most cases (Bunce, 2001 ; Jahnke, 2001 ; Pons
and Welschen, 2002 ). Applying some corrections for gas exchange leaks, Bunce (2001)
reported a significant reduction in respiration after a rapid increase in [CO2]. Moreover,
simultaneously switching the air isotopic composition from 12CO2 to pure 13CO2 when the [CO2]
was changed, showed that the efflux of 12CO2 (which represents net respiratory CO2 evolution
originating from respiratory substrates minus the CO2 being re-fixed by carboxylases, see below)
was reduced in leaves of C3 plants, but not C4, when the [CO2] increased (Pinelli and Loreto,
2003 ). These recent results illustrate that not all reports of direct inhibition of respiration by
elevated [CO2] have yet been reconciled with each other, but they do not show a clear proof that
the direct effect, as defined by Amthor (1991) exits either.
An alternate explanation for the apparent reduction in respiration rates upon an increase in [CO2]
(Bunce, 2001 ; Pinelli and Loreto, 2003 ) is an increase in dark CO2 fixation catalysed by
phosphoenolpyruvate carboxylase (PEPC), resulting in an apparent reduction of CO2 efflux
(Amthor, 1997 ; Drake et al., 1999 ). However, in Rumex and Glycine leaves, Amthor et al.
(2001) found an effect of elevated [CO2] on CO2 efflux in only a minority of experiments, with
no effect of elevated [CO2] on the O2 uptake of the same leaves, indicating at most a small effect
of elevated [CO2] on PEPC activity or respiration rates. Similar results were found for a variety of
species, involving 600 measurements, in which [CO2] increases did not alter the CO2 and O2 leaf
exchanges in the dark (Davey et al., 2004 ).
It seems that direct effects of [CO2] on mitochondrial enzymes may have no consequence on the
specific respiratory rate of whole tissues. Because mitochondrial respiratory enzymes are generally
‘in excess’ to the levels required to support normal tissue respiratory activity under most growth
conditions (Gonzalez-Meler and Siedow, 1999 ; Atkin and Tjoelker, 2003 ; Gonzalez-Meler and
Taneva, 2004 ), a minor inhibition of enzymatic activity by [CO2] will not translate to the overall
tissue respiration rate. A small increase in enzyme levels in plants exposed to elevated [CO2]
could be enough to compensate for the direct effects of CO2 on mitochondrial enzyme activity, as
it has been seen in leaves of some plants grown at high [CO2] (i.e. Griffin et al., 2001 ).
Another factor to consider is that increases in alternative pathway activity upon inhibition of
cytochrome oxidase by a change in [CO2] will result in unaltered dark respiratory rates (as
illustrated in Fig. 1). However, an increase in the non-phosphorylating activity of the alternative
pathway reduces the efficiency with which energy from oxidized substrates supports growth and
maintenance of plants (Gonzalez-Meler and Siedow, 1999 ; Gonzalez-Meler and Taneva, 2004 ).
Some studies have reported that the growth of plants exposed to elevated [CO2] only during the
night-time is altered (Bunce, 1995 , 2001 , 2002 ; Reuveni et al., 1997 ; Griffin et al., 1999 ).
Such altered growth patterns have been attributed, in part, to effects of [CO2] on plant respiration,
including increased activity of the non-phosphorylating alternative pathway. These CO2 effects are
considered next.
Are
there
direct
effects
of
CO2
on
whole
plants?
In the past, in some studies, plants have been grown at [CO2] elevated only at night-time to assess
the long-term consequences of the previously considered direct effects of CO2 on respiration rates.
Although most of these studies have found that growth patterns were altered in plants exposed to
high night-time [CO2], these effects cannot be attributed to direct effects of [CO2] on respiration
for reasons explained above. Bunce (1995) observed that the biomass and leaf area ratio of
Glycine max increased and that photosynthetic rates decreased in plants exposed to high nighttime [CO2] relative to plants grown at normal ambient [CO2]. Similarly, Griffin et al. (1999)
found that Glycine max exposed to high night-time [CO2] had lower leaf respiration rates and
greater biomass than plants grown at ambient or elevated [CO2]. Reuveni et al. (1997) speculated
that increases in the biomass of Lemna gibba grown at high night-time [CO2] relative to control
plants, was due to a reduction in alternative pathway respiration (although the alternative pathway
activity was not measured). Reduction in alternative pathway activity might more fully couple
respiration rates with growth and maintenance, enhancing growth. In contrast, Ziska and Bunce
(1999) showed that elevation of night-time [CO2] reduced biomass only in two of the four C4
species studied. In a later study, Bunce (2002) described that carbohydrate translocation was
reduced within 2 d of exposing plants to elevated night-time [CO2] when compared with plants
grown at ambient conditions day and night. These results suggest that elevated [CO2] may have
other uncharacterized direct effects on plant physiology that can have the consequence of reducing
energy demand for carbohydrate translocation, hence reducing the rate of leaf respiration. If so,
these new types of effects cannot be catalogued as direct effects of [CO2] on respiration but as
indirect effects, making more research in this area necessary.
In summary, even though rapid changes in [CO2] can inhibit the activity of some mitochondrial
enzymes directly, previously reported direct effects of [CO2] on tissue respiration are likely to be
due to measurement artefacts. Therefore, direct effects of [CO2] on specific respiration rates
(although not necessarily on respiration physiology) should be dismissed as having a major impact
on the amount of anthropogenic carbon that vegetation could retain. The role of the alternative
pathway in direct respiratory responses to elevated [CO2], although unresolved, would have little
impact, if any, on the general conclusion that direct effects of [CO2] on plant respiration are not to
be considered in plant growth or carbon cycle models. Other effects of long-term elevation of
night-time [CO2] can alter plant growth characteristics, and indirectly affect both respiration rates
and the relative activity of the cytochrome and alternative pathways. Therefore these effects
cannot be referred to as direct effects but rather as indirect effects of [CO2] on respiration rates.
Indirect
effects
Indirect CO2 effects represent changes in tissue respiration in response to plant growth at elevated
atmospheric [CO2] (Amthor, 1997 ), because of changes in tissue composition (Drake et al.,
1997 ). Other indirect effects of [CO2] on plant respiration include changes in growth or response
to environmental stress, as well as changes in the respiratory demand for energy relative to that of
plants grown at ambient [CO2] (Bunce, 1994 ; Amthor, 2000b ). Such indirect effects of plant
growth at elevated [CO2] can be detected as a reduction in CO2 emission (or O2 consumption)
from plant tissues when measured at ambient [CO2] (Gonzalez-Meler et al., 1996a ).
Indirect
effects
on
enzymes:
is
there
acclimation
of
respiration
to
elevated
[CO2]?
Acclimation of respiration to high [CO2] can be defined as the down- or up-regulation of the
respiratory machinery (i.e. amount of respiratory enzymes, number of mitochondria) irrespective
of changes in specific respiration rates when plants are grown at elevated [CO2] (Drake et al.,
1999 ). A change in the leaf respiratory machinery of plants grown at high [CO2] can be expected
because of (a) increases in carbohydrate content, (b) reduced photorespiratory activity and (c)
reductions in leaf protein content (15 % on average), when compared with plants grown at current
ambient [CO2] (Drake et al., 1997 ). These three processes can have different and sometimes
opposite effects on levels of respiratory enzymes and specific rates of respiration.
As atmospheric [CO2] rises, increased photosynthesis results in higher cellular carbohydrate
concentrations (Drake et al., 1997 ; Curtis and Wang, 1998 ). Increased carbohydrates can
stimulate the specific activity of respiration, due to the greater availability of respiratory substrates
(Azcón-Bieto and Osmond, 1983 ) and higher energy demand for phloem loading of
carbohydrates (Bouma et al., 1995 ; Körner et al., 1995 ; Amthor, 2000b ). Additionally,
increased tissue carbohydrate levels (as in the case of plants grown at high [CO2]) could result in
an increase of transcript levels of cytochrome oxidase (Felitti and Gonzalez, 1998 ; Curi et al.,
2003 ) and cytochrome pathway activity (Gonzalez-Meler et al., 2001 ). In contrast, a reduction
in photorespiratory activity in plants grown at high [CO2] will reduce the need for the
mitochondrial compartment, which may result in a reduction in mitochondrial proteins and
functions (Amthor, 1997 ; Drake et al., l999 ; Bloom et al., 2002 ). Not surprisingly, the
available studies show that levels and activity of respiratory machinery can either increase or
decrease in leaves of plants grown at high [CO2] with no concomitant effects on respiration rates
(Fig. 2).
FIG. 2. Relationship between the relative response of leaf
respiration and the leaf respiratory machinery of plants
grown at ambient and elevated [CO2]. Data extracted from
Azcon-Bieto et al. (1994) , George et al. (1996) ,
Gonzalez-Meler et al. (1996a) , Griffin et al. (2001 ,
2004) , Hrubec et al. (1985) , Tissue et al. (2002) , Wang
et al. (2004) and M. A. Gonzalez-Meler (unpubl. res.).
*
Respiratory machinery refers to changes in either number
of
View larger version (14K): mitochondria, or soluble or membrane enzyme activity in
[in
this
window] plant extracts from plants grown at either ambient or
[in
a
new
window] elevated [CO2] in field and laboratory experiments. Only
studies that have provided both respiration rates and
respiratory machinery from the same tissues or from
different tissues from the same plants are considered. Rates
of [CO2] evolution used are only those measured for plants
grown under ambient or elevated [CO2] at ambient [CO2]
conditions.
For instance, indirect effects of elevated [CO2] on respiration of leaves of Lindera benzoin and
stems of Scirpus olneyi were correlated with a reduction in maximum activity of cytochrome
oxidase (Azcon-Bieto et al., 1994 ). However, a reduction of respiratory enzyme activity was not
seen in rapidly growing tissues exposed to elevated [CO2] (Perez-Trejo, 1981 ; Hrubeck et al.,
1985 ). No acclimation to elevated [CO2] (i.e. reduction in cytochrome oxidase) has been
observed in leaves of C4 plants (Azcon-Bieto et al., 1994 ) or roots of C3 plants (Gonzalez-Meler
et al., 1996a ). In contrast, the number of mitochondria is consistently shown to increase in
response to plant growth at elevated [CO2] (Griffin et al., 2001 ; Wang et al., 2004 ). Overall
changes associated with increases or decreases in the respiratory machinery in leaves of plants
grown at elevated [CO2] (i.e. mitochondrial counts, cytochrome oxidase maximum activity) seem
to be independent of responses of the respiration rates to elevated [CO2] (Fig. 2; r2 = 0·, n.s.).
Therefore, changes in the mitochondrial machinery of plants grown in high [CO2] are associated
with altered mitochondrial function and biogenesis and are not necessarily related to energy
production. Anaplerotic functions of mitochondrial activity provide reductant and carbon
skeletons for the production of primary and secondary metabolites used in growth and
maintenance processes, as well as other processes such as nitrogen reduction and assimilation
(Gonzalez-Meler et al., 1996a ; Amthor, 1997 ; Bloom et al., 2002 ; Davey et al., 2004 ).
Vanoosten et al., (1992) reported increases in fumarase and malic enzyme activities in leaves of
Picea abies grown in high [CO2]. These increases contrasted with unchanged or decreased activity
of PEPC or glycolitic enzymes. Increase in enzyme activity of the tricarboxylic acid cycle with no
increase in respiration rates may support the export of carbon skeletons and reducing power
(mainly as malate) from the mitochondria to the cytosol for biosynthesis. This uncoupling between
an increase in mitochondrial numbers and no increase in respiratory activity of plants grown at
high [CO2] (Fig. 2) is further illustrated by the study of Tissue et al. (2002) in which a more than
two-fold increase in the leaf mitochondrial counts did not alter the maximum activity of
cytochrome oxidase in the same tissues. With the exception of two studies (i.e. Azcon-Bieto et al.,
1994 ; Tissue et al., 2002 ), there are no reports on the response of membrane-associated
mitochondrial enzymes to elevated [CO2] in leaves or roots of plants. More research is needed in
this area.
Indirect
effects
of
[CO2]
on
respiration
of
tissues
and
whole
plants
It has been historically accepted that leaf respiration is reduced as a consequence of plant growth
at elevated [CO2] (El Kohen et al., 1991 ; Isdo and Kimball, 1992 ; Wullschleger et al., 1992a ;
Amthor, 1997 ; Drake et al., 1997 ; Curtis and Wang, 1998 ; Norby et al., 1999 ). Poorter et al.
(1992) showed that leaf respiration was reduced, on average, by 14 % when expressed on a leaf
mass basis, but increased by 16 % on a leaf area basis. Curtis and Wang (1998) compiled
respiratory data for woody plants, and observed that growth at elevated [CO2] resulted in an 18 %
inhibition of leaf respiration (mass basis). In this section we will concentrate on the effects of
elevated [CO2] on respiration of above-ground tissues. It is worth noting, however, that root
respiration rates in the field are not altered by growth at elevated [CO2] during most of the plant’s
growing season (Johnson et al., 1994 ; Matamala and Schlesinger, 2000 ). Total ecosystem root
respiration may increase if root mass and/or turnover increases (Hungate et al., 1997 ; Hamilton
et al., 2002 ; George et al., 2003 ; Matamala et al., 2003 ).
Many previous reviews and meta-analyses (see above) have compared respiratory rates of plants
grown and measured at the [CO2] at which plants were grown. Therefore, these respiration
measurements are also susceptible by the gas exchange artefacts described by Jahnke and coworkers. Respiratory O2 uptake measured in closed systems is not susceptible to the measurement
artefacts described above. The O2 uptake of green tissues of plants grown at high [CO2] could be
increased, reduced or unaltered when compared with plants grown at ambient [CO2] (Table 1).
Davey et al. (2004) also showed that the O2 uptake of leaves of plants grown at high [CO2] was
slightly increased relative to control plants. Gas exchange leaks (Jahnke, 2001 ) should not be a
factor in determining the rates of CO2 emission rates when rates are measured and compared at
ambient [CO2] for plants grown at ambient and elevated [CO2]. A re-analysis of the literature
focused on leaf respiratory responses (on a leaf mass basis) of plants grown at ambient and
elevated [CO2] and measured at an ambient [CO2], suggests that specific leaf respiration rates will
be unaltered or even increased in plants grown at elevated [CO2] (Table 2). Therefore, the
generally accepted onclusion that respiration rates of plants grown at elevated [CO2] is reduced
relative to plants grown at ambient [CO2] should be re-evaluated (Amthor, 2000a ; Davey et al.,
2004 ; Gonzalez-Meler and Taneva, 2004 ). However, there is significant variability in the leaf
respiratory response to growth at elevated [CO2] when compared with plants grown at ambient
conditions, ranging from 40 % inhibition (Azcon-Bieto et al., 1994 ) to 50 % stimulation
(Williams et al., 1992 ). Considerations on the physiological basis by which the acclimation
response of respiration to elevated [CO2] varies, is considered next.
View this
table:
[in
this
window]
[in a new
TABLE 1. Indirect effects of [CO2] on the dark O2 uptake rates (on a leaf area
basis) at 25 °C of green tissues from Scirpus olneyi (C3), Spartina patens (C4),
Distchilis spicata (C4), Lindera benzoin (C3), Cinna arundinacea (C3), Quercus
geminata (C3), Quercus myrtifolia (C3) and Serenoa repens (C3) plants grown at
either normal ambient (A) or normal ambient + 340 mL L–1 atmospheric CO2
window]
(E), inside open-top chambers in the field
View
this TABLE 2. Indirect effects of the elevation in atmospheric [CO2] on the dark
CO2 efflux of leaves on a dry mass basis based on literature surveys
table:
[in
this
window]
[in a new
window]
In boreal species, reduction of respiration of plants grown at high [CO2] was related to changes in
tissue N and carbohydrate concentration (Tjoelker et al., 1999 ). Tissue N concentration often
decreases in plants grown at elevated [CO2] (Drake et al., 1997 ; Curtis and Wang, 1998 ). It is
expected that respiration rate would be lower in tissues having lower [N], because the respiratory
cost associated with protein turnover and maintenance is a large portion of dark respiration
(Bouma et al., 1994 ). Hence, the metabolic cost (i.e. respiratory energy demand) for construction
and maintenance of tissues with high protein concentration is greater than the cost for the
maintenance of the same tissue with low [N] (assuming there are no changes in rates of protein
turnover between plants grown at ambient and elevated [CO2]) (Amthor, 1989 ; Drake et al.,
1999 ). This idea was confirmed for leaves of Quercus alba seedlings grown in open-top
chambers in the field, where respiration was 21–56 % lower under elevated [CO2] than under
ambient [CO2] (Wullschleger and Norby, 1992 ). The growth respiration component of these
leaves was reduced by 31 % and the maintenance component by 45 %. These effects were
attributed to the reduced cost of maintaining tissues having lower [N]. Similar results were
obtained in leaves and stems of plants of other species (Wullschleger et al., 1992a , b , 1997 ;
Amthor et al., 1994 ; Carey et al., 1996 ; Griffin et al., 1996b ; Dvorak and Oplustilova, 1997 ;
Will and Ceulemans, 1997 ), with some exceptions (e.g. Wullschleger et al., 1995 ).
The growth component of respiration (Amthor, 2000b ) could also be reduced in plants grown at
elevated [CO2] as a result of altered tissue chemistry (Griffin et al., 1993 ). Based on the chemical
composition of tissues, Poorter et al. (1997) found that elevated [CO2] could reduce growth
construction costs by 10–20 %. Griffin et al. (1993 , 1996a) also observed reductions in
construction costs of Pinus taeda seedlings grown at elevated [CO2]. Hamilton et al. (2001)
reported that elevated [CO2] slightly reduced construction costs of leaves of mature trees
(including P. taeda) at the top of the canopy, but not at the bottom of the canopy. Such a small
reduction can be explained by reductions in tissue [N], as observed in leaves exposed to high
[CO2] at the top of the canopy. Changes in construction costs did not result in a decrease in the
leaf respiration rates of P. taeda trees exposed to elevated [CO2] (Hamilton et al., 2001 ).
The lack of long-term effects of increased [CO2] on specific plant respiration rates could also be
due to a lower involvement of the alternative pathway (Gonzalez-Meler and Siedow, 1999 ;
Griffin et al., 1999 ). Respiration through the alternative pathway bypasses two of the three sites
of proton translocation; so the free energy released is lost as heat, and is unavailable for the
synthesis of ATP. Respiration associated with this pathway will not support growth and
maintenance processes of tissues as efficiently as respiration through the cytochrome path. On the
other hand, the activity of the alternative pathway of respiration could decrease upon doubling
[CO2], masking increases in the activity of the cytochrome pathway and making respiration more
efficient, as it was the case in understorey trees grown under elevated [CO2] (Gonzalez-Meler and
Taneva, 2004 ).
Despite earlier reports, the responses of plant respiration to growth under high [CO2] are variable
and perhaps species specific, although the overall trend may be a moderate increase in respiration
rates of leaves of plants grown under elevated [CO2] relative to the ambient ones (Tables 1 and 2).
Ultimately, altered specific respiration rates of tissues will depend on the net balance between the
demand for ATP from maintenance (including phloem loading and unloading) and growth
processes and on carbon allocation patterns between sinks and source tissues. The fact that the
mitochondrial machinery has been shown to increase in leaves of plants grown at elevated [CO2]
suggests, however, a larger participation of mitochondria in functions other than oxidative
phosphorylation. Finally, indirect effects of [CO2] on tissue respiration rates can be augmented or
offset by changes in plant size and/or changes in carbon allocation between plant parts.
TOP
ABSTRACT
INTRODUCTION
DIRECT AND INDIRECT EFFECTS...
INTEGRATED EFFECTS OF ELEVATED...
ACKNOWLEDGEMENTS
LITERATURE
CITED
INTEGRATED EFFECTS OF ELEVATED [CO2] ON
PLANT RESPIRATION AT THE ECOSYSTEM LEVEL
Terrestrial ecosystems exchange about 120 Gt of carbon per year with the atmosphere, through the
processes of photosynthesis (leading to gross primary production, GPP) and ecosystem respiration
(Re) (Schlesinger, 1997 ). The difference between GPP and Re determines net ecosystem
productivity (NEP), the net amount of carbon retained or released by a given ecosystem.
Currently, the net exchange of C between the terrestrial biosphere and the atmosphere is estimated
to result in a global terrestrial sink of about 2 Gt of carbon per year (Gifford, 1994 ; Schimel,
1995 ; Steffen et al., 1998 ). Unfortunately, the effects of [CO2] on plant and heterotrophic
respiration at the ecosystem level are not well understood, despite their potential to control
ecosystem carbon budgets (Ryan, 1991 ; Giardina and Ryan, 2000 ; Valentini et al., 2000 ).
Annually, terrestrial plant respiration releases 40–60 % of the total carbon fixed during
photosynthesis (Gifford, 1994 ; Amthor, 1995 ) representing about half of the annual input of
CO2 to the atmosphere from terrestrial ecosystems (Schlesinger, 1997 ). Therefore the magnitude
of terrestrial plant respiration and its responses to [CO2] are important factors governing the
intrinsic capacity of ecosystems to store carbon. Plant respiration responses to high [CO2] may
stem from several mechanisms (in absence of direct effects on respiration rates): (a) indirect
effects; (b) changes in total plant biomass; and (c) changes in plant carbon allocation. If it is
confirmed that the response of overall terrestrial plant respiration to an increase in atmospheric
[CO2] is small (Tables 1 and 2), then changes in global plant respiration should be proportional to
changes in biomass. However, experimental evidence suggests otherwise, or that the response of
plant respiration at the ecosystem level to elevated [CO2] may be more a function of carbon
allocation patterns rather than just of increases in plant size (Drake et al., 1996 ; Hamilton et al.,
2002 ).
Attempts to scale [CO2] effects on mitochondrial or tissue respiration to the ecosystem level are
problematic because, unlike photosynthesis, little is known about applicable scaling methods for
plant respiration (Gifford, 2003 ). Current attempts to build respiratory carbon budgets at the
canopy level require knowledge of maintenance and growth respiration (see above), as well as
tissue respiratory responses to light, temperature and [CO2] (Amthor, 2000b ; Gifford, 2003 ;
Turnbull et al., 2003 ) or have been based on theoretical respiration-to-photosynthesis ratios (i.e.
Norby et al., 2002 ). The respiration-to-photosynthesis ratio was not affected by elevated [CO2] in
soybean plants (Ziska and Bunce, 1998 ) but was reduced in pine forests stands (Hamilton et al.,
2002 ) when compared with ambient controls. The limited available data on the effects of
elevating the [CO2] on intact field ecosystems on plant and total ecosystem respiration are
summarized in Tables 3 and 4.
View this TABLE 3. Contribution of CO2-induced reduction in ecosystem respiration (Re)
to total net ecosystem productivity in a C3-dominated salt-marsh ecosystem
table:
[in
this exposed to elevated [CO2] since 1988
window]
[in a new
window]
View
table:
[in
window]
[in
a
window]
this TABLE 4. The effect of elevated [CO2] on ecosystem-level plant gas
exchange of forested ecosystems
this
new
The contribution of CO2-induced changes in ecosystem respiration to annual NEP of an intact salt
marsh exposed to elevated [CO2] in open-top chambers is shown in Table 3. In this ecosystem,
elevation of atmospheric CO2 by 350 µL L–1 over ambient levels consistently reduced night-time
ecosystem respiration in C3 and C4 community stands (Drake et al., 1996 ). Net ecosystem
productivity increased in stands exposed to elevated [CO2] because the decrease in respiration was
accompanied by increases in GPP. The reduction in annual respiration in this ecosystem
represented from one-fifth to one-third of the difference in NEP between ambient and elevated
open-top chambers (Table 3). Ecosystem-level plant respiration in forest stands, however, may not
be reduced by elevated [CO2] as in the case of the salt marsh (Table 4).
In forests exposed to elevated [CO2] using free air CO2 enrichment (FACE), GPP was stimulated
by 15–36 % (Table 4) over ambient stands, resulting in consistent increases in net primary
productivity (NPP). Increases in GPP and NPP seemed to be accompanied by variable responses
on whole-plant respiration, which could be increased by as much as 20 % (Table 4). For instance,
Hamilton et al. (2002) found that elevated [CO2] increased forest NPP by 27 % without any
increase in total plant ecosystem respiration. This result can be explained by a possible decrease in
specific respiration rates at high [CO2] accompanied by larger biomass of plants grown at elevated
[CO2]. In a Popolus deltoides plantation at Biosphere 2 centre, elevated [CO2] increased standlevel plant respiration by 40 % when compared with ambient (Table 4). The total increase in
stand-level plant respiration was larger than the CO2 stimulation of NPP, suggesting that, in this
case, specific respiration rates and plant size both contributed to the increase in plant respiration of
trees grown in high [CO2] (R. J. Trueman and M. A. Gonzalez-Meler, unpubl. res.).
Some studies suggest that increases in ecosystem-level plant respiration in ecosystems exposed to
elevated [CO2] mainly occur in below-ground plant tissues (Hungate et al., 1997 ; Lin et al.,
2001 ; Hamilton et al., 2002 ), which in turn may stimulate soil respiration rates (Zak et al.,
2000 ; Pendall et al., 2003 ). This requires a substantial proportion of the additional C
assimilated by plants growing at elevated [CO2] to be allocated to roots for growth and turnover
(Johnson et al., 1994 ; Hungate et al., 1997 ; King et al., 2001 ; Matamala et al., 2003 ). Greater
plant C allocation below ground can theoretically increase the contribution of root respiration to
total soil respiration because of greater root biomass relative to ambient [CO2]. However,
Hamilton et al. (2002) reported increases in total root respiration in a forest exposed to elevated
[CO2], where root mass, turnover (Matamala et al., 2003 ) or respiration rates (Matamala and
Schlesinger, 2000 ; George et al., 2003 ) were not affected by the CO2 treatment. The apparent
disparity may be due to the different methodologies employed in these studies and in the
contribution of rhizosphere activity to soil CO2 efflux, emphasizing the need for more coordinated
research in building ecosystem carbon budgets using multiple approaches, especially below
ground.
In conclusion and contrary to what was previously thought, specific respiration rates are generally
not reduced when plants are grown at elevated [CO2]. This is because direct effects of [CO2] on
respiratory enzymes are inconsequential to specific respiration rates of tissues. A re-analysis of the
literature comparing respiration of leaves of plants grown at ambient [CO2] with leaves of plants
grown at elevated [CO2] when rates are measured at the same [CO2], indicate that leaf respiration
on average will not be greatly changed by increasing atmospheric [CO2]. Increases in growth rates
appear to be compensated for by changes in tissue chemistry that affect growth and maintenance
respiration. If specific rates of respiration are not affected by growth at elevated [CO2], respiration
from the terrestrial vegetation in a high-CO2 world should be proportional to changes in plant
biomass. However, whole ecosystem studies show that canopy respiration does not increase
proportionally to increases in biomass when natural ecosystems are exposed to elevated
atmospheric [CO2]. Field studies also suggest that a larger proportion of plant respiration takes
place in the root system under elevated [CO2] conditions. Fundamental information is still lacking
on how respiration and the processes supported by it are physiologically controlled, thereby
preventing sound interpretations of what seem to be species-specific responses of respiration to
elevated [CO2]. Therefore the role of plant respiration in augmenting the sink capacity of
terrestrial ecosystems is still uncertain.
TOP
ABSTRACT
INTRODUCTION
DIRECT AND INDIRECT EFFECTS...
INTEGRATED EFFECTS OF ELEVATED...
ACKNOWLEDGEMENTS
LITERATURE
CITED
ACKNOWLEDGEMENTS
This review is part of a lecture given at the International Congress of Plant Mitochondria held in
Perth in July 2002, partly sponsored by Annals of Botany. M.A.G.-M. specially thanks Jeff
Amthor for his work in this area, his thoughts, discussions and specific comments in parts of this
manuscript. Authors also wish to thank Joaquim Azcon-Bieto, Bert Drake, Evan DeLucia, Steve
Long, Miquel Ribas-Carbo and Jim Siedow for their discussion on the effects of elevated [CO2]
on respiration of organelles, plants and ecosystems over the years. We acknowledge financial
support by US Department of Agriculture (M.A.G.-M.), UIC fellowship (L.T.), and Sigma Xi and
provost awards to L.T. and R.J.T.
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TOP
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INTRODUCTION
DIRECT AND INDIRECT EFFECTS...
INTEGRATED EFFECTS OF ELEVATED...
ACKNOWLEDGEMENTS
LITERATURE CITED
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Plant Physiol, October 2001, Vol. 127, pp. 607-614
Insertional Mutants of Chlamydomonas reinhardtii That Require Elevated CO2 for Survival1
Kyujung Van, Yingjun Wang, Yoshiko Nakamura, and Martin H. Spalding*
Interdepartmental Plant Physiology Major (K.V., Y.W., M.H.S.) and Department of Botany
(K.V., Y.W., Y.N., M.H.S.), 353 Bessey Hall, Iowa State University, Ames, Iowa 50011
TOP
ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
ABSTRACT
Aquatic photosynthetic organisms live in quite variable conditions of CO2 availability. To survive
in limiting CO2 conditions, Chlamydomonas reinhardtii and other microalgae show adaptive
changes, such as induction of a CO2-concentrating mechanism, changes in cell organization,
increased photorespiratory enzyme activity, induction of periplasmic carbonic anhydrase and
specific polypeptides (mitochondrial carbonic anhydrases and putative chloroplast carrier
proteins), and transient down-regulation in the synthesis of Rubisco. The signal for acclimation to
limiting CO2 in C. reinhardtii is unidentified, and it is not known how they sense a change of CO2
level. The limiting CO2 signals must be transduced into the changes in gene expression observed
during acclimation, so mutational analyses should be helpful for investigating the signal
transduction pathway for low CO2 acclimation. Eight independently isolated mutants of C.
reinhardtii that require high CO2 for photoautotrophic growth were tested by complementation
group analysis. These mutants are likely to be defective in some aspects of the acclimation to low
CO2 because they differ from wild type in their growth and in the expression patterns of five low
CO2-inducible genes (Cah1, Mca1, Mca2, Ccp1, and Ccp2). Two of the new mutants formed a
single complementation group along with the previously described mutant cia-5, which appears to
be defective in the signal transduction pathway for low CO2 acclimation. The other mutations
represent six additional, independent complementation groups.
TOP
ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
INTRODUCTION
Acclimation to changed environmental conditions is a key to survival for all organisms. In
response to perceived environmental signals, organisms may exhibit specific adaptive changes,
such as changes in the expression of key genes to survive specific environmental changes.
Because CO2 can vary substantially in aquatic habitats and represents the major substrate for
photosynthetic CO2 fixation via the enzyme Rubisco, CO2 concentration is an important
environmental signal in aquatic photosynthetic organisms including cyanobacteria and
Chlamydomonas reinhardtii.
Unlike terrestrial higher plants, aquatic photosynthetic organisms can face difficulties in acquiring
CO2. Because the CO2 diffusion rate in water is much slower than that in air (Badger and
Spalding, 2000 ), the CO2 supply to Rubisco in these aquatic photosynthetic organisms can
become limited. C. reinhardtii and other aquatic photosynthetic organisms have a genetic program
to allow them to acclimate to low CO2. This acclimation includes induction of a CO2concentrating mechanism (CCM) that allows the cells to acquire CO2 efficiently by increasing the
CO2 concentration around Rubisco under limiting CO2 conditions (Badger et al., 1980 ; for
review, see Spalding, 1998 ; Kaplan and Reinhold, 1999 ).
Along with the induction of the CCM, C. reinhardtii shows adaptive changes to limiting CO2
conditions, such as changes in cell organization (Geraghty and Spalding, 1996 ), increased
photorespiratory enzyme activity (Marek and Spalding, 1991 ), induction of periplasmic carbonic
anhydrase (CA) (pCA1, encoded by the Cah1 gene; Fujiwara et al., 1990 ; Fukuzawa et al.,
1990 ; Ishida et al., 1993 ), mitochondrial CA (mtCA, encoded by the Mca1 and Mca2 genes;
Eriksson et al., 1996 ; Geraghty and Spalding, 1996 ), and putative chloroplast carrier protein
(Ccp, encoded by the Ccp1 and Ccp2 genes; Geraghty et al., 1990 ; Ramazanov et al., 1993 ;
Chen et al., 1997 ), and transient down-regulation in the synthesis of Rubisco (Coleman and
Grossman, 1984 ; Winder et al., 1992 ).
The signal for acclimation to limiting CO2 in C. reinhardtii is unidentified. It is not known how
they sense a change of CO2 availability, whether by CO2 concentration directly or indirectly via a
cellular process such as carbohydrate metabolism. Whatever the limiting-CO2 signal, it must be
transduced into the changes in gene expression observed during acclimation, such as expression of
Cah1. A powerful way to identify components of the CCM and of the signal transduction pathway
for low CO2 acclimation is through the analysis and characterization of mutants specifically
defective in growth in limiting CO2, like the ca-1, pmp-1, and cia-5 mutants (Spalding et al.,
1983a ; 1983b ; Moroney et al., 1989 ). Using advances in nuclear transformation of
C. reinhardtii (Kindle, 1990 ), a collection of insertionally generated high CO2-requiring (HCR)
mutants unable to grow in limiting CO2 was obtained and is described here.
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ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
RESULTS
Generation and Isolation of Mutants
Using glass bead transformation (Kindle, 1990 ; Davies et al., 1994 ), CC425 (Table I) was
complemented by transformation with p-Arg7.8 (Debuchy et al., 1989 ) to generate a pool of
insertional mutants on CO2-minimal medium. Cells from each of more than 7,000 transformant
colonies were suspended in air-minimal medium and grown on plates in high CO2 (5% [v/v] CO2
in air), normal air, and low CO2 (50-100 µL L 1 CO2). HCR mutants, defined as those showing
little or no growth either in normal air or in low CO2, should include mutants, like cia-5, that are
defective in acclimation to limiting CO2, as well as those with functional defects in the CCM.
Sixteen putative HCR mutants were identified, and eight of those are described here (Table II).
Table I. Strains of C. reinhardtii used in the study
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Table II. Characteristics of HCR mutants
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General Characteristics of HCR Mutants
The eight HCR mutants and their general characteristics are shown in Table II and Figure 1.
When grown in high CO2 on agar, all HCR mutants except HCR105 were indistinguishable from
the wild type (Fig. 1). The eight HCR mutants could be divided into four groups based on their
apparent high CO2 requirement for photoautotrophic growth. The first group, including HCRP34,
HCR209, and HCR90, showed a leaky HCR phenotype in air but a stringent phenotype in low
CO2. The second group, including HCR86 and HCR105, showed a stringent HCR phenotype both
in air and in low CO2. HCR89 and HCR95, comprising the third group, had a leaky HCR
phenotype both in air and in low CO2. HCR3510 lacked a significant growth phenotype in air but
had a stringent phenotype in low CO2.
Figure 1. Spot tests for growth response to different CO2
concentrations for wild-type strains (CC849 and ars301),
four previously described HCR mutants (cia-5, ca-1, pmp1, and pgp-1), and eight new HCR mutants. Plates were
kept either at high CO2 (5% [v/v] CO2), at air level of CO2,
or at low CO2 (50-100 µL L 1) for 10 d.
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Genetic Characteristics of HCR Mutants
Seven of the eight HCR mutants were found by Southern analysis (data not shown) to contain
only one copy of the Arg7 insert, and the presence of vector sequences was confirmed in six
mutants (Table II). The presence of vector sequences provides an opportunity for the cloning of
sequences flanking the insert by plasmid rescue (Quarmby and Hartzell, 1994 ).
Selected random progeny and/or tetrads from HCR mutants were tested in crosses with another
arg2 mutant (CC1068, Table I) for linkage of the Arg insert with Arg+ and HCR phenotypes
(Table II). Five of the eight mutants showed cosegregation of the single Arg insert with the HCR
phenotype, suggesting that the Arg insert is responsible for the HCR phenotype in these five
mutants. In two of the mutants, HCR95 and HCR105, the inserts did not cosegregate with the
HCR phenotype, indicating that insertion of the Arg plasmid was not directly responsible for the
HCR phenotype in these two mutants. In HCR209, which has two inserts, cosegregation crosses
were not conclusive, but other evidence (see below) suggests the two inserts are tandemly
arranged and are responsible for the phenotype.
Heterozygous vegetative diploids, generated in crosses with CC1068 and selected by their
resistance to both kanamycin and streptomycin, were used to determine the dominant/recessive
nature of the HCR phenotype of each mutant. Based on growth tests of the heterozygous diploids,
the mutant phenotype of all eight HCR mutants was judged to be recessive.
Complementation Group Analysis
Crossing with the various known mutants such as cia-5, ca-1, pmp-1, and pgp-1 should help
identify new alleles of previously characterized mutants. If any wild-type colonies appear under
low CO2 conditions (50-100 µL L 1 CO2) after mating with HCR mutants, this indicates they are
not allelic to each other, because these known mutants also show HCR phenotypes.
Rapid allelism tests were used to place the various HCR mutants into different complementation
groups. Complementation analysis was tested with the eight HCR mutants (Table II) along with
cia-5, ca-1, pmp-1, and pgp-1 (Table I). Only crosses between cia-5 × HCRP34, cia-5 × HCR209,
and HCRP34 × HCR209 failed to generate wild type colonies. Thus, HCR3510, HCR86, HCR89,
HCR90, HCR95, and HCR105 each define a new HCR locus. HCRP34 and HCR209 have been
confirmed as defective in the same locus as cia-5 by comparison of the sequence of the DNA
flanking the inserts with a cloned cia-5 gene (Xiang et al., 2001 ) and by complementation with a
cloned cia-5 gene (data not shown).
Liquid Growth Experiments
Growth experiments showed patterns of high CO2 requirement for photoautotrophic growth
consistent with those seen in spot tests (Fig. 1). Active, 1-d-old air-adapted cells were inoculated
into liquid minimal medium with similar starting cell densities (5 × 104 cells ml 1), grown with no
aeration, and the cell densities measured daily at the same time of day for 10 d. HCRP34 and
HCR209, judged to be allelic to cia-5, grew very similar to cia-5 in air (Fig. 2A). The growth rates
of HCR86 and HCR90 also were only slightly better than that of cia-5 in air (Fig. 2B), but the
growth rates of HCR89 and HCR95 were intermediate between wild type (ars301; see Table I) and
cia-5 (Fig. 2C). HCR105 was able to grow slightly in air but bleached within a few days (Fig. 2B).
HCR3510, which showed a wild-type phenotype in air on agar, also grew as well as wild type
(ars301) in air in liquid culture (Fig. 2C). Chlorophyll content also was measured in these cultures
along with cell density, and the growth curves based on chlorophyll content showed the same
pattern as those of cell density (data not shown).
Figure 2. Liquid cell growth curves for wild type (ars301), cia-5, and HCR
mutants grown at pH 7 on an orbital shaker without aeration. A, HCRP34 and
HCR209. B, HCR86, HCR90, and HCR105. C, HCR89, HCR95, and
HCR3510. The growth curves shown are averages of three independent
growth
experiments.
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Accumulation of Low CO2-Inducible Transcripts
Because the expression of low CO2-inducible polypeptides (pCA1, mtCA1, mtCA2, Ccp1, and
Ccp2) has been reported to change differentially during acclimation to limiting CO2 (Villarejo et
al., 1996 , 1997 ; Eriksson et al., 1998 ), accumulation of these three transcripts also was
analyzed. The cia-5-like mutants, HCRP34 and HCR209, showed no detectable Cah1 mRNA,
Mca1 and Mca2 mRNA, and Ccp1 and Ccp2 mRNA (Fig. 3A; data shown only for HCRP34).
HCR90, which showed a leaky HCR phenotype in air but a stringent phenotype in low CO2, had
reduced expression of only Mca1 and Mca2 mRNA (Fig. 3B). In separate, long-term experiments,
the expression of the other genes was somewhat variable, but only Mca1 and Mca2 showed
reproducibly decreased mRNA abundance (data not shown). HCR3510, which showed a wildtype phenotype in air but a stringent HCR phenotype in low CO2, had normal expression of these
genes compared with wild type (CC849; see Table I; Fig. 3A). However, HCR95 showed a much
different pattern of expression for these three genes. From cells exposed for 2 h to air, Cah1
mRNA of HCR95 was detected at normal levels, whereas much-reduced levels of Mca1 and Mca2
mRNA and Ccp1 and Ccp2 mRNA were detected relative to wild type (Fig. 3B). After 6 h, wild
type showed the same or increased levels of these three mRNAs, but expression of all three
mRNA in HCR95 was dramatically reduced (Fig. 3B), suggesting only a transient induction of
their expression in this mutant. In separate, long-term experiments, this apparent transient
induction in HCR95 also was confirmed up to 24 h (data not shown). The other HCR mutants
(HCR86, HCR89, and HCR105) did not show reproducibly different patterns of expression for the
three low CO2-inducible transcripts relative to wild type (data not shown).
Figure 3. Northern-blot analyses for wild type (CC849)
and HCR mutants. A, HCRP34 and HCR3510. B, HCR90
and HCR95. Total RNA (10 µg per lane) was isolated 2 h to
6 h after transfer of cells to air levels of CO2 from high
CO2. Cah1 mRNA was probed with the 1.4-kb BglII and
NcoI fragment of Cah1 cDNA (Van and Spalding, 1999 ).
Mca1 and Mca2 mRNA was probed with the full-length
Mca2 cDNA (Eriksson et al., 1996 , 1998 ). Ccp1 and
Ccp2 mRNA was probed with the 1.2-kb EcoRI and HindIII
fragment of Ccp1 G1 (Chen et al., 1997 ). The rRNA was
probed with 25S and 5.8S rDNA (Marco and Rochaix,
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TOP
ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
DISCUSSION
HCR mutants have been useful for investigation of various processes, both in algae and in higher
plants. HCR mutants with defects in several of the enzymes of the photorespiratory pathway have
been isolated in the C3 plants Arabidopsis (Somerville and Ogren, 1982 ) and barley (Hordeum
vulgare) (Joy et al., 1992 ; Leegood et al., 1996 ; Wingler et al., 1999 ). These photorespiratory
mutants exhibited lethality (HCR phenotype) in air levels of CO2 for various reasons, including
accumulation of toxic intermediates during photorespiration and depletion of exchangeable
nitrogen in photorespiratory intermediates. In C. reinhardtii, the photorespiratory mutant pgp-1
(lacks PGPase) has a HCR phenotype, indicating that the oxygenase activity of Rubisco was not
completely suppressed by operation of the CCM and that photorespiratory mutants in
C. reinhardtii also are lethal in air levels of CO2 (Suzuki et al., 1990 ; Spalding, 1998 ).
Mutants defective in functional components of the CCM also exhibit an HCR phenotype in
C. reinhardtii (Spalding et al., 1983a , 1983b ; Moroney et al., 1986 ; Suzuki and Spalding,
1989 ; Funke et al., 1997 ; Karlsson et al., 1998 ) and cyanobacteria (Price and Badger, 1989 ;
Ogawa, 1991 , 1992 ; Marco et al., 1993 ; Ohkawa et al., 1998 ; Price et al., 1998 ). Isolation
and characterization of the C. reinhardtii mutants, ca-1 and pmp-1, demonstrated the requirement
for active transport and accumulation of Ci (Badger et al., 1980 ; Spalding et al., 1983b ) and for
a thylakoid lumen CA (Spalding et al., 1983a ; Funke et al., 1997 ; Karlsson et al., 1998 ) for
function of the CCM. Another C. reinhardtii HCR mutant, cia-5, exhibits no apparent low-CO2
acclimation responses, such as induction of CCM, up-regulation of low CO2-inducible
polypeptides, up-regulation of photorespiratory enzymes, or down-regulation of Rubisco
biosynthesis (Moroney et al., 1989 ; Marek and Spalding, 1991 ; Spalding et al., 1991 ; Burow
et al., 1996 ). This mutant is thought to be defective in the signal transduction pathway for
acclimation to limiting CO2. The gene responsible for this mutation (Cia5) has been cloned
recently (Fukuzawa et al., 2001 ; Xiang et al., 2001 ), and its characterization suggests it may
encode a transcription factor. The identification of this important gene opens the way for more
rapid progress in delineation of the signal transduction pathway for acclimation to limiting CO2.
Because many changes involved in acclimation to limiting CO2 conditions appear to be controlled
at different gene expression levels, it is possible that mutations in several different loci might yield
signal transduction mutants like cia-5 with HCR phenotypes. Thus, the HCR phenotype should be
a good indicator of nonacclimation to low CO2 as well as for a dysfunctional CCM, so isolation of
HCR mutants should be helpful for identification of loci required for either function of the CCM
or for signal transduction leading to low CO2 acclimation.
Among the eight new HCR mutants described here, six represent new complementation groups
and the other two represent new alleles of the previously described cia-5 locus. The patterns of
growth and of low CO2-inducible transcript accumulation for HCRP34 and HCR209 were similar
to those of cia-5, and complementation group analyses confirmed that the three are allelic. As new
alleles of cia-5, HCRP34 and HCR209 may prove valuable in understanding the function of the
gene product from this important locus.
Other than for HCRP34 and HCR209, the growth responses to air and low CO2 varied among
these new HCR mutants, as did the pattern of accumulation of limiting-CO2-inducible genes.
HCR90, which showed a stringent HCR phenotype in low CO2 and grew only slightly better than
cia-5 in air (Fig. 2B), had reproducibly reduced expression of only one pair of the limiting-CO2inducible transcripts, Mca1 and Mca2. No disruption of the structural gene for either Mca1 or
Mca2 was found in genomic Southern blots probed with the Mca1 and Mca2 promoter region
(data not shown), so HCR90 may be defective in a regulatory component that preferentially
affects expression of Mca1 and Mca2. HCR86, which has a growth phenotype very similar to
HCR90, showed limiting-CO2-inducible transcripts accumulations that were not reproducibly
different from those of wild type (data not shown). The leaky phenotype in low CO2 of HCR89
and HCR95 was supported by their growth patterns (Fig. 2C), but only HCR95 reproducibly
showed reduced level of low CO2-inducible transcripts (Fig. 3B).
HCR3510 showed no significant differences from wild type in terms of low CO2-inducible
transcript accumulation, suggesting it is unlikely to be defective in the limiting-CO2-responsive
signal transduction pathway. The growth phenotype of this mutant, near wild-type growth in
normal air but a stringent phenotype in low CO2, suggests a defect in a functional component of
the CCM (or another pathway required for acclimation to limiting CO2) that is essential in very
low CO2 but not in air levels of CO2.
The advantage of using insertional mutagenesis to generate mutants lies in the use of the inserted
DNA as a “tag” to clone the disrupted gene, but of course this only works if the insert
cosegregates with the mutant phenotype, i.e. if the insert is responsible for the mutation. As
judged by the Arg+ phenotype, the Arg7 inserts in mutants HCRP34, HCR3510, HCR86, HCR89,
and HCR90 cosegregate with the HCR phenotype (Table II), suggesting the insert caused the
mutation in each of these strains. As indicated above, both HCRP34 and HCR209 are allelic to
cia-5 and the insert in each has been confirmed to disrupt the cia-5 gene. Thus, we know the defect
in both these mutants, even though cosegregation of the Arg+ and HCR phenotypes has not been
demonstrated for HCR209.
It is unfortunate that the inserts in mutants HCR95 and HCR105 do not cosegregate with the HCR
phenotype, so identification of the disrupted gene responsible for the HCR phenotype in these
mutants will have to be accomplished without the aid of an insertional tag. The three remaining
tagged mutants (HCR3510, HCR86, and HCR 90) remain as viable candidates for identification of
novel genes essential for acclimation of C. reinhardtii to limiting CO2. Cloning of the disrupted
genes in these three HCR mutants is in progress.
TOP
ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
MATERIALS AND METHODS
Cell Strains and Culture Conditions
All Chlamydomonas reinhardtii strains (Table I) were grown as previously described (Geraghty et
al., 1990 ). Cells were cultured on an orbital shaker under aeration with 5% (v/v) CO2 in air (high
CO2-grown cells) or no aeration (air-adapted cells). For experiments monitoring the accumulation
of low CO2 inducible transcripts, cell cultures were switched from aeration with 5% (v/v) CO2 to
aeration with normal air for 2 h to 6 h. For growth on solid media, cells were maintained under
5% (v/v) CO2 in air (high CO2), normal air, or 50 to 100 µL L 1 CO2 (low CO2).
Generation and Isolation of Mutants
Glass bead transformations were performed as described previously (Van and Spalding, 1999 ).
To generate a pool of insertional mutants on CO2-minimal medium, CC425 (Table I) was
transformed with linearized p-Arg7.8 (Debuchy et al., 1989 ) containing the structural gene
(Arg7) for argininosuccinate lyase to complement the arg2 mutation. Each of more than
7,000 colonies was screened by spot tests to identify HCR mutants. After replica plates with
transformants were made, each plate was placed in high CO2 and air or high CO2 and low CO2.
Mutants identified in this primary screen as having HCR phenotypes were screened again by
western immunoblots of extracellular protein to identify mutants in which pCA1 expression was
decreased or absent (Van and Spalding, 1999 ).
Spot Growth Tests and Growth Experiments
For spot growth tests, actively growing cells were suspended to similar cell densities in minimal
medium, spotted (10 µL) onto minimal agar plates, and grown in different concentrations of CO2
for 10 d (Harris, 1989 ).
For liquid growth experiments, active, 1-d-old air-adapted cells were inoculated into liquid
minimal medium at similar cell densities (5 × 104 cells ml 1). The cultures were grown on an
orbital shaker without aeration for the next 10 d. The cell density was determined using a
hemacytometer (Reichert Scientific Instruments, Buffalo, NY; Harris, 1989 ). Chlorophyll content
was estimated after extraction with 96% (v/v) ethanol (Wintermans and De Mots, 1965 ).
DNA- and RNA-Blot Analysis
Southern- and northern-blot analyses were performed as described by Van and Spalding (1999) .
Total RNA was purified with TRIzol reagent (Life Technologies, Gaithersburg, MD) from airinduced cells exposed to limiting CO2 (aeration with normal air) and Hybond N+ nylon transfer
membrane (Amersham Pharmacia Biotech Inc., Piscataway, NJ) was used for blotting. After
phoporimager analysis of each northern blot (Molecular Dynamics, Piscataway, NJ), total RNA
amounts were normalized to hybridization with 25S and 5.8S rRNA (Marco and Rochaix, 1980 )
using ImageQuaNT (Molecular Dynamics).
Genetic Analyses
All matings were performed by crossing insertionally generated mutants with various strains
(Table I) according to the protocol of Harris (1989) . To isolate vegetative diploids, gametes from
HCR mutants (sr-u-2-60) and CC1068 (nr-u-2-1) were induced under nitrogen stress, mated, and
the mating mixture spread onto kanamycin-containing medium to select for expression of the
plastid-encoded kanamycin resistance (nr-u-2-1) transmitted from the mating-type minus parent.
Putative diploids (surviving colonies) were verified by selection for simultaneous expression of
the plastid-encoded streptomycin resistance (sr-u-2-60) from the mating-type plus parent and by
DNA quantity in flow cytometry (performed at the Iowa State University Cell Facility, Ames).
Complementation group analyses required construction of mating type minus strains of each HCR
mutant (both new and previously described mutants). Mating type minus strains of cia-5, ca-1,
pmp-1, and pgp-1 were generated by crossing with CC124 (Table I). CC1068 (Table I) was used
for generating mating type minus strains from all new HCR mutants, except HCR95 and HCR105.
After crossing each of the seven new HCR mutants and the four known mutants with each other,
the progeny from each cross were tested for photoautotrophic growth in low CO2 (50-100 µL L
1
). Because all HCR mutants required elevated CO2 for survival, wild-type colonies were observed
in low CO2 only if the cross generated wild-type recombinant progeny.
FOOTNOTES
Received April 9, 2001; returned for revision June 4, 2001; accepted July 2, 2001.
1
This work was supported by the U.S. Department of Agriculture National Research Initiative
(grant nos. 97-35100-4210 and 99-35100-7569 to M.H.S.). This is journal paper no. J-19297 of
project no. 3578 of the Iowa Agriculture and Home Economics Experiment Station (Ames) and
was supported by the Hatch Act and State of Iowa funds.
•
Corresponding author; e-mail mspaldin@iastate.edu; fax 515-294-1377.
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ABSTRACT
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
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Plant Physiology, January 2004, Vol. 134, pp. 520-527
This Article
10.1104/pp.103.030569
ENVIRONMENTAL STRESS AND ADAPTATION
Respiratory Oxygen Uptake Is Not Decreased by an
Instantaneous Elevation of [CO2], But Is Increased
with Long-Term Growth in the Field at Elevated [CO2]1
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Phillip A. Davey2, Stephen Hunt, Graham J. Hymus3,
Evan H. DeLucia, Bert G. Drake, David F. Karnosky
and Stephen P. Long*
Departments of Crop Sciences and Plant Biology,
University of Illinois, Urbana, Illinois 61801 (P.A.D.,
S.P.L., E.H.D.); Department of Biology, Queen’s
University, Kingston, Ontario, K7L 3N6, Canada (S.H.);
Smithsonian
Environmental
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Edgewater, Maryland 21307 (G.J.H., B.R.D.); and
School of Forest Resources and Environmental Science,
Michigan
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ABSTRACT
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE
CITED
ABSTRACT
Averaged across many previous investigations, doubling the CO2 concentration ([CO2]) has
frequently been reported to cause an instantaneous reduction of leaf dark respiration measured as
CO2 efflux. No known mechanism accounts for this effect, and four recent studies have shown
that the measurement of respiratory CO2 efflux is prone to experimental artifacts that could
account for the reported response. Here, these artifacts are avoided by use of a high-resolution
dual channel oxygen analyzer within an open gas exchange system to measure respiratory O2
uptake in normal air. Leaf O2 uptake was determined in response to instantaneous elevation of
[CO2] in nine contrasting species and to long-term elevation in seven species from four field
experiments. Over six hundred separate measurements of respiration failed to reveal any decrease
in respiratory O2 uptake with an instantaneous increase in [CO2]. Respiration was found
insensitive not only to doubling [CO2], but also to a 5-fold increase and to decrease to zero. Using
a wide range of species and conditions, we confirm earlier reports that inhibition of respiration by
instantaneous elevation of [CO2] is likely an experimental artifact. Instead of the expected
decrease in respiration per unit leaf area in response to long-term growth in the field at elevated
[CO2], there was a significant increase of 11% and 7% on an area and mass basis, respectively,
averaged across all experiments. The findings suggest that leaf dark respiration will increase not
decrease as atmospheric [CO2] rises.
A quantitative analysis of prior studies weighted for replication and experimental variation
concluded that a doubling of atmospheric CO2 concentration would decrease respiratory CO2
efflux by 18% in woody plants (Curtis and Wang, 1998 ). This decrease is considered the result of
a direct instantaneous effect of increased CO2 concentration and a longer term indirect effect due
to changes in leaf composition with long-term growth at elevated CO2 (for review, see Drake et
al., 1999 ). In an analysis of 45 species, 36 showed an average 15% instantaneous reduction in net
respiratory CO2 efflux per unit leaf area (Rd,CO2) on transfer to elevated [CO2] (for review, see
Amthor, 1997 ). Terrestrial plant respiration releases 10 times more carbon per annum than fossil
fuel combustion (Amthor, 1997 ). Therefore, a 15% to 20% decrease in foliar respiration might
increase the potential of terrestrial vegetation to sequester carbon into biomass, providing a partial
amelioration to the rate of increase of atmospheric [CO2] (for review, see Gonzalez-Meler and
Siedow, 1999 ).
However, there is considerable variation in the reported instantaneous effects of elevated [CO2] on
Rd,CO2 with some studies reporting a large decrease and others reporting no change (Bunce, 2002 ;
Bruhn et al., 2002 ). Furthermore, whereas some biochemical reactions within plant respiratory
metabolic pathways are sensitive to [CO2], none of those identified exert sufficient metabolic
control to account for the reported decreases in respiration (Gonzalez-Meler and Siedow, 1999 ).
Four developments now suggest that the decreases observed when measuring Rd,CO2 from leaves
in gas exchange systems incorporating infrared CO2 analyzers may be the result of artifacts in the
measuring system. Amthor (2000 ) showed that very thorough sealing and use of an enlarged
cuvette enclosing the leaf greatly reduced the apparent instantaneous effect of doubling [CO2] on
Rd,CO2, although a small significant effect still remained. Jahnke (2001 ) systematically analyzed
and eliminated potential artifacts within a gas exchange system and subsequently failed to detect
any response in bean (Phaesoleus vulgaris) and poplar (Populus tremula) leaves when corrections
were applied. Bouma et al. (1997 ) and Burton and Pregitzer (2002 ) similarly demonstrated that
respiratory efflux of CO2 from roots was independent of measurement CO2 concentration.
Assuming common dark respiratory metabolism between roots and leaves, this casts further doubt
on the possibility of a direct response in the leaf. Avoiding the potential artifacts of CO2 exchange
measurement by determining dark respiratory O2 uptake (Rd,O2), Amthor et al. (2001 ) showed no
significant change in respiration of five Rumex crispus leaves in sharp contrast to earlier
measurements of Rd,CO2 in the same species (Amthor et al., 1992 ).
Measurement of O2 uptake has three important advantages over measurement of CO2 efflux for an
instantaneous effect of change in [CO2] on respiration. First, the gas being measured is not the gas
being altered in concentration, avoiding any need for instrument recalibration. Second, the
concentration gradient of [O2] between the cuvette enclosing the leaf and the surrounding air is
unaltered when [CO2] is changed. Finally, O2, unlike CO2, is not easily absorbed and adsorbed by
surfaces in the gas exchange system. Yet, even when the effect of elevated [CO2] on O2 uptake has
been examined, findings have been variable. In contrast to Amthor et al. (2001 ), Reuveni et al.
(1993 ) measured a decrease in respiratory O2 uptake under elevated [CO2]. Controversy over the
presence or absence of a direct effect of [CO2] on Rd,CO2 continues. Some recent studies of Rd,CO2,
despite controlling for errors likely to arise from leaks, continue to report a significant
instantaneous decrease in Rd,CO2 on elevation of [CO2]. For example, decreases have been reported
of 19% for rice (Oryza sativa) canopies on elevation of [CO2] from 350 to 700 µmol mol-1 (Baker
et al., 2000 ); 31% to 79% for Scots pine (Pinus sylvestris) needles also on elevation from 370 to
700 µmol mol-1 (Jach and Ceulemans, 2000 ); 16% to 40% for eight species of crop and crop
weeds on increase from 60 to 1,000 µmol mol-1 (Bunce, 2001 ); an 11% average for leaves of 12
grass species on elevation from 360 to 1,300 µmol mol-1 (Tjoelker et al., 2001 ); 18% in the
second leaf of soybean (Glycine max) on elevation from 350 to 1,400 µmol mol-1 (Bunce, 2002 );
and 4.5% decrease for seedling shoots of the tropical rainforest tree Tectona grandis on elevation
from 370 to 600 µmol mol-1 (Holtum and Winter, 2003 ). Precautions taken in measurement were
sufficient for Bunce (2002 ) to rebut the idea that the direct effect of elevated [CO2] on respiration
was an artifact: “These precautions coupled with observed effects of dark [CO2] treatments on
mass accumulation and translocation, suggest that the observed changes in respiration were real.”
Although these decreases are smaller than those suggested by earlier studies, and some use larger
increases in [CO2] to elicit an effect, they continue to suggest direct instantaneous inhibition of
leaf dark respiration by increase in [CO2]. An added uncertainty is whether an instantaneous
suppression of respiration by elevated [CO2] is limited to certain species or conditions (Drake et
al., 1999 ). Although maintaining the same [CO2] outside the cuvette to that inside will minimize
diffusive leaks, it will not eliminate the substantial problem of CO2 absorption/desorption from
surfaces in the gas exchange system (Bloom et al., 1980 ; Long et al., 1996 ; Jahnke, 2001 ) or
the mass flow of CO2 between the cuvette and the outside through the intercellular air space when
only part of the leaf is enclosed (Jahnke and Krewitt, 2002 ). Thus, some of these further artifacts
may still affect even these very careful studies. Clearly, we are some way from any consensus
over the existence, or otherwise, of a direct effect of increase in [CO2] on leaf respiration in the
dark.
If the artifacts suggested by Amthor (2000 ), Jahnke (2001 ), and Jahnke and Krewitt (2002 )
apply to the effect of an instantaneous increase in [CO2] on Rd,CO2, then the same errors would at
least in part apply when these systems are used to measure the effect of long-term acclimation to
elevated [CO2] on Rd,CO2. Plants grown in elevated [CO2] in the long-term (1-10 years) typically
show decreased nitrogen content and increased photosynthesis, growth, leaf mass per unit area,
nonstructural carbohydrate concentrations (Drake et al., 1997 ), and large increases in leaf
mitochondrial numbers (Robertson et al., 1995 ; Griffin et al., 2001 ; Tissue et al., 2002 ). With
the exception of nitrogen content, these are all factors that might be expected to lead to increased
leaf respiration. However, a quantitative survey using meta-analytical analysis concluded that
long-term growth at elevated [CO2] decreased Rd,CO2 significantly and by 18% (Wang and Curtis,
2002 ).
Recently, high-sensitivity dual lead-gold (fuel cell) detectors have been developed that allow
measurement of small differences in [O2] (approximately 1-2 µmol mol-1) between the inlet and
outlet air streams of a leaf cuvette in normal air (Willms et al., 1997 , 1999 ; Hunt, 2003 ). The
objectives of this study were to use such a measurement system to answer two questions: Can the
findings of Amthor et al. (2001 ) of no instantaneous effect of doubling of [CO2] on leaf Rd,O2 be
extended to a wide range of species, growth conditions, and measurement [CO2]? Using a range of
long-term experiments in which a wide range of plants have been grown under elevated [CO2] in
the field (Table I) is Rd,O2 decreased or unaffected as a result of acclimation to elevated [CO2]?
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Table I. Species, [CO2] treatments, plant and leaf developmental stage at the
time of measurement
Locations for material growing in the field and microclimate for controlled
environment grown plants are given under “Materials and Methods”.
References providing a description of each of the long-term field elevated
[CO2] experiments used are included.
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RESULTS
There was no significant instantaneous effect of [CO2] on leaf Rd,O2 in any of the nine species
studied (Fig. 1; Tables II) and III). The results for the ANOVA were, for CO2, F3,332 = 0.032, P
= 0.99; and for species x CO2, F24,332 = 0.528, P = 0.97. A complete lack of any effect of [CO2]
was evident in all species not only on doubling [CO2], but when [CO2] was increased more than
five times above the current ambient concentration or decreased to zero (Fig. 1; Tables II and III).
The extremely high probabilities for accepting the null hypothesis (P > 0.97) eliminate any
possibility of a Type II error, i.e. a difference undetected because of variability. This is also
reflected in the combined regression of response relative to ambient [CO2] (Fig. 1), where the
probability that the response is independent of [CO2] was P = 0.99 (F3,360 < 0.001). This lack of
response of respiration to elevated [CO2] was independent of treatment method (F3,360 = 0.222, P
= 0.88), developmental stage (F3,360 = 0.174, P = 0.92), beginning or end of night (F3,360 =
0.638, P = 0.59), and the [CO2] at which the plants had been grown (F3,100 = 0.080, P = 0.97).
Although there was no interaction with [CO2], absolute rates of respiration generally decreased
with time in the dark.
Figure 1. The instantaneous effect of change in [CO2],
from the current ambient concentration, on leaf respiratory
O2 uptake in the dark. Respiration is plotted as the
percentage of change (±1 SE) relative to the rate of O2
consumption at current ambient [CO2]. Each mean
illustrated is based on measurements of 274 plants and nine
species (Table I). Leaves were in darkness and their
temperature was maintained at 25°C. Results for an analysis
View larger version (13K): of variance (ANOVA) are given in the results text, and
[in
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[in
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Table II. The response of leaf respiration to changes in [CO2] for plants
grown at current [CO2] only.
The mean dark leaf O2 uptake on an area basis (micromoles per minute per
second), and its SE (n = 6), are given for each of the four species at the
beginning and end of the 10-h night.
Table III. The response of leaf respiration to changes in (CO2) for species
grown at long-term elevated (CO2) experiments
The mean respiration rate and its SEM are given on an area basis and a dry weight
basis. n indicates the number of plots or open-top chambers from which
measurements of each treatment were made. Bold values show the critical longterm comparison of leaves grown and measured at their growth concentrations.
An asterisk indicates that leaves grown and measured at elevated (CO2) have
significantly different rates (P < 0.05) to those grown and measured at current
ambient (CO2).
Despite the lack of an instantaneous effect, long-term growth at elevated [CO2] did affect
respiration. Leaves grown and measured at elevated [CO2] had the same or a significantly higher
Rd,O2 than those grown and measured at current ambient [CO2] (Table III). The largest increases in
respiration were for Quercus geminata (23%) and soybean (22%) on a leaf area basis. By contrast,
there was no increase in Acer saccharum, Betula papyrifera, and Populus tremuloides (Table
IIIa). When all values from the long-term experiments were combined, there was a significant
increase in Rd,O2 of 11% and 7% on a leaf area and mass basis, respectively, and relative to
controls (Fig. 2).
Figure 2. The effect of long-term (lifetime or >3 years)
growth at elevated [CO2] in the field on leaf respiration.
The data points show mean leaf respiratory O2 uptake
(±90% confidence intervals) on an area and mass basis for
View larger version (8K): leaves grown at elevated [CO2] relative to controls grown at
[in
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using meta-analytical statistical methods.
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DISCUSSION
Six hundred separate measurements of different leaves, encompassing four long-term field CO2
enrichment experiments revealed no evidence of any decrease in Rd,O2 in response to
instantaneous elevation of [CO2]. This was consistent for seven tree and two crop species, the
majority sampled from long-term elevated [CO2] experiments where plants have developed
throughout their life or for several seasons under elevated [CO2]. This lack of response was
irrespective of developmental stage, growth conditions, and growth [CO2]. Bunce (2001 )
reported that the sensitivity of Rd,CO2 to an instantaneous elevation of [CO2] increased with
duration of the dark period. However, no effect on Rd,O2 was evident here (Tables II and III),
regardless of whether measurements were made at the start or end of the natural dark period.
Jahnke (2001 ) similarly found no effect of [CO2] on Rd,CO2 even after 72 h of darkness. Tjoelker
et al. (2001 ) noted that the instantaneous decrease in Rd,CO2 with elevated [CO2], although minor
(1.8%) in response to increase from 360 to 700 µmol mol-1, was far more substantial (11%) on
increase from 360 to 1,300 µmol mol-1. However, a 5-fold increase above current ambient [CO2]
or removal of CO2 from the atmosphere around the leaf did not alter Rd,O2 (Fig. 1). These findings
support the conclusions of Amthor et al. (2001 ) and of Jahnke (2001 ) and extend them to more
species and to major long-term [CO2] field manipulation studies. Although the results show no
effect of instantaneous elevation of [CO2] on Rd,O2, this cannot rule out some effect on apparent
Rd,CO2 due to non-respiratory processes, for example, CO2 uptake into carboxylic acids. However,
in one of the very few studies to measure CO2 and O2 exchange at different [CO2], Amthor et al.
(2001 ) found no effect of elevated [CO2] on respiratory quotient.
It has been suggested that the variability of response to instantaneous increase in [CO2] apparent
in the literature may be the result of species differences (Drake et al., 1999 ; Baker et al., 2000 ;
Hamilton et al., 2001 ). Without definitive testing, it is impossible to be certain on this point.
However, this study contains a diverse range of species and functional types (including C3 and C4
photosynthetic types, herbaceous and woody forms, two major crops, and seven forest dominant
gymnosperm and angiosperm species) that individual exceptions exist now seem somewhat
improbable. Moreover, we deliberately selected species previously reported to show some of the
largest inhibitions of respiration by elevated CO2 with previous methods and technology: soybean
40% (Bunce, 1995 ), Pinus taeda 14% (Teskey, 1995 ), Quercus rubra 5.6% (Amthor, 2000 ),
and maize (Zea mays) 46% (Cornic and Jarvis, 1972 ).
Any errors associated with measuring the instantaneous response of respiratory CO2 efflux to
elevated [CO2] may be equally applicable to measurement of the long-term effects of growth at
elevated [CO2] on Rd,CO2. The significant 7% increase in Rd,O2 on a mass basis, averaged across
seven species from four long-term experiments (Fig. 2), contrasts sharply with the average 18%
decrease reported by Wang and Curtis (2002 ) in their meta-analysis of Rd,CO2 on a mass basis
measured in long-term experiments. The significant increase on a mass, as well as an area basis,
suggests that increased respiration per unit leaf area is not just the result of the increased mass per
unit area that is commonly observed in leaves grown at elevated [CO2] (Drake et al., 1999 ).
However, as noted above, this mean increase masks marked differences between species in this
long-term response (Table III).
To our knowledge, this is the first study to use this alternative method to investigate the effects of
long-term growth at elevated [CO2] on respiration. The increase in respiration shown here is
consistent with the widely reported increase in leaf-soluble carbohydrates with long-term growth
at elevated [CO2] (Moore et al., 1999 ). Azcon-Bieto and Osmond (1983 ) showed that increase
in leaf nonstructural carbohydrate content produced by manipulation of photosynthesis during the
photoperiod increased subsequent dark respiration rates. Therefore, a similar response might be
expected when elevated [CO2] increases photosynthesis and leaf nonstructural carbohydrate
content during the photoperiod.
In conclusion, instantaneous elevation of [CO2] had no effect on leaf O2 uptake in these nine
species. This further confirms the suggestion that reports of respiratory inhibition, based on
measurements of CO2 uptake, are the result of experimental artifacts and not the result of any
sensitivity of plant respiration to the [CO2] at the time of measurement. A 15% to 20% reduction
of terrestrial plant respiration with a doubling of atmospheric [CO2] concentration would represent
some amelioration, at least temporarily, in the rate of rise in the global atmospheric [CO2] (for
review, see Gonzalez-Meler and Siedow, 1999 ). It can no longer be assumed that the direct effect
of elevated [CO2] on plant respiration will reduce future ecosystem CO2 efflux. On the basis of the
long-term field manipulations of [CO2] studied here, a small increase in respiration per unit leaf
area should be expected—this will amplify the effect of any increase in leaf area due to growth at
elevated [CO2].
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MATERIALS AND METHODS
LITERATURE
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MATERIALS AND METHODS
Growth Conditions
The species, developmental stage, growth conditions, and experimental site are listed in Table I.
Seven out of nine species were grown at current ambient (368 µmol mol-1) and at an elevated
[CO2] (550 or 700 µmol mol-1). Two of the species were measured at more than one
developmental stage; soybean (Glycine max) at vegetative and podfill stages, and Pinus taeda at
juvenile and mature stages. Soybean was grown in controlled environment- and open-field
conditions (FACE).
Oxygen Analysis System
Leaf O2 uptake rate was determined in an open gas exchange system incorporating a dual-cell
oxygen analyzer (S-3A/DOX; AEI Technologies, Pittsburgh) described by Willms et al. (1997 ),
and incorporating a stainless steel and glass leaf gas exchange cuvette (MPH-1000; Campbell
Scientific, Logan, UT). The analyzer incorporates a novel differential oxygen sensor obtained by
modifying two electrochemical fuel cells (model KE-25; Figaro USA, Wilmette, IL), placing one
in a reference gas stream and the other in an analytical gas stream, and then ensuring that the two
gas sensors were exposed to an identical temperature and pressure (Willms et al., 1997 ). The O2
fuel cells contain a Teflon membrane through which O2 can diffuse, and a weak acid electrolyte
bathes a lead anode and a gold cathode electrode. In the environment of the sensor, the O2 oxidizes
the lead, forming PbO, which dissolves into the solution and generates a small current. The
difference in output of the two sensors is directly proportional to the [O2] between the air streams.
The selective diffusive properties of Teflon and the specific electrochemistry ensure that the
differential sensor responds directly only to O2 in air. However, changes in partial pressures of
CO2 and water vapor due to respiration and transpiration will affect that of O2 causing an indirect
effect (Willms et al., 1999 ). For this reason, these gases are removed from the air stream before
the O2 sensors (Hunt, 2003 ). The analyzer was secured to a vibrationless table top and additional
layers of insulation were wound around the O2 measurement cells and instrument casing to further
improve resolution. The differential resolution of the modified dual-cell O2 analyzer was 1 to 2
µmol mol-1 against a background [O2] of 210 mmol mol-1. The cuvette was large enough to
enclose the entire leaf, leaflet, or cohort of needles; typically, 50 to 120 cm2 of leaf were enclosed.
This avoided errors associated with gas flow between the cuvette and surrounding air via the
intercellular air space that Jahnke and Krewitt (2002 ) have shown to result from the use of
cuvettes that only enclose part of the leaf. The cuvette (MPH-1000; Campbell Scientific) has been
described in detail previously (Bingham et al., 1980 ), adapted from the earlier design of Bingham
and Coyne (1977 ). Typical boundary layer conductances to water vapor are 2.5 to 4 mol m-2 s-1
(calculated from Bingham and Coyne, 1977 ; Bingham et al., 1980 ). A gas exchange system (LI6400; LI-COR, Lincoln, NE) was used to provide a controlled [CO2] to the cuvette, and air flow
rate was measured by a pair of flow meters (F350; AEI Technologies), calibrated as described
previously (Long et al., 1996 ). The gas flow across the leaf was maintained at 2.5 cm3 s-1. This
flow rate and the enclosed leaf area resulted in a [O2] of 40 to 80 µmol mol-1. The entry air
streams to the oxygen analyzer were scrubbed of water and CO2 by magnesium perchlorate and
soda lime. Column sizes were calculated to remove all CO2 and to be sufficient to dry the air
stream to the equilibrium vapor pressure of magnesium perchlorate (approximately 0.1 Pa). The
efficacy of these columns was tested by switching dry air (<0.01 kPa water vapor) with a
humidified air stream (2.0 kPa water vapor) at the cuvette inlet; no change in [O2] from 0 µmol
mol-1 was recorded. Similar tests showed that the soda lime column was effective in preventing
interference from differences in [CO2] between the cuvette and reference air streams.
The response time of the gas exchange system was assessed using two cylinders of compressed air
found to differ in [O2] by 80 µmol mol-1. Air from one of these cylinders was fed to the empty
cuvette inlet and also the reference air stream until a constant [O2] of 0 µmol mol-1 was obtained
at the analyzer. Air from the second cylinder was then switched in at the cuvette inlet and the time
taken to obtain a constant [O2] of 80 µmol mol-1 at the analyzer was recorded. The change was
98% complete within 10 min and complete within the resolution of the analyzer at 15 min.
Therefore, it was assumed that 20 min was adequate to record any response of respiratory O2
efflux to an instantaneous change in [CO2].
Respiration Measurements
Respiration was measured in the last 2.5 h of the night in all species and additional measurements
were made in the first 2.5 h of the night for three species (Table I). Measurements were made at
each location listed in Table I. However, because of its size and stability, the gas exchange system
was housed in a field laboratory at each site and leaves were detached shortly before measurement.
Petioles were cut under water and their cut ends were kept immersed in water until measurements
were complete. The leaves remained turgid throughout. For pot-grown soybean (Table I), parallel
measurements on attached and detached leaves were made; no significant differences in
respiration or its response to [CO2] were found. Leaves were equilibrated in the dark to 25°C
before measurement. For a single replicate measurement, respiration was measured at a [CO2] of
2,000 µmol mol-1, and then again at 700, 550, 360, and 0 µmol mol-1 CO2. To avoid systematic
error, each alternate replicate measurement was started at 0 µmol mol-1 CO2 and was then
switched to 368, 700, 550, and 2,000 µmol mol-1 CO2. Species from the FACE experiments at
Duke Forest, Rhinelander, and the University of Illinois, and the open-top chamber experiment at
Merritt Island were measured at 550 µmol mol-1 [CO2] rather than at 700 µmol mol-1 [CO2]
because this was the growth concentration (Table I). After each step-wise change in [CO2], a 20min waiting period ensured adequate time for any instantaneous response in Rd,O2. Each set of
measurements took 80 min to complete, and leaves were maintained in the dark at a leaf
temperature of 25°C for the entire measurement period. After each set of measurements, leaf area
was measured by digital imaging and the leaves were then dried to a constant weight at 80°C. Leaf
O2 evolution rates were calculated on a leaf area and a dry mass basis by adapting the CO2 uptake
equations of von Caemmerer and Farquhar (1981 ) for open system gas exchange measurements.
Statistical Analysis
In the four long-term field experiments, replicate number was determined by the number of
treatment plots (Table III). This avoided pseudoreplication. A minimum of three repeat
measurements were taken in each treatment plot of each experiment, and their pooled values
provided the individual sample measure. Only these data were used to assess the effect of longterm growth at elevated [CO2] on respiratory O2 uptake. Outside of these long-term elevated [CO2]
studies, measurements were made from six randomly selected plants each of P. taeda seedlings,
Q. rubra mature trees, and maize flowering plants growing under current ambient [CO2] in the
open. Soybean was grown in two separate controlled environment studies, and here, replicates
represent the number of separate chambers used (Table I).
ANOVA was used to test the effect of immediate changes of [CO2] on leaf respiration rate for all
nine species combined (SYSTAT, Evanston, IL). This tested the effect of species, growth
conditions, developmental stage, time of measurement, and growth [CO2] on the instantaneous
response to elevated [CO2]. In addition, a regression analysis of response relative to ambient
[CO2] was tested (SYSTAT).
Because of the very different experimental designs, durations, and treatment procedures of the
experiments in which plants were grown at elevated [CO2], and the possibility that species may
respond differently to long-term growth in elevated [CO2], the pooled effect of all field studies
was assessed by meta-analytical statistical techniques (Hedges et al., 1999 ; Ainsworth et al.,
2002 ; Morgan et al., 2003 ) using a software package (MetaWin, Rosenberg et al., 2000). This
allows determination of the consensus of studies differing in design. The natural log of the
response ratio (respiratory O2 uptake for leaves grown at elevated [CO2]/respiratory O2 uptake for
leaves grown at the current ambient [CO2]) was used as the metric for analysis (Hedges et al.,
1999 ). A mixed-model analysis was used based on the assumption of random variation in effect
sizes between individual measurements. A weighted parametric analysis was used (Gurevitch and
Hedges, 1999 ), and each individual observation was weighted by the reciprocal of the mixedmodel variance (Hedges et al., 1999 ). If the response ratio was not significantly different from
unity, growth at elevated [CO2] failed to change leaf O2 uptake from that at current ambient [CO2].
However, if significantly different from 1, growth at elevated [CO2] on average altered O2 uptake
across the studies.
ACKNOWLEDGMENTS
We thank Henry Ginsberg and Joseph Veltre of AEI Technologies, and Nicholas Dowling of
Qubit Systems for technical help and modifications to the O2 analysis system; Jeff Amthor,
Miquel Gonzalez-Meler, Andrew Leakey, and Shawna Naidu for comments on draft versions of
the manuscript; Carlos Pimentel and Kate George for the supply of plant material; and Lisa
Ainsworth for advice on the meta-analysis.
Received July 20, 2003; returned for revision August 20, 2003; accepted September 25, 2003.
FOOTNOTES
1
This work was supported by the U.S. Department of Energy’s Office of Science (to B.E.R.), as
were the long-term elevated [CO2] experiments in Florida, North Carolina, and Wisconsin.
2
Present address: Department of Biological Sciences, University of Essex, Colchester, CO4 3SQ,
UK.
3
Present address: Dipartimento di Scienze dell’ Ambiente Forestale e delle Sue Risorse,
Università della Tuscia, Via S. Camillo de Lellis, 01100 Viterbo, Italy.
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Corresponding author; e-mail stevel@life.uiuc.edu; fax 217-244-7563.
TOP
ABSTRACT
RESULTS
DISCUSSION
MATERIALS AND METHODS
LITERATURE CITED
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