COMPARATIVE GROWTH AND PROPAGULE VIABILITY OF LOUISIANAHARVESTED BLACK MANGROVE, AVICENNIA GERMINANS
A Thesis
Submitted to the Graduate Faculty
of Nicholls State University
in Partial Fulfillment
of the Requirements for the Degree
Master of Science in Marine and Environmental Biology
By
William Finney
Bachelor of Science, Renewable and Sustainable Resources, College of Applied Life
Science at University of Louisiana at Lafayette, 2002
Fall 2011
CERTIFICATE
This is to certify that the thesis entitled “Comparative Growth and Propagule
Viability of Louisiana-harvested Black Mangrove, Avicenia germinans” submitted for
the award of Master of Science to the Nicholls State University is a record of authentic,
original research conducted by Mr. William Finney under our supervision and guidance
and that no part of this thesis has been submitted for the award of any other degree
diploma, fellowship, or other similar titles.
APPROVED:
SIGNATURE:
DATE:
Quenton Fontenot, Ph.D.
Associate Professor of Biological
Sciences
Committee Chair
___________________________
____________
Allyse Ferrara, Ph.D.
Associate Professor of Biological
Sciences
Committee Member
___________________________
____________
Dr. Gary LaFleur
Associate Professor of Biological
Sciences
Committee Member
___________________________
____________
Mr. Gary Fine
Research Associate and
Plant Materials Specialist
Committee Member
___________________________
____________
1
ACKNOWLEDGEMENTS
I would like to thank the following people and organizations who made this research
possible:
Dr. Quenton Fontenot
Mr. Gary Fine
Dr. Allyse Ferrara
Dr. Gary LaFleur
Louisiana Native Plant Initiative USDA NRCS
Coalition to Restore Coastal Louisiana (Funding)
Nicholls State University
Jefferson Parish, Louisiana
Barataria-Terrebonne National Estuary Program
Ms. Shelley Sparks
Mr. Christopher Finney
Fellow students: Tim Clay, Sara Shields, Cynthia Fox, and Justin Merrifield
ii
ABSTRACT
As the integrity of coastal salt marsh in Louisiana has degraded, efforts to rebuild
land have intensified. Native vegetation or erosion resistant materials must be used to
stabilize land rebuilt by natural sediment deposition or by relocation of dredge material.
Black mangrove, Avicennia germinans, is a species native to Louisiana that is ideal for
coastal stabilization because of its high salt-tolerance and resistance to wave energy. The
commercial production of black mangrove in Louisiana is currently limited by costs
related to the plant’s lack of both frost tolerance and seed dormancy. In an effort to
increase black mangrove propagation efficiency, seed viability and vigor were compared
among harvest date and pericarp removal methods.
Two thousand eight hundred black mangrove seeds were harvested in October,
November, and December 2009 from the Louisiana coast. Seeds from the October
harvest were soaked for 48 hours in water of different salinities to remove pericarps (seed
coat). Subsequent harvests were soaked in freshwater, while pericarps were peeled by
hand from some seeds of each harvest while still dry. From the soaked treatments, the
floating and sinking seeds were separated and planted as different treatments. All seeds
were weighed, measured and planted three days after harvest. Seed viability and vigor
were recorded and compared to buoyancy, method of pericarp removal, and time of
harvest.
Data indicate no relationship between salinity of soak water or buoyancy to
viability or vigor; however harvest date may influence both viability and vigor. Seeds
harvested in October and November germinated more quickly than the December harvest,
although there was no difference in total germination. Peeled seeds also germinated more
slowly than soaked seeds for all harvests, but with no difference in total germination.
Seeds harvested in October and November had similar growth, and both grew more
vigorously than the December harvest.
iii
TABLE OF CONTENTS
CERTIFICATE .................................................................................................................... i
ACKNOWLEDGEMENTS................................................................................................ ii
ABSTRACT....................................................................................................................... iii
TABLE OF CONTENTS................................................................................................... iv
LIST OF TABLES...............................................................................................................v
LIST OF FIGURES ........................................................................................................... vi
LIST OF TERMS ABBREVIATIONS ............................................................................ vii
INTRODUCTION ...............................................................................................................1
METHODS ..........................................................................................................................9
RESULTS ..........................................................................................................................15
DISCUSSION ....................................................................................................................24
CONCLUSION..................................................................................................................29
LITERATURE CITED ......................................................................................................30
BIOGRAPHICAL SKETCH .............................................................................................34
iv
LIST OF TABLES
Table 1. Number of replicates (10 seeds per replicate) for each treatment combination for
the October harvest…………………………………………………………….14
Table 2. Mean (±SE) seed weight (g) and length (mm) for seeds from the October
harvest. ..............................................................................................................16
Table 3. Mean (±SE) seed length (mm) and weight (g) for floaters (0 ppt), sinkers
(0 ppt), and peeled seeds for October, November, and December harvests. ......19
v
LIST OF FIGURES
Figure 1.
Global distribution Avicennia germinans and distribution in Louisiana ........2
Figure 2.
Basic post-germination anatomy of an Avicennia germinans propagule.......4
Figure 3.
Mean daily greenhouse temperature (°C) recorded by constant temperature
readers located in the center, and outside raceways for the duration of the
growout period. ..............................................................................................11
Figure 4.
Cumulative percent germination through 48 DSH for floaters, sinkers and
peeled treatments for black mangrove seeds harvested 1 October 2009…… 17
Figure 5.
Mean (±SE) percent germination per treatment by 28 DSH for the 1 October
2009 harvest. ................................................................................................18
Figure 6.
Cumulative percent germination for early, middle and late season harvest
seed through 48 DSH .....................................................................................20
Figure 7.
Mean (±SE) percent germination of black mangrove seeds 48 days since
harvest on 1 October 2009, 1 November 2009, or 1 December 2009………21
Figure 8.
Mean (± SE) percent germination by 15 March, 2010 for seeds harvested 1
October 2009, 1 November 2009, or 1 December 2009. …………………. 22
Figure 9.
Mean (± SE) height for early, middle, and late harvests on March 15, 2010
for floaters, sinkers, and peeled seeds…..………………………………….23
vi
LIST OF TERMS AND ABBREVIATIONS
Anoxic = a condition of very low dissolved oxygen in soil or water, below 0.5 mg/L
Cotyledon = a seed leaf which provides stored energy for embryonic growth until true
leaves are formed on the epicotyl.
Crypto-viviparous seed. = propagules that germinate within the pericarp of a seed, often
while still attached to the parent plant
DSH = Days since harvest
Epicotyl = the phototropic, upward-projecting embryonic stem of a mangrove, from
which true leaves emerge
Geotropic = growing in response to gravity; gravitropism
Germination = the embryonic growth of a plant. In this study, germination is defined by
the exposure of the epicotyl beyond the cotyldeon
Germplasm = plant tissues from which new plants can be grown
Halophyte = a plant adapted to live in soil and water with high salt concentrations.
Hypocotyl = the geotropic stem extension between the root radical and the cotyledon
Isotherm = geographic boundary or contour line demarcating contiguous regions of
common climatic temperatures
ppt = parts per thousand
Pericarp = fleshy outer covering of the black mangrove propagule; a seed coat
Propagule = the viviparous and recalcitrant seed of the black mangrove
Radical = embryonic root of a plant, the root tip of the hypocotyl
Recalcitrant seed = seed that cannot survive typical storage of dehydration or freezing
Spartina alterniflora, S. patens = Salt marsh cordgrasses native to Louisiana that often
share coastal habitat with black mangrove, Avicennia germinans.
Xylem embolism = the formation of an air bubble in the xylem of a vascular plant that
interrupts evapo-transpirative uptake of water and minerals.
vii
INTRODUCTION
Mangroves are an association of approximately 69 species of tropical woody
halophytes from at least 16 families (Lewis 2000; Kathiregan and Bingham 2001;
Lacerda 2001). Mangrove propagules are tidally dispersed and have developed
physiological mechanisms to tolerate inundation, anoxic soils, and varying degrees of
salinity (Lewis et al. 2005; Duke et al. 1998). Some of these adaptations render
mangroves vulnerable to freezing temperatures (Lugo and Snedaker 1974; Duke et al.
1998), and are therefore restricted to tropical and subtropical intertidal zones
approximately 30º north and south of the equator (Figure 1), where they form unique
global marine habitats (Tomlinson 1986).
Red mangrove Rhizophora mangle, white mangrove Laguncularia racemosa, and
black mangrove Avicennia germinans are the three mangrove species native to the
southeastern United States (Tomlinson 1986). However, only the black mangrove occurs
naturally outside of Florida (McMillan and Sherrod, 1986). The northern-most population
of black mangrove in the United States is restricted to the salt marshes of the Mississippi
River delta between Atchafalaya Bay and the mouth of the Mississippi River in Louisiana
(Patterson et al. 1993; Visser et al. 1998). Spartina alterniflora and S. patens marshes
coincide with the Louisiana population of black mangroves (Patterson et al. 1993; Visser
et al. 1998), which may limit the distribution of mangrove propagules (Kathiregan and
Bingham 2001).
1
Figure 1. Global distribution of Avicennia germinans and distribution in Louisiana
(inset). Arrow indicates harvest site at Fourchon, Louisiana (29o06’50.00”N,
90o11’04.74”W). Distributional limiting isotherms are demarcated by 28o north and
south latitudinal meridians (isotherm = 20o Celsius; Duke et al. 1998).
2
Black mangroves produce recalcitrant seeds, dispersed by tidal currents
(Tomlinson 1986). Despite a reliance on buoyancy for dispersal, some seeds do not float
(McKee 1995). Seeds of black mangrove are crypto-viviparous, and do not have a
dormant stage resulting in an unorthodox germination possible while still attached to the
parent plant (Tomlinson 1986; Farnsworth 2000). Black mangrove seeds possess a
hydrophilic pericarp, which once seed is abscised, is shed upon contact with water
(Tomlinson 1986). The seeds are dispersed tidally and may remain viable while afloat in
agitated seawater for up to one year, although viability decreases over time (USDA
2009). Typical black mangrove germination initiates when a propagule comes to rest on
a suitable substrate such as a tidal mud flat, beach, or within a Spartina spp. marsh
(Lewis 2000). This epigeal germination is first observed with the extension of a geotropic
root radical into the soil where the seed comes to rest. The hypocotyl extends to become
a vertical stalk supporting the cotyledon (Tomlinson 1986). The propagule then extends
its epicotyl, from which the first true leaves emerge, allowing the plant to
photosynthesize independent of the seed (Figure 2). The cotyledons then desiccate, and
abscise (Tomlinson 1986).
As facultative halophytes, mangroves do not require salt to survive, but are
tolerant of prolonged exposure to saline environments up to 48 ppt (Alleman and Hester
2010), where they can out compete other woody species (Hoff 2002). The black
mangrove’s large xylem vasculature allows the uptake of saltwater, which creates an
increased vulnerability to freeze induced xylem embolism (Stuart et al. 2006; Salisbury
and Ross 1992).
3
Figure 2. Basic post-germination anatomy of an Avicennia germinans propagule.
4
Frost intolerance prevents mangroves from surviving far beyond the tropics
(Stuart et al. 2006). In Louisiana, black mangroves occupy the northern-most limit of
their temperature requirements beyond the limiting isotherm (20o Celsius; Duke1998).
Prolonged or frequent exposure to freezing temperatures can kill a stand of black
mangrove (Turner and Lewis 1997). Even the mild winters in Louisiana cause stunted
growth in the northern-most population (Visser et al. 1998). In the tropics, the black
mangrove is a substantial tree and can grow to 20 m in height (Lugo 1974). In a
Louisiana Forestry Commission Bulletin, Brown (1945) described black mangroves near
the mouth of the Mississippi River reaching approximately 7 m tall. However, the largest
extant trees observed during this research were less than 4 m in height at Port Fourchon,
Louisiana. Fluctuations in distribution and individual plant size likely occur relative to
periodic freeze events (Tomlinson 1986).
Inundation and anoxic soils are also stressors to the black mangrove (Hoff 2002).
Prolonged inundation can stress black mangroves to the point of mortality, while
drainage may produce dry conditions that allow competitive species to overtake the
habitat (Turner and Lewis 1997). Black mangrove roots survive inundation and anoxic
soils by producing lenticels-covered pneumatophores that extend above the soil and water
surfaces and conduct atmospheric oxygen to the underground root system through (Lugo
and Snedaker 1974; Tomlinson 1986). The pneumatophores also trap detritus, enabling
the cycling of nutrients (Feller et al. 1999) and in doing so, create a vital nursery habitat
for many fish and marine invertebrates (Duke et al. 1998; Lacerda et al. 2001).
Mangrove habitats in south Louisiana are currently vulnerable to subsidence and
sea level rise (Maul 1993; Reed 1995). As the integrity of coastal salt marsh in Louisiana
5
degrades, black mangrove habitat is threatened (Reed 1995). The construction of roads
and navigation canals within the coastal marshes has altered hydrology, and effectively
blocked the tidal influence of the Gulf of Mexico in mangrove habitats (Lugo and
Snedaker 1974). As a result, mangrove stands may be permanently inundated or drained;
neither of which are conducive to survival and recruitment (Turner and Lewis 1997).
As public awareness of land loss and economic impacts related to hurricanes has
increased, efforts to restore coastal habitats have intensified (Reed 1995). Numerous
governmental and non-profit organizations are attempting to restore sedimentary
processes lost by the channelization of the Mississippi River (LCWCRTF 2006; Reed
1995). The cost of coastal restoration projects is often a limiting factor, therefore natural
restorative processes are most desirable (Reed 1995; Lewis 2000). Millions of dollars are
spent each year in Louisiana to build land in degraded coastal wetlands (Reed 1995;
Neumann et al. 2000). Land built by natural sediment deposition or by relocation of
dredge material must be stabilized with vegetation or erosion resistant materials (Reed
1995; Lewis 2000).
The most efficient means of sustainable shoreline stabilization is often the
establishment of native vegetation (USDA 2009). As dredged soils and sediment are
relocated to counteract subsidence and sea level rise, vegetation must be established to
hold these new lands (Lewis 2000). However, the variety and availability of native salt
tolerant vegetation is often limited, and a combination of characteristics should be
considered in species selection (Lewis 1998). The resilience of mangroves in the coastal
environment makes them an ideal species for wetland and barrier island restoration (Hoff
2002; Lewis 2000). Woody structure and salt tolerance make black mangrove one of the
6
most effective buffers against storm surge among Louisiana’s native coastal vegetation
(Patterson et al. 1993).
Coastal restoration projects often require mangroves for revegetation purposes
(Lewis 2000), but the wholesale cultivation of mangroves in Louisiana may be limited by
costs related to the plant’s lack of seed dormancy and cold tolerance (USDA 2009). The
majority of black mangrove cultivated for restoration purposes in the United States is of
Florida germplasm (Lewis 2000), but the limitations of tidal dispersal and isolation of the
mangrove colonies in Louisiana suggest that plants used for restoration in this most
northern population be of its native germplasm.
Historically, Louisiana nurseries specializing in coastal vegetation have avoided
black mangrove propagation due to difficulties associated with the harvest, transport and
storage of viviparous seed. For example, greenhouses are needed to prevent losses due to
freezing. Therefore, methods that may increase the availability of Louisiana germplasm
A. germinans include identifying seed viability, indicators to increase yield, and
characterization of growth patterns that may allow for the elimination of seed storage and
increased efficiency of germination in northern climates.
In Louisiana, black mangroves flower from May to August and fruit from late
August through December. The recalcitrant seeds are vulnerable to desiccation and
cannot be reliably stored for long periods by chilling or drying (Duke et al. 1998).
Commercial cultivation of this species in Louisiana requires that seeds be harvested in
the fall and immediately planted in a greenhouse, where temperatures must be maintained
above freezing. The lack of consistent harvest techniques among cultivators has
7
historically resulted in low yields of black mangrove when attempted (USDA-NRCS
2009).
The goal of this research was to identify the optimal time of season to harvest
seed and the methods of seed treatment that resulted in the highest germination rate and
most vigorous seedling growth to improve the commercial harvest and propagation
efficiency of Avicennia germinans using native Louisiana germplasm. Specific
objectives of this study were to:
1. Compare germination rate of seeds that had pericarps removed by hand or had
pericarps removed by soaking seeds for 48 hrs in a 0, 5, 10, 15, or 20 ppt saline
solution,
2. Compare the germination rate of seeds harvested early (October 1), mid
(November 1), or late (December 1) in the harvest season that either had their
pericarps removed by hand or by soaking seeds for 48 hrs in freshwater.
3. Compare the size of seeds collected either early (October 1), mid (November 1),
or late (December 1) in the harvest season,
4. Compare the seedling height 48 days after seed harvest for seeds harvested early
(October 1), mid (November 1), or late (December 1) in the harvest season that
had pericarps removed by hand or by soaking seeds for 48 hrs in freshwater, and
5. Compare the seedling height on the 15 March following seed harvest for seeds
harvested early (October 1), mid (November 1), or late (December 1) in the
harvest season that had pericarps removed by hand or had pericarps removed by
soaking seeds for 48 hrs in freshwater.
8
METHODS
Seed Collection
Black mangrove seeds were harvested from mature black mangroves near Port
Fourchon, LA (Figure 1) on 1 October 2009, 1 November 2009, and 1 December 2009.
The seeds were transported to Nicholls State University in dry buckets and were
maintained indoors at room temperature for one day. The day of harvest was assigned
day zero for all time-delineated treatments.
Seed Preparation
Seeds undergoing water-removal of pericarps were soaked in shaded outdoor
tanks and hand-peeled seeds were stored indoors in dry buckets (18.9L) indoors until
peeling, to initiate all treatments one day after harvest. Soaked seeds were placed in one
of five 150 L tanks of 0, 5, 10, 15, or 20 ppt salinity. Instant Ocean® (Spectrum Brands
Inc. Madison Wisconsin) sea salt formula was used for all salinity treatments. Seeds
were placed into each of the solutions on the first day after harvest (day 1), to remove
pericarps and establish buoyancy. Seed buoyancy was determined after 48 hours, as
“floaters” or “sinkers”. Dry seeds, were peeled of their pericarps by hand on the day that
the soaked seeds were removed from their salt solutions and were categorized as
“peeled.” The three pericarp removal treatments will be referred to as ‘floaters’,
‘sinkers’ and ‘peeled’.
9
Seed Planting
Prior to planting, individual cells (“cells” 6.4 cm x 25cm: D40-L Deepot
Conetainer® Stuewe & Sons Inc. Tangent, OR) were filled with Pro-Mix BX® (Premier
Tech Horticulture Ltd. Quebec, Canada) planting medium (sphagnum moss, Vermiculite
mix). Cells were placed in raceway tables in a greenhouse at the Nicholls State
University farm. The depth of raceways was maintained to submerge the lower half of
each cell, allowing the planting medium to saturate with water prior to planting, thus
simulating natural germination conditions (Alleman and Hester 2010). Three Hobo®
constant temperature readers (Onset Computer Corp. Cape Cod, MA) were placed among
raceways in the center and both end tanks and were used to record water temperature at
30 minute intervals for the duration of the experiment. Thermostatically controlled
heaters and exhaust fans were set to maintain air temperatures between 15 and 37 ºC
(Figure 3; Kao et al. 2004).
After the pericarps were removed, the seeds were weighed (g), measured (mm)
along the longest axis, and then placed on top of the planting medium. One seed was
placed in each cell, similarly oriented, standing vertically to facilitate equal growth of the
radicle into the medium. Treatments were arranged in rows of ten seeds, and distributed
randomly among the raceway tables. Each row of ten cells represented an experimental
unit replicate for each treatment combination.
10
1-Oct
28-Oct
25-Nov
22-Dec
19-Jan
15-Feb
15-Mar
Figure 3. Mean daily greenhouse temperature (°C) recorded by constant
temperature readers located in the center, and outside raceways for the duration of
the grow-out period.
11
Germination
Black mangrove produce crypto-viviparous propagules, therefore germination
was recorded as the day post-harvest when epicotyl’s true leaves were visible beyond the
cotyledon. Propagule viability was recorded daily for the first 128 days for each trial, so
that cumulative germination over time could be compared among treatments.
Optimal Pericarp Removal Method
Analysis of variance (ANOVA) was used to compare percent germination through
day 28 among the salinity pretreatments of the 1 October harvest to determine
pretreatment methods for harvests on 1 November and 1 December. Percent germination
was calculated for each row (replicate), and then the percent germination for each
treatment was calculated as the mean of each row. The number of sinkers was limited for
all salinities, resulting in fewer replicates than that of floaters, therefore the design was
not balanced (Table 1). However, there were a minimum of four replicates for each
sinker treatment.
Effect of Harvest Date on Germination
Percent germination was not different between buoyancies of varied soak
salinities on day 28 of the 1 October harvest, therefore only 0 ppt salinity soaked seeds
were used to compare treatments for all harvests (Figure 4). To determine the effect of
harvest date on germination, percent germination was compared among harvest dates and
12
treatments, either peeled, floater (0 ppt), and sinker (0 ppt) seeds. There were 36
replicates for the 0 ppt October floaters, 4 replicates for the 0 ppt October sinkers and 24
replicates for the October peeled (Table 1). The November and December harvest used
10 replicates for each treatment combination. ANOVA was used to compare mean
percent germination on day 48 and on 15 March among all treatments. 15 March was
used to compare percent germination and seedling size among all treatments based on the
date that would allow for earliest field planting of frost-sensitive black mangroves on the
Louisiana coast (Salisbury and Ross 1992; Koss 1988; NOAA 2005).
Seedling Vigor
Seedling vigor was quantified by seedling height, measured as stem length (mm)
from the soil surface to the terminal bud. Mean height of plants for each harvest date was
measured at a minimum of three intervals during the growing period. Seedlings from the
October harvest were measured at 62, 75, and 165 days since harvested (DSH). Seedlings
from the November harvest were measured at 43, 70, 92 and 133 DSH and seedlings
from the December harvest were measured at 79, 104, and 126 DSH. Seeds that had not
germinated were not included in the seedling vigor analyses.
13
Table 1. Number of replicates (10 seeds per replicate) of each pericarp removal method
(soaked, peeled) treatment combinations and resulting buoyancy in varying soak
salinities for the October harvest of black mangrove seeds.
Treatment
Soaked
Peeled
0ppt
5ppt
10ppt
15ppt
20ppt
Floaters
36
35
33
33
34
-
Sinkers
4
5
7
4
4
-
Peeled
-
-
-
-
-
24
14
RESULTS
Optimal Pericarp Removal Method for Germination
The mean weight and length of seeds among the treatments varied slightly (Table
2). However, the largest and smallest seeds were sinkers and there was no evident seed
size trend among treatments. Percent germination was not linear over time and became
asymptotic by approximately 40 days post-harvest (Figure 4). By day 28, the percent
germination for the peeled treatment was among the lowest rates (Figures 4 and 5).
Germination rates for the floaters and sinkers soaked in 0 ppt treatments were not
different from other salinities; therefore, subsequent germination trials only used 0 ppt
soak salinity and hand-peeling for pericarp removal (Figure 4).
Effect of Harvest Date on Germination
Mean seed size varied among treatments and harvest dates, but there were no
obvious trends among pericarp removal treatment, soak salinity, or harvest date (Table 3)
Germination rate was not linear for seeds harvested in October, September, or December
(Figure 6). Seeds harvested in October germinated faster than seeds harvested in
December (Figure 7; ANOVA). However, by 15 March, there was very little difference
for percent germination among treatments (Figure 8; ANOVA).
Seedling Vigor
The mean stem length of seedlings harvested in October and November was not different
among treatments by 15 March, but all treatments from the December harvest had smaller
seedlings on that date (Figure 9).
15
Table 2. Mean (±SE) seed weight (g) and length (mm) for seeds from the October
harvest. Peeled seeds had pericarps removed without soaking in water. The floaters and
sinkers had pericarps removed by soaking in 0, 5, 10, 15, or 20 ppt saline solution. Means
that share a letter are similar.
Treatment
`
0 ppt
5 ppt
10 ppt
15 ppt
20 ppt
Peeled
Floaters
Weight
Sinkers
Peeled
3.0 ± 0.08 ab
2.9 ± 0.37 ab
-
N=36
N=4
3.3 ± 0.23 ab
2.8 ± 0.15 ab
N=35
N=5
3.1 ± 0.10 ab
3.5 ± 0.72a
N=33
N=7
2.9 ± 0.09 ab
2.2 ± 0.22 b
N=33
N=4
2.8 ± 0.11 ab
2.6 ± 0.24 ab
N=34
N=4
-
-
3.2 ± 0.07 ab
N=24
0 ppt
5 ppt
10 ppt
15 ppt
20 ppt
Peeled
28.9 ± 0.28
Length
29.1 ± 1.33 a
a
N=36
N=4
29.9 ± 0.28 a
29.4 ± 0.55 a
N=35
N=5
28.4 ± 0.37 a
28.7 ± 0.49 a
N=33
N=7
27.6 ± 0.38 a
24.7 ± 0.89 b
N=33
N=4
28.7 ± 0.29 a
27.3 ± 1.02 ab
N=34
N=4
-
-
29.4 ± 0.22 a
N=24
16
Percent Germination
Days Since Harvest
Figure 4. Cumulative percent germination through 48 DSH for floaters, sinkers and
peeled treatments for black mangrove seeds harvested on 1 October 2009. Sinkers
exposed to 0 ppt are represented by the large dashed black line, floaters exposed to 0 ppt
are represented by the small dashed black line, and the peeled seeds are represented by
the solid black line. All other soak salinity and buoyancy treatment combinations are
represented by the small gray lines. The vertical line indicates day 28 post harvest.
17
Percent Germination
A
AB
ABC ABC ABC
AB
ABC
ABC
ABC
BC
C
F0
F5
F10
F15
F20
S0
S5
S10
S15
S20
P
Treatment
Figure 5. Mean (±SE) percent germination per treatment by 28 DSH for the 1 October
2009 harvest. F=floaters, S=sinkers, and P=peeled. The number following either F or S
represents the salinity used to remove the pericarp. Means that share a letter (ABC) are
not different.
18
Table 3. Mean (±SE) seed length (mm) and weight (g) for floaters (0 ppt), sinkers (0
ppt), and peeled seeds for October, November, and December harvests. Means that share
a letter are similar.
Treatment
October
Harvest Date
November
December
Length (mm)
Floaters
Sinkers
Peeled
Floaters
Sinkers
Peeled
28.6 ± 0.28 ab
30.2 ± 0.29 a
27.4 ± 0.42 b
N=10
N=10
N=10
28.4 ± 1.33 ab
28.9 ± 0.34 ab
28.9 ± 0.50 ab
N=10
N=10
N=10
30.2 ± 0.29 ab
28.6 ± 0.29 ab
29.7 ± 0.31 a
N=10
N=10
N=10
Weight (g)
3.3 ± 0.09 a
2.4 ± 0.13 b
N=10
N=10
N=10
2.9 ± 0.37 ab
3.0 ± 0.09 ab
2.9 ± 0.12 ab
N=10
N=10
N=10
3.2 ± 0.07 a
3.0 ± 0.07 ab
3.2 ± 0.11 a
N=10
N=10
N=10
ab
3.0 ± 0.08
19
Figure 6. Cumulative percent germination for October, November and December
season harvested seed through 48-DSH. All soak treatments were 0 ppt. October harvest
is represented by black lines, November harvest treatments are light grey lines and
December harvest treatments are the dark grey lines. Dry peeled seeds are short dashes,
floaters are solid lines, and sinkers are long dashes.
20
Percent Germination
A
A
A
A
B
C
BC
B
C
Oct
Nov
Dec
Harvest Date
Figure 7. Mean (±SE) percent germination of black mangrove seeds 48 days since
harvest on 1 October 2009, 1 November 2009, or 1 December 2009. Floaters are black
bars, sinkers are gray bars, and peeled seeds are white bars. Means that share a common
letter are similar across all harvest dates.
21
Harvest Date
Figure 8. Mean (± SE) percent germination by 15 March, 2010 for seeds harvested 1
October 2009, 1 November 2009, or 1 December 2009. Floaters are represented by black
bars, sinkers are grey bars and peeled seeds are white bars. Means that share a letter are
similar across all harvest dates.
22
200
(mm)
HeigHeight
ht (mm)
A
A
AA
160
A
A
A
A
AA
A
A
B
120
B
B
B
B
80
40
0
Octt
Oc
Nov
Nov
Harvest Date
Dec
Dec
Harvest Date
Figure 9. Mean (± SE) height for October, November, and December harvests on 15
March, 2010 Floaters are light grey bars, sinkers are dark grey bars, and peeled seeds are
black bars.
23
Discussion
The worldwide fragmentation of mangrove populations is likely the result of the
division of contiguous ranges by continental drift or fluctuations in global temperature
(Dodd et al. 2000), which may account for the occurrence of Avicennia germinans on
three continents and two oceans (Figure 1; Duke et al. 1998). Kao et al. (2003) found
that growth of Avicennia marina in Thailand was inhibited below 15 ºC, and McMillan
(1971) found that A. germinans populations in the northern Gulf of Mexico would not
root below 15 ºC. This temperature coincides with the average low water temperature at
the global extremes of mangrove distribution. The isolated population in the northern
Gulf of Mexico appears to be delineated by the limitations associated with cryptoviviparous seed distribution by tidal currents and the regional temperatures.
Despite the presence of suitable substrate and intertidal habitat, black mangrove
does not occur on the Gulf coast of Florida north of Cedar Key (29.10º N), where the
average January water temperature is 14º C (NOAA 2005). In Texas, black mangrove
distribution is limited to south of Freeport (28.89º N), where average water temperatures
are below 15º C for three months of the year. However, at Grand Isle, Louisiana
(29.29º N), the effluent of the Mississippi River and Gulf currents maintain average
temperatures above 16 ºC year round (NOAA 2005), which has allowed the
establishment an “island population” of dwarfed black mangroves in the Barataria and
Terrebonne estuaries associated with the floodplain of Bayou Lafourche.
Once a major distributary of the Mississippi River, Bayou Lafourche (French for
‘the fork’) could not be distinguished from the main channel during high water by
24
boatmen upriver as recently as 300 years ago (Butler 1934). Bayou Lafourche was
dammed and cut off from the main channel in 1903 to prevent flooding, resulting in
stagnation and saltwater intrusion. Beginning in the 1950’s, a siphon pump reintroduced
only 5.7 m3/s of river water to the channel, which remains today. Levees built along the
main channel of the Mississippi during the 19th century have prevented the flooding of
the Barataria Estuary for over a century (Snedden et al. 2007). This loss of periodic
flooding of the Mississippi and its distributaries has resulted in the subsidence and
erosion of 90-115 km² of estuarine wetlands annually since 1956 (Reed 1995). The
conversion of coastal wetlands to open water significantly increases the inland impact of
storm surge associated with tropical storm systems and coastal flooding from sea level
rise (Reed 1995).
In his 1960 “Monograph of Avicennia”, Moldenke challenged a colleague who
suggested that mangroves lacked economic value by proposing that members of
Avicennia are of tremendous importance to humans and the economy based on the
historic and increasing trend of human development of coastal areas in tropical and
subtropical climates. This statement was made unaware of the future demand for
vegetated wetlands in response to the expansive wetland loss associated with the deltalobe abandonment of the Mississippi River. The Barataria-Terrebonne National Estuary
Program (BTNEP), on behalf of a 1995 panel of coastal scientists, listed as the primary
objective of coastal restoration efforts in Louisiana, “the creation and enhancement of
vegetated wetlands” (Reed 1995).
Despite the usefulness of black mangrove seedlings for coastal restoration in
Louisiana, commercial sources of Louisiana germplasm seedlings remain scarce. Low
25
survival of field plantings has discouraged many organizations from pursuing the
inclusion of black mangroves in restoration efforts. Avicennia germinans and other
mangrove cultivars have long been grown for restoration purposes in Florida and
elsewhere globally (Lewis 2000), but the work of McMillan and Sherrod (1986) and
Markley et al. (1982) suggests that black mangrove demonstrate an increased frost
tolerance at northern latitudes. This would indicate that seedlings of Florida stock may
be unsuitable for restoration efforts in Louisiana, resulting in decreased survival during
cold winters and the potential dilution of frost tolerant genotypes in durations of warmer
winter trends.
Seed size is commonly used as an indicator of seed viability and vigor for many
plant species (Murali 1997), but the relationship is highly varied and therefore typically
abandoned as reliable outside of anecdotal agricultural applications (Stanton 1984). Seed
size and buoyancy have been examined in mangrove species as indicators of seedling
performance, but primarily as factors influencing seed dispersal (Rabinowitz 1978a,
1978b, 1978c). The studies of Rabinowitz (1978) suggest that the buoyancy and seed
size of Avicennia and other mangrove species may determine the zonation of seed
deposition but make no indication of viability and vigor. While seed size varied slightly
among harvests and trials, there was no difference in germination rate among seed size in
the trials reported here. Therefore, the selection of seed during commercial harvest of
black mangroves need not differentiate by size.
Rabinowitz (1978a) states that once abscised from the parent plant, black
mangrove propagules must undergo an obligate dispersal period, during which most are
buoyant until stranded on the tideline. Alleman and Hester (2011) investigated the
26
duration of buoyancy and its effect on viability in Louisiana black mangroves and found
that viability decreased in propagules that lost buoyancy after 45 days.
Average salinity in the mangrove habitats of south Louisiana is typically about 8
ppt but can vary from 5 to 20 ppt (NOAA 2006) during the fall, when black mangroves
are releasing seed. Also, salinity can vary widely as hurricanes and flood events may
introduce large volumes of seawater or freshwater to the coastal estuaries. The studies of
Rabinowitz (1978a,b,c) and McKee (1995) indicate that soak salinity may affect the
duration of viability during dispersal. However, results of this study indicate no
meaningful relationship between buoyancy and salinity of soak water to viability or
vigor, although these trials do suggest that harvest date may influence viability and vigor.
Time of harvest is commonly used in horticulture and agriculture to predict seed
viability and crop yield, observing the relationship between maturity of seed and viability
(Basra 1995). However, correlations between early and late harvests are not consistent
among species. Late harvest and subsequent seed maturity have been shown to positively
influence seed quality and vigor in soybeans Glycine max (TeKrony et al. 1984) as well
as other cultivated crops and trees (Basra 1995), while maximum viability and vigor
occurs earlier in seed maturation for tomatoes Solanum lycopersicum (Demir and Ellis
1992). Relationships between propagule maturity and viability among viviparous
mangrove genera are poorly defined (Farnsworth 2000)
Seeds harvested in October and November germinated sooner after harvest than
the December harvest, but had no difference in total germination by 15 March. Seeds
harvested in October and November had grown to a similar size by 15 March, and both
27
grew more vigorously than the December harvest. Day length and temperature likely
influence late season harvested seeds germination and growth rates. Further study is
warranted to determine the relationship between day length, temperature and propagule
performance. Peeled seeds germinated later than soaked seeds for all harvests, but with
no difference in total germination. Mean peeled seed stem length was nominally greater
for all harvests by 15 March.
Agitation of soak water at varied salinities has been observed to delay
germination in Avicennia germinans (McMillan, 1971) when it was assumed that
stabilization initiated germination. The delay of germination displayed in peeled seeds
for all harvests may indicate some other stimulus besides immobilization as an initiator of
germination in this species such as seed moisture, pericarp abscission, and seed
abscission. The potential for delayed germination of crypto-viviparous seed is
justification for further investigation.
28
CONCLUSION
This research has determined that salt is not necessary in the soaking process of
pericarp removal, enabling the commercial grower to defray the cost of salt until salt
hardening of plants is required for field planting. Earlier season harvest results in quicker
germination and subsequently larger plants at the time of earliest field planting, but will
require longer maintenance of seedling growing conditions. Later harvests result in
smaller plants by frost date, but less greenhouse time with no difference in germination
viability. Germination can be delayed for a short time by hand peeling the pericarps from
dry seeds with equal or greater germination and growth rates, but is more labor intensive
than soaked pericarp removal.
In consideration of these findings, it is recommended that the commercial grower
collect and pot seeds in October and November if intended field-planting is within one
year of harvest or if a greenhouse is available. This technique enables propagules to
achieve robust growth before freezing temperatures are likely and that seedlings are tall
enough to withstand field conditions. If field-planting after one year or if no greenhouse
is available, growers may consider harvesting late and or peeling pericarps by hand while
still dry. This most labor intensive method can allow propagules to avoid the longest
periods of freezing temperatures and reduce the duration of artificial heating required.
29
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33
BIOGRAPHICAL SKETCH
On a cold rainy night, in the hinterlands of Orleans Parish, shortly after the new year of
1977, William “Billy” Finney began his life. He grew up wrangling the fauna of the
Mississippi River batture, and headed out for New Mexico after high school. There he
took odd ranch jobs and picked up the banjo. He returned to Louisiana periodically often
enough to complete a B.S. in Renewable and Sustainable Resources from the University
of Louisiana at Lafayette. After college, Billy again headed out west, this time to pursue
a career as a National Park Ranger. Heeding the call of Hurricane Katrina, he returned
home to the swamp to help the U.S. National Park Service interpret the nature the
disaster. Then, while paddling in the Atchafalaya Basin on his birthday, Billy met Dr.
Gary LaFleur of Thibodaux, who encouraged him to enroll in the graduate program of the
Department of Biological Sciences at Nicholls State University. Upon completion of his
master’s degree, Billy will continue his work with the U.S. National Park Service as a
biologist.
34