Artisanal Multiplication of Entomopathogenic Nematodes
in Ecuador for the control of Phyllophaga sp.
Francisco Báez, Patricio Gallegos, César Asaquibay, Marcía Oña
NATIONAL AUTONOMOUS INSTITUTE FOR AGRICULTURAL RESEARCH (INIAP) - ECUADOR
ABSTRACT
In Ecuador, entomogenous nematodes have considerable potential in managing soil pests, especially Phyllophaga sp. in
5. The test was maintained for 14 days after inoculation with EPN.
perennial crops such as blackberry (Rubus glaucus). Wide adoption of beneficial nematodes by farmers is currently
6. Larval mortality and root weight of each blackberry plant was evaluated.
challenged by the need for economically feasible, local, artisanal mass production of efficacious nematode
7. Mortality of larvae and weight of root blackberry plants were analyzed using LSD FISHER comparison method (InfoStat
species/strains.
I/S Version 2010).
Nine strains of Steinernema and Heterorhabditis nematodes from the lNlAP collection, were evaluated greenhouse tests
B. ARTISANAL MULTIPLICATION OF EPN
using blackberry plants infested with Phyllophaga sp. The best nematode was Heterorhabditis bacteriophora strain
1. Compost beds, 2.5 m long and 1.25 m wide, were established with wooden boards to retain the material.
H-04D. It caused 70% larvae mortality and increased production of blackberry roots (10.8 gram roots per plant versus 2.7
2. Compost was a mix of local materials: one part organic matter (droppings of small animals, chickens, or cattle) with
three parts soil). All organic material used in this study was completely decomposed.
grams in the untreated controls).
Artisanal production of this nematode used compost, into which 100 larva Phyllophaga sp. from farmers’ fields, were
3. Introduction of 100 Phyllophaga sp. larvae and 15 G. mellonela larvae, infected with EPN.
placed. Then 15 larval Galleria mellonela larvae infected with the nematode were added. Infective juveniles (IJs), were
4. Compost was protected with mulch to prevent sunlight, low temperatures, and to maintain humidity.
found up to 90 days after start of the test production. The nematode population that can be obtained by this method
5. Compost was moistened weekly to keep moisture close to water field capacity.
depends on the number of larvae of Phyllophaga sp. used at the beginning of the preparation of the compost. One larva
6. Subsequent, satellite production could be made by mixing 15 kg of composted material infested with EPN (or 15
can produce up to 32,000 IJs. In addition, the farmer will be able to establish composting and multiply more nematodes
infected Phyllophaga larvae) with the contents of the new compost along with 100 healthy larvae of Phyllophaga sp.
from the inoculum available. Soil and compost from the nematode can be used in crops for the control of pest
RESULTS AND CONCLUSIONS
mentioned.
INTRODUCTION
Larval Phyllophaga sp. are important in perennial Ecuadorean crops such as blackberry (Rubus glaucus). Blackberry is a
highly attractive fruit for the Ecuadorean, and due to its low production area, it is not exported in large volumes but
consumed domestically. The Department of Plant Protection of lNlAP, reported a reduction in the root area of the plant,
with fewer and shorter branching, causing decreased fruit yield. (INlAP 2012).
In Ecuador, the limiting factor in wide-scale use of beneficial nematodes (EPN) is the
difficulty of mass production and subsequent distribution to farmers.
Currently,
these nematodes are produced in the laboratory, by methods difficult for
Figure 1. Mortality of larvae of Phyllophaga sp. after exposure to nine
strains of nematodes. Error bars represent S.E.M.
Figure 2. Weight of root blackberry plants infected with larvae of
Phyllophaga sp. Error bars represent S.E.M.
Ecuadorean farmers. The method requires extraction of nematodes from larvae of
A. SELECTION OF THE BEST STRAIN NEMATODE
Galleria mellonella by White trapping (Castillo et al., 2012). Subsequently, these
We determined the best strain to be Heterorhabditis bacteriophora HO4-D in the greenhouse tests. It resulted in 70% larval
nematodes are distributed for application to control larvae of soil, especially
mortality under the condition of the test (Figure 1) and significantly increased blackberry plant root mass (10.8 grams )
Phyllophaga sp. A basic artisanal production method is needed if nematodes are to
compared with the control treatment (2.7 grams per plant) (Figure 2).
have greater adoption.
.
B. ARTISANAL MULTIPLICATION OF EPN
Blackberry plant root affected by
Phyllphoga sp.
Prior to development of artisanal multiplication, an efficacious nematode strain had
to be identified through efficacy tests evaluating 9 EPN from lNlAP for the control of
larval Phyllophaga sp. These trials were conducted in blackberry plants through
deliberate infestations of the pest. A compost based artisanal production system was
then devised and evaluated with the most efficacious EPN.
Establishment of the compost bed
Nematode-infected Phyllphoga sp. larva
(left) and healthy larva (right)
Preparation of compost by manual mixing
Placement of EPN-infected larval Phyllophaga
sp. and G. mellonella in the compost.
Artisanal production of EPN is possible using Phyllophaga sp. larvae, in compost. On average, the mortality of the larvae in
METHODOLOGY
compost was 78.3% and the population of IJs / larva reached a maximum of 32,000 individuals, as determined by the
A. SELECTION OF THE BEST ENTOMOPATHOGENIC NEMATODE STRAIN
laboratory extraction method (White trap). At the end of the study, on day 90 after the trial was begun, the floor of the
1. Efficacy tests were conducted with 2-month-old blackberry plants placed in 1.5 L plastic pots.
compost was sampled and we found an average density of 25 IJs / gram of soil. The source of organic material for the
2. Field collected five larval Phyllophaga sp. collected from field (1.0 to 2.0 g each) were placed in each pot and
production of compost can be chicken (Gallus gallus domesticus), guinea pig (Cavia porcellus) and rabbit (Oryctolagus
maintained for 7 days.
3. For the test were used two plastic pots per treatment more a untreated control.
4. The pots were inoculated with one of 9 EPN nine strains (Table 1), at a ratio of ten infective juveniles (IJs) per gram of
soil. (The IJs used in this study were extracted from larvae of Phyllophaga sp., previously inoculated in the laboratory).
Table 1. EPN evaluated in the selection of the best entomopathogenic nematodes strain.
Collection Region
Strain Designation
Species
Chimborazo – La Delicia
H04 - D
Heterorhabditis bacteriophora
Tungurahua – Huachi
2-TH
Heterorhabditis bacteriophora
Carchi – Monteverde
CB-13
Steinernema feltiae
Chimborazo – Guayllabamba
H01-G
Steinernema feltiae
Cotopaxi – Chambapongo
CT-08
Steinernema feltiae
Carchi – Chután Bajo
CH-06
Steinernema feltiae
Cotopaxi – Pataín Norte
CT-13
Heterorhabditis bacteriophora
Chimborazo – Rayoloma
H03-R
Steinernema feltiae
Cotopaxi – Chanchaló
CT-06
Steinernema feltiae
cuniculus), all of which are commonly reared on Ecuadorean farms. The compost method can be easily adopted by Ecuadorean
farmers.
REFERENCES
Castillo, C.; Gallegos, P.; Asaquibay, C.; Oña, M. eds. 2012. Guìa de prospección y producción de nematodos
entomopatógenos. INIAP, EESC, Departamento Nacional de Protección Vegetal. Quito, Ecuador. Manual Técnico Nº 88. 15 p.
INIAP (Instituto Nacional Autónomo de Investigaciones Agropecuarias). Departamento Nacional de Protección Vegetal.
2012. Informe Anual 2012. Estudio de la acción de los insectos de suelo en la marchitez descendente de la mora (Rubus
glaucus) en la provincia de Tungurahua. Cutuglagua, Ecuador.
Lemma Ebssa.; Eugene M. Fuzy.; Matthew W. Bickerton.; Albrecht M. Koppenhöfer. 2012. Host density effects on
efficacy of entomopathogenic nematodes against white grub (Coleoptera: Scarabaeidae) species, Biocontrol Science and
Technology, 22:1, 117-123, DOI: 10.1080/09583157.2011.643767
Ricci, M.; Glazer, I.; Campebll, J.F.; Gaugler, R. 1996. Comparison of bioassays to measure virulence of different
entomopathogenic nematodes, Biocontrol Science and Technology, 6:2, 235 – 246 DOI: 10.1080/09583159650039421
CONTACT
Francisco Báez: francisco.baez@iniap.gob.ec, Patricio Gallegos: patricio.gallegos@iniap.gob.ec