Interactions of a Rhabditis sp. on the virulence of Heterorhabditis and Steinernema in Puerto Rico.
Article Type: Report
Subject: Nematoda (Research)
Authors: Garcia, Jose Miguel
Jenkins, David A.
Chavarria, Jose A.
Shapiro-Ilan, David I.
Goenaga, Ricardo
Pub Date: 09/01/2011
Publication: Name: Florida Entomologist Publisher: Florida Entomological Society Audience: Academic Format: Magazine/Journal Subject: Biological sciences Copyright: COPYRIGHT 2011 Florida Entomological Society ISSN: 0015-4040
Issue: Date: Sept, 2011 Source Volume: 94 Source Issue: 3
Topic: Event Code: 310 Science & research
Geographic: Geographic Scope: Puerto Rico Geographic Code: 1U0PR Puerto Rico
Accession Number: 298614273
Full Text: Entomopathogenic nematodes (EPN) have been widely used to control pests in the soil, but the efficacy of EPNs can be affected by a variety of factors, among these, competitive interactions with other nematode species, e.g., free-living bactivorous nematodes (Duncan et al. 2003). Surveys conducted separately by D. A. J. and J. M. G. indicated that a free-living nematode, Rhabditis sp., was commonly found in agricultural soils of Puerto Rico and emerged from cadavers of insects, indicating it might be an entomopathogen or an opportunistic invader (Jenkins, unpub.). Therefore, we tested the hypothesis that the native nematode would impact the efficacy of EPN's used to control soil pests in Puerto Rico, either by augmenting the efficacy of the EPNs or by reducing their efficacy through competition. In addition to affecting the mortality caused by the EPNs, the Rhabditis sp. could potentially affect the rate at which target organisms were killed by the EPNs.

The bioassay methods used were based on those described by Shapiro & McCoy (2000a, b) and Jenkins et al. (2007). Nematodes and insects were obtained through soil surveys or laboratory cultures. Soil samples were taken from 5 citrus orchards in the University of Puerto Rico Agricultural Experiment Stations at Isabela and Adjuntas with soil probes sterilized with 10% bleach solution. Each sample was divided into five containers, 400 grams of soil (wet weight) per container and five larvae of Galleria mellonella L. (Lepidoptera: Pyralidae) were placed into each container. Soil was moistened to field capacity. After 7 days the larvae were extracted and evaluated for mortality. Dead larvae were placed in White traps (Kaya & Stock 1997) to collect, quantify and identify any nematodes coming from the cadavers. These nematodes were then placed on G. mellonella larvae to demonstrate infection and thereby satisfy Koch's postulates. EPNs were not found in any of the soil samples collected, but most of the soil samples did yield an unidentified Rhabditis sp. Nematodes were stored in tap water at 13[degrees]C. Cultures of Heterorhabditis bacteriophora Oswego strain, and Steinernema glaseri NJ93 strain were obtained from the laboratory of D. S. I. and subsequently reared in G. mellonella in Puerto Rico. Diaprepes abbreviatus larvae were obtained from Florida Division of Plant Industry.

To assess whether nematode interactions were additive, antagonistic or synergistic we used the analyses outlined in Nishimatsu and Jackson (1998). The nature of the interaction was determined by comparing expected percentage mortality of D. abbreviatus to observed mortality, adjusted using Abbott's formula (Abbott 1925), and subjecting these comparisons to [chi square] tests. Expected mortality was derived from the formula [P.sub.E] = [P.sub.O] + (1 - [P.sub.O])([P.sub.1]) + (1 - [P.sub.O])(1 - [P.sub.1])([P.sub.2]), where [P.sub.E] is the expected mortality, [P.sub.O] is the control mortality, [P.sub.1] is the mortality from 1 nematode sp. treatment alone, and [P.sub.2] is the mortality from the other nematode sp. applied alone. The [chi square] value was derived from the formula [chi square] = [([L.sub.O] - [L.sub.E]).sup.2]/[L.sub.E] + [([D.sub.O] - [D.sub.E]).sup.2]/[D.sub.E], where [L.sub.O] is the number of living larvae observed, [D.sub.O] is the number of dead larvae observed, and [D.sub.E] is the number of dead larvae expected (Nishimatsu and Jackson 1998; SAS 2003). Interactions were deemed additive if the [chi square] value was less than 3.84, antagonistic if the [chi square] value was greater than 3.84 and the observed mortality from the combination was less than the expected mortality from the combination, and synergistic if the [chi square] value was greater than 3.84 and the observed mortality from the combination was greater than the expected mortality from the combination.

There was no difference between the expected and observed mortalities when the nematodes were combined. No interaction was observed between Rhabditis sp. and the EPNs at the doses we assayed, indicating that the native and common Rhabditis sp. is not likely to interfere with applications of the EPNs we assayed (Table 1). Heterorhabditis bacteriophora and S. glaseri caused higher mortality of D. abbreviatus than either the control or the Rhabditis sp. treatment. At the doses tested there was no difference between the mortality caused by Rhabditis sp. and the control treatment. However, in the second trial, 10 days after treatment the combination of S. glaseri and Rhabditis sp. caused higher mortality than the control; whereas neither the S. glaseri treatment nor the Rhabditis treatment applied alone was different from the control. This may indicate some potential for enhanced suppression of the target pest when the 2 nematodes are in the soil simultaneously. On the other hand, when applied in conjunction with S. glaseri, which is visibly different in size from the Rhabditis sp., infective juveniles of Rhabditis sp. were observed emerging from cadavers, indicating that the 2 species may compete for resources within the host.

SUMMARY

A Rhabditis sp. of nematode was collected from soil and then assayed for virulence to last instar larvae ofDiaprepes abbreviatus, both alone and in conjunction with the entomopathogenic nematode species, Steinernema glaseri and Heterorhabditis bacteriophora. When Rhabditis sp. was applied at low doses (10 infective juveniles per [cm.sup.2]), we did not detect a significant difference between the mortality in the Rhabditis sp. treatment and the control treatment. However, cadavers from soil that had been treated with the Rhabditis sp. yielded Rhabditis sp. nematodes, indicating that it is an opportunistic invader of cadavers. When applied with either S. glaseri or H. bacteriophora at low doses, Rhabditis sp. nematodes had no detectable impact on the virulence of the other nematodes, either in total mortality caused or in the speed of the mortality. Because S. glaseri is so much larger than the Rhabditis sp., it was easy to distinguish which nematode emerged from cadavers exposed to both, and in several cases Rhabditis sp. nematodes were detected emerging from cadavers, demonstrating its opportunistic nature.

REFERENCES CITED

ABBOTT, W. S. 1925. A method of computing the effectiveness of an insecticide. J. Econ. Entomol. 18: 265-267.

DUNCAN, L. W, GRAHAM, J. H., DUNN, D. C., ZELLERS, J., MCCOY, C. W., AND NGUYEN, K. 2003. Incidence of endemic entomopathogenic nematodes following application of Steinernema riobrave for control of Diaprepes abbreviatus. J. Nematol. 35: 178-186.

JENKINS, D. A., SHAPIRO-ILAN, D., AND GOENAGA, R. 2007. Virulence of entomopathogenic nematodes against Diaprepes abbreviatus in an Oxisol. Florida Entomol. 90: 401-403

KAYA, H. K., AND STOCK, S. P. 1997. Techniques in insect nematology In L. A. Lacey [ed.], Manual of Techniques in Insect Pathology. Academic, San Diego, CA. 281-324.

NISHIMATSU, T., AND JACKSON, J. J. 1998. Interaction of insecticides, entomopathogenic nematodes, and larvae of the western corn rootworm (Coleoptera: Chrysomelidae). J. Econ. Entomol. 91: 410-418.

SAS. 2003. Version 9.1. SAS Institute, Cary, North Carolina.

SHAPIRO, D. I., AND MCCOY, C. W. 2000a. Susceptibility of Diaprepes abbreviatus (Coleoptera: Curculionidae) larvae to different rates of entomopathogenic nematodes in the greenhouse. Florida Entomol. 83: 1-9.

SHAPIRO, D. I., AND MCCOY, C. W. 2000b. Virulence of entomopathogenic nematodes to Diaprepes abbreviatus (Coleoptera: Curculionidae) in the laboratory. J. Econ. Entomol. 93: 1090-1095.

JOSE MIGUEL GARCIA (1), DAVID A. JENKINS (2), JOSE A. CHAVARRIA (3), DAVID I. SHAPIRO-ILAN (4), AND RICARDO GOENAGA (2)

(1) Instituto de Investigaciones Agropecuarias y Forestales (IDIAF) Centro de Tecnologias Agricolas, Santo Domingo, Republica Dominicana

(2) USDA-ARS, Tropical Agriculture Research Station, 2200 Ave. P.A. Campos, Mayaguez, PR 00680-5470

(3) University of Puerto Rico, Department of Environmental Sciences, PR 00680

(4) USDA-ARS-Southeastern Fruit and Tree Nut Laboratory, Byron, GA 31008
TABLE 1. MEAN ([+ OR -] SEM) PROPORTION OF DIAPREPES ABBREVIATUS
LARVAE SURVIVING AFTER INOCULATION WITH VARIOUS NEMATODE SPECIES AND
COMBINATIONS. MEANS FOLLOWED BY THE SAME LETTER WITHIN A COLUMN WERE
NOT DETERMINED TO BE SIGNIFICANTLY DIFFERENT.

                                                 Mean proportion
                                             surviving [+ or -] SEM
                                              Days post inoculation

                                                        2
Trial 1
  Control                                     1.00 [+ or -] 0.00 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.92 [+ or -] 0.02 a
  H. bacteriophora (10 IJs/[cm.sup.2])        0.82 [+ or -] 0.02 a
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.82 [+ or -] 0.05 a
  S. glaseri (10 IJs/[cm.sup.2])              0.84 [+ or -] 0.08 a
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.90 [+ or -] 0.05 a

Trial 2
  Control                                     0.88 [+ or -] 0.05 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.96 [+ or -] 0.03 a
  H. bacteriophora (10 IJs/ [cm.sup.2])       0.94 [+ or -] 0.03 a
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.94 [+ or -] 0.04 a
  S. glaseri (10 IJs/[cm.sup.2])              0.94 [+ or -] 0.04 a
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.96 [+ or -] 0.03 a

                                                 Mean proportion
                                             surviving [+ or -] SEM
                                              Days post inoculation

                                                        5
Trial 1
  Control                                     0.98 [+ or -] 0.02 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.88 [+ or -] 0.02 ab
  H. bacteriophora (10 IJs/[cm.sup.2])        0.40 [+ or -] 0.04 d
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.42 [+ or -] 0.05 d
  S. glaseri (10 IJs/[cm.sup.2])              0.70 [+ or -] 0.11 bc
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.64 [+ or -] 0.12 c

Trial 2
  Control                                     0.72 [+ or -] 0.04 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.84 [+ or -] 0.03 a
  H. bacteriophora (10 IJs/ [cm.sup.2])       0.30 [+ or -] 0.04 b
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.34 [+ or -] 0.03 b
  S. glaseri (10 IJs/[cm.sup.2])              0.68 [+ or -] 0.05 a
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.66 [+ or -] 0.03 a

                                                 Mean proportion
                                             surviving [+ or -] SEM
                                              Days post inoculation

                                                       10
Trial 1
  Control                                     0.78 [+ or -] 0.08 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.76 [+ or -] 0.06 a
  H. bacteriophora (10 IJs/[cm.sup.2])        0.16 [+ or -] 0.01 c
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.16 [+ or -] 0.07 c
  S. glaseri (10 IJs/[cm.sup.2])              0.38 [+ or -] 0.10 b
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.38 [+ or -] 0.04 b

Trial 2
  Control                                     0.62 [+ or -] 0.04 a
  Rhabditis sp. (10 IJs/[cm.sup.2])           0.68 [+ or -] 0.07 a
  H. bacteriophora (10 IJs/ [cm.sup.2])       0.24 [+ or -] 0.04 c
  H. bacteriophora [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.30 [+ or -] 0.05 bc
  S. glaseri (10 IJs/[cm.sup.2])              0.48 [+ or -] 0.07 ab
  S. glaseri [+ or -] Rhabditis sp.
    (5 IJs/[cm.sup.2])                        0.34 [+ or -] 0.08 bc
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