Virulence of Three Entomopathogenic Nematode Species on Corn Stalk Borer ( Sesamia Cretica Led

Full Length Research Paper

Virulence of three entomopathogenic nematode species on corn stalk borer ( Sesamia cretica Led. Lep: Noctuidae) at different temperatures

Ugur Gozel* and Cigdem Gunes

Department of Plant Protection, Faculty of Agriculture Canakkale Onsekiz Mart University, 17020 Canakkale, Turkey.

Accepted 15 May, 2013

Three Turkish strains of entomopathogenic nematodes, Steinernema carpocapsae , Steinernema feltiae and Heterorhabditis bacteriophora were tested in the laboratory for their virulence on the last instar larvae of the corn stalk borer Sesamia cretica . Mortality rates of last instar S. cretica larvae were determined for each nematode species under four different temperature treatments. Accordingly, at 15ºC, mortality of S. cretica exposed to for S. carpocapsae , S. feltiae and H. bacteriophora was 48, 56 and 14%, respectively. This ratio at 20°C was 62, 76 and 50%, at 25°C, 82, 90 and 90%, and at 30ºC (the highest experimental temperature application) mortality was 82, 92 and 94%. These obtained results show that S. carpocapsae and S. feltiae yield similar results at all applied temperatures and their effectiveness increased as the temperature increased. Although H. bacteriophora achieved low mortality at low temperatures, it infected and attained very high levels of mortality at relatively high temperatures.

Key words: Sesamia cretica , Steinernema carpocapsae , Steinernema feltiae , Heterorhabditis bacteriophora , virulence assay.

INTRODUCTION

Entomopathogenic nematodes are known to be insect 1994). parasite since the 17 th century, but until the 1930’s their Now that the true costs of pesticide use on human necessary importance as biological control agents was health and the environment are becoming better poorly explored. However, after the 1970’s, after the understood, alternative methods to control harmful negative effects of pesticides against to insects were insects are increasingly becoming in greater demand. determined, important research on entomopathogenic Among these methods, one of the groups especially nematodes has been carried out at an ever increasing accentuated is entomopathogenic nematodes, known rate, until it is now a common biological control agent obligate insect pathogens that are commercially prepared among the alternative methods to chemical control and are effective in controlling some target insect pests (Smart, 1995). The use of entomopathogenic nematodes (Burnell and Stock, 2000; Liu et al., 2000; Nguyen and against target insect pests now forms the basis for a Duncan, 2002; Nguyen et al., 2004a, b; Qiu et al., 2004, significant body of research (Nickle, 1984; Kaya, 1985; 2005). Klein, 1990; Wouts, 1991; Georgis and Meanweiler, Entomopathogenic nematodes having a wide host

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range effectively suppress their hosts and are naturally infection by observing the condition and colour of the cadavers. In available under nearly in all environmental conditions, addition to this method, all infected S. cretica larvae were dissected increasing their potential as successful biological control in Ringer’s solution. In this way, the presence of entomopathogenic nematodes, in cadavers, was determined. Thus, all dead larvae agents (Gaugler and Kaya, 1990; Koppenhofer, 2000; encountered in the study were examined in this manner for the Nguyen et al., 2004a). Currently, entomopathogenic existence of entomopathogenic nematodes. nematodes are applied with success against many target pests in biological control (Kaya and Gaugler, 1993; Fenton et al., 2000; Koppenhofer, 2000). Statistical analysis

In this study, the effectiveness of three different Mortality percentage of corn stalk borer for each variable was entomopathogenic nematodes, Steinernema controlled by Abbotts’s formula initially and then determined for carpocapsae Weiser, Steinernema feltiae Filipjev and significance via one-way ANOVA Duncan (P<0.05). Heterorhabditis bacteriophora Poinar, isolated from Turkish soils, were tested on last instar larvae of one of the most important pests in corn production in Turkey. RESULTS Accordingly, Sesamia cretica Lederer was tested for its mortality by these three species at four different We determined the virulence of S. carpocapsae, S. feltiae temperatures under laboratory conditions. and H. bacteriophora to corn stalk borer under laboratory conditions at four different temperatures. At four different temperatures and the same application rate, we show MATERIALS AND METHODS that S. cretica infection and mortality among the three

Isolation and identification of the nematodes nematode species is correlated with temperature. While the virulence of each entomopathogenic nematode Turkish isolates of S. carpocapsae , S. feltiae and H. bacteriophora species varied according to temperature, mortality rates were used in the present study. The nematodes were isolated in in corn stalk borer larvae were observed to be highest at different cities from the Western part of Turkey. These species have higher temperatures. However, there was significant been identified by morphometrics and sequencing of D2/D3 and ITS region of rDNA using molecular identification methods (Gozel variation in the mortality rates caused by the three and Gunes, 2008). different entomopathogenic nematode species at the different temperatures. For S. carpocapsae , the lowest mortality rate occurred with 48% at 15°C. The mortality Propagation of nematodes rate in corn stalk borer at 20ºC was 62%. The highest

The nematodes were propagated in the last instar larvae of the mortality rates occurred at 25 and 30ºC (82 and 86% greater wax moth, Galleria mellonella (Lep.: Pyralidae), as respectively; Figure 1). On corn stalk borer S. feltiae described by Dutky at al. (1964). The emerged infective juveniles mortality was 56% at 15ºC, 76% at 20ºC and 90% at (IJs) were stored in Ringer solution (9.0 g NaCl, 0.42 g KCI, 0.37 g 25ºC. The highest mortality caused by S. feltiae was 90% CaCI 2, 0.2 g NaHCO 3 and 1 lt distilled water) in culture flasks for 5 at 30ºC (Figure 2). days in at 4 to 5°C prior to use in the experiment (Solomon et al., On corn stalk borer at 15ºC H. bacteriophora killed 14% 1999). of the corn stalk borer larvae, the lowest among the three nematode species tested at this temperature. At 20ºC Test organisms corn stalk borer mortality increased to 50%. This trend continued as temperature increased, resulting in mortality Last instar larvae of Corn Stalk Borers ( S. cretica , Lep.: Noctuidae) rates of 90% at 25ºC and 94% at 30 ºC (Figure 3). were used as the test insect of this study. Larvae of were collected from corn fields found in Canakkale in August 2008. Field-collected For comparative purposes the mortality of the three larvae were mass produced on corn in controlled rooms at 25±1°C entomopathogenic nematode species is presented in and % 70±5 moisture. Figure 4. At 15ºC the most effective species on corn stalk borer, was S. feltiae producing a 56% mortality on corn stalk borer. S. carpocapsae was the second effective Bioassays species at this temperature producing 48% mortality. The Infective juveniles (500 individual nematodes) of each least effective nematode species at this temperature was entomopathogenic nematode species were applied to the larvae of H. bacteriophora. corn stalk borers in 250 µl distilled water in a 4 cm diameter plastic At 20ºC, there was an increase in mortality rates on Petri dishes lined with Whatman filter paper along with one corn corn stalk borer larvae produced by all nematode species stalk borer larva. Experiments were repeated 50 times for each of nematode species at each temperature. Control Petri dishes (for tested. The most effective species was at this every temperature) consisted of 250 µl of distilled water applied to temperature was S. feltiae . It was followed by S. corn stalk borer larvae. carpocapsae with a mortality rate of 62%. H. bacteriophora was the species that most increased its effectiveness with the increase of temperature of 5ºC Determination of the infection producing a mortality rate of 50% at this temperature. At After 5 days, dead S. cretica larvae were examined for nematode 25ºC, the third highest temperature we tested, there 2180 Afr. J. Agric. Res.

Figure 1. Effect of Steinernema carpocapsae on Sesamia cretica at four different temperatures.

Figure 2. Effect of S. feltiae on S. cretica at four different temperatures.

were no statistical differences among the activities of all entomopathogenic nematodes to the corn stalk borer at three species ( S. carpocapsae caused 82% of mortality, four different temperatures revealed that mortality rates S. feltiae and H. bacteriophora caused 90% of mortality). were a function of temperature. Among the three The highest temperature of applied in experiment nematode species tested, the lowest effectiveness (30ºC) yielded the highest efficiency producing a mortality occurred at 15ºC whereas the highest effectiveness rate of 94% in H. bacteriophora followed by S. feltiae with occurred at 30ºC, the highest temperature used in a mortality rate of 92%. The lowest mortality occurred in experiment. S. carpocapsae at 86%. However, at this temperature the The lowest mortality rate on corn stalk borer larvae was differences in mortality were not statistically important. the effectiveness of H. bacteriophora at 15ºC. However, the same species at 30ºC produced a 94% mortality rate, was the highest in the experiment. With the increase in DISCUSSION temperature, H. bacteriophora was the species that increased its effectiveness the most. We show that at the This study of virulence of the three different lowest temperature this species was the least effective, Gozel and Gunes 2181

Figure 3. Effect of Heterorhabditis bacteriophora on S. cretica at four different temperatures.

Figure 4. Effect of three entomopathogenic nematode species on S. cretica at four different temperatures.

but that it also was the most effective at the highest In another study, González-Ramírez et al. (2000), temperatures. These findings are consistent with those of performed laboratory studies to determine the El-Wakeil and Hussein, who obtained similar results on effectiveness of H. bacteriophora on Mocis latipes their study of S. cretica exposed to H. bacteriophora (Guenée) (Lep.: Noctuidae). On its larvae, pupae and (realizing a 97% of mortality one week post infection). prepupae, they found mortality that ranged from 2182 Afr. J. Agric. Res.

22.5 to 100% depending on the temperature. Fenton A, Norman R, Fairbairn JP, Hudson PJ (2000). Modelling the The second species used in experiment, S. efficacy of entomopathogenic nematodes in the regulation of invertebrate pests in glasshouse. J. Appl. Ecol. 37:309-320. carpocapsae was least effective at 15ºC, and showed the Gaugler R, Kaya HK (1990). Entomopathogenic nematodes in biological highest effectiveness at 30ºC which was again the control. CRC Press, Boca Raton, FL. highest temperature in experiment with a mortality ratio of Georgis R, Manweiler SA (1994). Entomopathogenic nematodes: A 86%. The effectiveness of S. carpocapsae increased with developing biological control technology. Agric. Zool. Rev. 6:63-94. González-Ramírez M, Lezama-Gutiérrez R, Molina-ochoa J, Rebolledo each increase in temperature (although differences DO, López-Edwards M, Pescador-Rubio A (2000). 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