An improved method to determine the biological activity (LC50) of the granulovirus PoGV in its host Phthorimaea operculella

Granulovirus (PoGV) is a promising candidate to substitute for chemical insecticides in integrated pest management (IPM) of the potato tuber moth, Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae). A procedure to assess pathogenicity (LC₅₀) and its standardization is crucial for research into the use of pathogens as biocontrol agents. In the present study, an egg‐dip bioassay method was developed and its precision tested in a series of six bioassays over a time period of 18 months. Bioassays were carried out at 25 °C incubation temperature. The probit model as a statistical analysis method for the interpretation of concentration responses proved better than the exponential model. LC₅₀ values ranged from 2.3 × 10⁶ to 10⁷ granules ml^?1 with a mean of 5 × 10⁶ granules ml^?1 and regression slopes varied between 0.73 and 1.05 with a mean of 0.84. LC₅₀ values and slopes displayed no significant differences according to their 95% confidence limits. The rate of harvested infected larvae showed a linear increase with increasing concentration on a log‐log scale. The mean natural mortality was 15% (SE ± 1.8%) and can be considered as a main factor producing variability among bioassays. The advantages of this bioassay method and its application in PoGV research are discussed.     

Potential of a granulovirus isolate to control <Emphasis Type="Italic">Phthorimaea operculella</Emphasis> (Lepidoptera: Gelechiidae)

In this study a Brazilian granulovirus strain, PhopGV, isolated from the potato tuber moth (PTM) Phthorimaea operculella, was investigated regarding its potential for biological control and in vivo production. The relationship between mortality of P. operculella larvae and virus concentration was determined at different temperatures on potato tubers and susceptibility of P. operculella to PhopGV was also determined on potato leaves. Virulence of PhopGV to P. operculella was not affected by temperatures from 18 to 30°C. The median lethal concentration (LC₅₀) of larvae fed on potato foliage treated with PhopGV was not higher than that verified with larvae fed on treated tubers. Optimal conditions for production of virus-infected larvae were obtained by using the virus suspensions of 41 × 10⁵, 6.3 × 10⁵ and 62 × 10⁵ OBs ml⁻¹ at 18, 24 and 30°C, which resulted in 32.0, 31.4 and 34.8% of infected larvae collected, respectively. The maximum percentage of infected larvae recovered from tubers was not affected by temperature. However, time for production of virus-infected larvae was longer at 18°C and shorter at 30°C. Persistence of PhopGV was determined on stored tubers and we observed that the virus remained effective for at least two months, causing up to 84.2% mortality of P. operculella at 1 × 10⁷ OBs ml⁻¹. The pathogen was also highly virulent to tomato pinworm, Tuta absoluta, inflicting high percentage of mortality, delaying larval growth and inhibiting pupation. This Brazilian PhopGV strain has potential to control PTM larvae on potato tubers at a broad range of temperature and can be produced in vivo using virus-treated tubers.     

Effects of temperature on the activity and kinetics of the granulovirus infecting the potato tuber moth Phthorimaea operculella Zeller (Lepidoptera: Gelechiidae)

The granulovirus infecting the potato tuber moth (PoGV) is an important biocontrol agent, especially for managing the pest in rustic potato storerooms. For efficient propagation and use of baculoviruses in pest control strategies, information on the effects of temperature on virus multiplication and activity is crucial. The interaction between PoGV infection and incubation temperature on P. operculella was studied in laboratory bioassays by determining the survival, yield of virus-infected larvae, and the kinetics of virus in vivo increase. Bioassays for LC₅₀ determination by using the egg-dip method were repeated over a period of six years in controlled incubation chambers at six constant temperatures ranging from 16 to 28 °C. Additionally, at temperatures of 17 and 24 °C the kinetics of virus development and increase in larva were assessed in destructive time-series experiments. Three different virus concentrations were used for inoculation. Control mortality was significantly temperature-dependent and was well described by a second-order polynomial function, with lowest mortality at 25 °C (20%) and highest at 16 °C (>60%). LC₅₀ values and slopes of probit-mortality curves were not significantly different between temperatures. Numbers of virus-infected larvae increased exponentially with increasing log-concentration of virus inoculum; an effect of temperature was not evident. Virus granules per larva correlated highly with larval age and larval weight. Multiple regression revealed minor direct effects of temperature on virus numbers; however, with decreasing temperature, larval weight and hence virus numbers increased. As a result, temperature is an important factor to be considered in virus-production facilities. Rearing temperature in virus-production facilities should be maintained at temperatures around 24 °C.     

Pathogenicity of entomopathogenic fungus,Metarhizium anisopliae MET-GRA4 isolate on dengue vectors,Aedes albopictus and Aedes aegypti mosquito larvae (Diptera: Culicidae)

Considering the rapid transmission of the dengue virus, substantial efforts need to be conducted to ward-off the epidemics of dengue viruses. The control effort is depending on chemical insecticides and had aroused undesirable conflicts of insecticide resistance. Here, we study the entomopathogenic fungus, Metarhizium anisopliae as a promising new biological control agent for vector control. The pathogenicity effects of Metarhizium anisopliae against field and laboratory strains of Aedes albopictus and Aedes aegypti larvae were tested using the larvicidal bioassay technique. The results demonstrate that the treatments using M. anisopliae isolate MET-GRA4 were highly effective and able to kill 100% of both Ae. albopictus and Ae. aegypti mosquito larvae at a conidia concentration of 1 × 10?/ml within 7 days of the treatment period. The fungus displayed high larvicidal activity against laboratory and field strain of Ae. aegypti larvae with LC₅₀ values (9.6 × 10³/ml, 1.3 × 10³/ml) and LC₉₅ values (1.2 × 10?/ml, 5.5 × 10⁵/ml) respectively. For Ae. albopictus, LC₅₀ values for laboratory and field strains were (1.7 × 10⁴/ml, 2.7 × 10⁴/ml) and the LC₉₅ values were (2.1 × 10?/ml, 7.0 × 10⁵/ml) respectively. Interestingly, the susceptibility of field strain towards M. anisopliae was higher as compared to the laboratory strain Aedes larvae. In which, the causative agents of all the dead larvae were verified by the virulence of M. anisopliae and caused morphological deformities on larval body. The findings from this study identify this isolate could be an effective potential biocontrol agent for vector mosquitoes in Malaysia.     

Biological activity of Hyposidra talaca nucleopolyhedrovirus against all instar stages of a destructive tea pest Hyposidra talaca (Lepidoptera: Geometridae) in India

Hyposidra talaca (Walker) (Lepidoptera: Geometridae) is the most harmful pest of northeastern tea hub of India that devastates the tea production by feasting on the tea leaves. Hyposidra talaca nucleopolyhedrovirus (HytaNPV) is a natural enemy of the aforesaid pest, as it poses great obstruction in the multiplication of the pest by causing significant larval mortality. The study was undertaken to screen the virus activity against first to fifth instar H. talaca larvae. Early instar stages are found more susceptible than the late stages as they tend to reflect highest LC₅₀ value for fifth instar as 4.3 × 10⁷ POB/ml and lowest LC₅₀ value for first instar as 7 × 10⁴ POB/ml within seven days of inoculation. LT₅₀ values vary between 2.47 and 8.45 days for neonates to fifth instar. The high record of virulence of HytaNPV indicates its bright prospect as a useful microbial biopesticide.     

Susceptibility of Heliothis armiger to a commercial nuclear polyhedrosis virus

The susceptibility of Heliothis armiger larvae of different ages to a commercial nuclear polyhedrosis virus (NPV), Elcar, was determined by bioassay. The median lethal dosage (LD₅₀) increased 150-fold during the first week of larval life at 25°C, i.e., during development to early fourth instar, but daily feeding rate and thus potential virus acquisition also increased. A linear relationship was determined between log LD₅₀ and larval length, indicating that larval length constitutes a useful index for estimating the susceptibility of larval populations. Median lethal times (LT₅₀s) were similar for larvae tested at ages of 0 to 7 days and ranged from 3.6 to 8.0 days at 30°C. The amount of virus produced in a single, infected neonate was equivalent to 1.4 × 10⁶ LD₅₀s for neonates, a 900,000-fold increase on the dose supplied. The data support the practice of directing the NPV against neonates, but, on the basis of larval susceptibility alone, the age of larvae at treatment may not always be critical.     

Time-concentration-mortality response of potato tuber moth pupae to the biocontrol agent Cordyceps tenuipes

ABSTRACT

The potato tuber moth (PTM) Phthorimaea operculella is a critical potato pest. Larvae infest both foliage and tubers and mature larvae pupate in the soil or other safe places. Cordyceps tenuipes, an entomopathogenic fungus, infect lepidopteran pupae. To determine the effectiveness of this fungus as a biocontrol agent for PTM, we evaluated the time-concentration-mortality (TCM) response of PTM pupae to C. tenuipes using the following bioassays: (1) direct immersion in conidial suspensions, (2) incubation in sterilised or (3) unsterilised soilpremixed with conidia, and (4) incubation in unsterilised soil drenched with conidial suspensions to simulate field conditions. Fungal infection caused 100%, 83.3%, 73.3%, and 85.0% mortality of PTM pupae in assays 1–4, respectively. At 10⁸ conidia/mL or conidia/g concentration, assays 1 and 4 had short lethal times (LT₅₀) of 2.2 and 2.6 days compared with 3.7 and 4.8 days for assays 2 and 3, respectively. On day 7 after inoculation, assays 1 and 4 also had low lethal concentrations (LC₅₀) of 1.69 × 10³ conidia/mL and 1.10 × 10⁵ conidia/g compared with those of assays 2 and 3, which showed low virulence, with LC₅₀ of 3.50 × 10⁵ and 3.60 × 10⁶ conidia/mL, respectively. Our results demonstrate that C. tenuipes is a promising candidate for PTM biocontrol at the pupal stage. Drenching the soil surface with conidial suspensions may be the most effective method of field application.     

Lethal effects of Gliocladium virens,Beauveria bassiana and Metarhizium anisopliae on the malaria vector Anopheles albimanus (Diptera: Culicidae)

Control of Anopheles albimanus, the main vector of malaria on the coast of the State of Chiapas, is based mainly on application of chemical insecticides, which has resulted in resistance to most registered insecticides. Strategies for biological control may provide sustainable alternatives. We report on the lethal effects of a native isolate of Gliocladium virens on An. albimanus larvae and adults, compared to that of strains of Beauveria bassiana and Metarhizium anisopliae. Conidial suspensions of G. virens, B. bassiana and M. anisopliae cultured on Sabouraud agar were tested in bioassays with An. albimanus larvae and adults. Mosquito larvae were more susceptible to all fungi, compared to adults. On early and late instar larvae, M. anisopliae showed the most pathogenic effect (LC₅₀ of 1.4×10⁵ conidia/mL in early instars; 1.1×10⁵ conidia/mL in late instars), followed by G. virens (LC₅₀ of 3.3×10⁵ conidia/mL in early instars and 3.5×10⁶ conidia/mL in late instars). Metarhizium anisopliae sensu lato and the native G. virens could be considered good choices for An. albimanus control in southern Mexico.     

The susceptibility ofTribolium castaneum [Col.: Tenchrionidae] toFarinocystis tribolii [Protozoa: Schizogregarinida]

The LT₅₀ ofFarinocystis tribolii Weiser to larvae ofTribolium castaneum (Herbst) increased with the age of the insect indicating that older larvae were relatively more tolerant to the infection though there was 100 % mortality ultimately. The adults were less susceptible than larvae and between sexes, females were more susceptible than males. The number of spores produced increased with the stage of the larvae, but there was no variation in the size of spores in the different instars. The LC₅₀ on 20th and 40th day of inoculation were 1.4×10⁷ and 2.1×10⁶ respectively. Mortality-time due toF. tribolii was shorter at 35 °C than at 25 °C. Sporulation occurred earlier at 35 °C than at 25 °C.     

Effect of host plant on susceptibility of whitefly Bemisia tabaci (Homoptera: Aleyrodidae) to the entomopathogenic fungus Beauveria bassiana (Ascomycota: Hypocreales)

We determined host plant effect on susceptibility of whitefly Bemisia tabaci to the entomopathogenic fungus Beauveria bassiana under controlled conditions. Insects were reared on cucumber, eggplant, tomato or cabbage. Fungal suspensions of 1×10⁴, 10⁵, 10⁶, 10⁷ and 10⁸ conidia/mL were applied on second-instar nymphs. Nymphal survival significantly differed among different host plant species on which the nymphs were reared. Ten days after inoculation with 1×10⁸ conidia/mL, percent survival was 4.2±0.7, 9.6±0.4, 13.4±0.8, and 24.3±0.9% on cucumber, eggplant, tomato and cabbage, respectively. Average survival times of nymphs were also significantly influenced by host plant species. After inoculation with 1×10⁸ conidia/mL, survival times were 4.8±0.15, 6.0±0.11, 5.7±0.13, and 6.2±0.08 days for nymphs reared on cucumber, eggplant, tomato, and cabbage, respectively. Virulence also differed depending on host plant species; 10 days after inoculation, LC₅₀ values were 4.6×10⁴, 1.6×10⁵, 4.2×10⁵ and 2.1×10⁶ conidia/mL on cucumber, eggplant, tomato and cabbage, respectively. Nymphs on cucumber showed highest susceptibility.     

Selection of promising fungal biological control agent of the western flower thrips Frankliniella occidentalis (Pergande)

Aims: Larval stages of Frankliniella occidentalis are known to be refractory to fungal infection compared with the adult stage. The objective of this study was to identify promising fungal isolate(s) for the control of larval stages of F. occidentalis. Methods and Results: Ten isolates of Metarhizium anisopliae and eight of Beauveria bassiana were screened for virulence against second‐instar larvae of F. occidentalis. Conidial production and genetic polymorphism were also investigated. Metarhizium anisopliae isolates ICIPE 7, ICIPE 20, ICIPE 69 and ICIPE 665 had the shortest LT₅₀ values of 8·0–8·9 days. ICIPE 69, ICIPE 7 and ICIPE 20 had the lowest LC₅₀ values of 1·1 × 10⁷, 2·0 × 10⁷ and 3·0 × 10⁷ conidia ml^?1, respectively. Metarhizium anisopliae isolate ICIPE 69 produced significantly more conidia than M. anisopliae isolates ICIPE 7 and ICIPE 20. Internally transcribed spacers sequences alignment showed differences in nucleotides composition, which can partly explain differences in virulence. Conclusion: These results coupled with the previous ones on virulence and field efficacy against other species of thrips make M. anisopliae isolate ICIPE 69 a good candidate. Significance and Impact of the Study: Metarhizium anisopliae isolate ICIPE 69 can be suggested for development as fungus‐based biopesticide for thrips management.     

Toxicity of spinosad and Beauveria bassiana to the black cutworm,and the additivity of subletal doses

The main objective of this study was to assess the susceptibility of the black cutworm (Agrotis ipsilon) to the biopesticide spinosad and to a commercial formulation (GHA strain) of the entomopathogenic fungus, Beauveria bassiana. Secondly, we quantified the effects of sublethal doses of spinosad on a number of A. ipsilon fitness parameters, and interactions resulting from simultaneous applications of sub-lethal doses of spinosad and B. bassiana. Under laboratory conditions, A. ipsilon third instar larvae were highly susceptible to spinosad, with an estimated LC₅₀ of 50 ppm. The entomopathogenic fungus, B. bassiana had a lower efficacy with an estimated LC₅₀ of 7×107 spores mL^?1. Topical applications of 5, 7.5 and 10 ppm of spinosad on third instar larvae reduced larval size and increased time to pupation and to emergence. However, pupal and adult weights were not significantly different between treated and control individuals. Additivity was observed from most spinosad–B. bassiana combinations tested, thus indicating compatibility between products. We concluded that spinosad is a promising tool for controlling black cutworm larvae alone or in combination with other products.     

Susceptibility of apple clearwing moth larvae,Synanthedon myopaeformis (Lepidoptera: Sesiidae) to Beauveria bassiana and Metarhizium brunneum

Apple clearwing moth larvae, Synanthedon myopaeformis (Lepidoptera: Sesiidae) were found to be susceptible to infection by two entomopathogenic fungi: an indigenous fungus isolated from S. myopaeformis cadavers and identified as Metarhizium brunneum (Petch); and Beauveria bassiana isolate GHA. In laboratory bioassays, larvae exhibited dose related mortality after exposure to both the M. brunneum and Beauveria bassiana with 7 day LC₅₀'s of 2.9×10⁵ and 3.4×10⁵ spores/mL, respectively. Larval mortalities caused by the two isolates at 1×10⁶ spores/mL were not significantly different and 73% of the M. brunneum-treated, and 76% of the B. bassiana-treated larvae were dead 7 days post treatment, with LT₅₀'s of 5.5 and 5.1 days, respectively.     

Nosema whitei,a microsporidan pathogen of some species ofTribolium

The pathogenicity ofNosema whitei was studied using a dose-mortality technique; larvae ofTribolium castaneum were reared for the duration of each experiment in flour mixed with known numbers of spores. The susceptibility of each of the first 5 larval instars was compared. The LD₅₀ (for mortality after 20 days) increased consistently from the first instar (1.8×10⁶ spores/g) to the fifth instar (1.0×10¹⁰ spores/g). The slopes of the probit lines increased consistently as age increased (from b=1.1 to b=3.9). Two factors which reduce the development time ofT. castaneum, high temperature and high humidity, both reduced the pathogenicity ofN. whitei. Thus pathogenicity decreased as the temperature was increased fram 25°C (LD₅₀=4.2×10⁶) through 30°C (LD₅₀=1.3×10⁷) to 35°C (LD₅₀=3.2×10⁶), also pathogenicity decreased consistently as humidity was increased fram 10%, through 30, 50, 70% to 90% R.H. Adults, emerging fromNosema free larvae, became infected only when exposed to a very high dose (2×10¹⁰ spores/g for 14 days from the day of emergence). Infected larvae were treated for 1 hr. at 45°C in an attempt to cure the infection. The infected larvae were not cured, rather the treatment had an adverse alfect on their survival.

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Résumé La pathogénicité deNosema whitei a été étudiée en élevant des larves deT. castaneum dans de la farine mélangée à des quantités connues de spores. La sensibilité des larves diminue uniformément en fonction de l'age; La DL₅₀ varie de 1,8×10⁶/g (1^er stade) à 1,0×10¹⁰ spores/g (5^e stade). Deux facteurs, qui accélèrent le développement deT. castaneum, des températures et des humidités élevées, réduisent tous les deux la pathogénicité deN. whitei. Les adultes ne peuvent être infectés qu'en les exposant à la dose extrêmement élevée de 2×10¹⁰ spores/g. Un traitement par la chaleur (45°C pendant une heure) n'a pas réussi à guérir les larves.

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This work financed by a Science Research Council (U.K.) studentship is based on a thesis submitted for a degree of Ph. D. at the University of Newcastle-upon-Tyne.     

Virulence of a South African isolate of the <Emphasis Type="Italic">Cryptophlebia leucotreta</Emphasis> granulovirus to <Emphasis Type="Italic">Thaumatotibia leucotreta</Emphasis> neonate larvae

Surface inoculation dose–response and time–response bioassays and detached fruit bioassays were conducted with a novel South African isolate of the Cryptophlebia leucotreta granulovirus (CrleGV-SA) against Thaumatotibia leucotreta (Meyrick) (Lepidoptera: Noctuidae) neonate larvae. LC₅₀ and LC₉₀ values were estimated to be 4.095 × 10³ and 1.185 × 10⁵ OBs ml⁻¹, respectively. LT₅₀ and LT₉₀ values were estimated to be 4 days 22 h and 7 days 8 h, respectively, categorising the virus as a fast or type 2 granulovirus. There was a conspicuous difference in behaviour between larvae on inoculated diet and untreated diet, resulting in a significant reduction in penetration of diet. Bioassays on detached Navel oranges revealed LC₅₀ and LC₉₀ values of 9.310 × 10⁷ and 1.515 × 10⁹ OBs ml⁻¹, when using data on numbers of larvae per fruit rather than on numbers of infested fruit. Field trials will be conducted.     

A laboratory evaluation of the pathogenicity of Bacillus popilliae var. rhopaea, the agent of milky disease in Rhopaea verreauxi (Coleoptera: Scarabaeidae)

Two methods of infection, i.e., feeding known numbers of spores and rearing larvae in contaminated peat, were used to bioassay the susceptibility of Rhopaea verreauxi to Bacillus popilliae var. rhopaea at 23°C. The susceptibility of the three larval instars was similar as measured by the ID₅₀ and IC₅₀ values. However, within an instar, newly molted larvae were less susceptible than mature larvae when infected by the contaminated peat method. It is suggested that this was due to reduced food intake. The range of ID₅₀ values for all bioassays with R. verreauxi larvae were 1.1 × 10⁷ to 4.0 × 10⁷ spores per larva, and IC₅₀ values were 3.4 × 10⁶ to 5.0 × 10⁷ spores per g of contaminated peat. The slope of the probit line was always low (0.6 to 1.8) except for young first-instar larvae infected by contaminated peat when the slope was 4.0. Disease per se did not affect food intake, though intake was reduced at high doses of contaminated peat. Young larvae often died without developing symptoms but, with increasing age, infected larvae were more likely to develop symptoms. Bioassays with Othnonius batesi and Rhopaea morbillosa indicated a much lower susceptibility per os than for R. verreauxi. It is concluded that the potential for using B. popilliae var. rhopaea to control R. verreauxi is high, but the bacillus is unlikely to be of value in control of O. batesi or R. morbillosa.     

Effects of the IGRs compounds,lufenuron and fenoxycarb on Leptinotarsa decemlineata (Say) (Col.: Chrysomelidae)

In this research work, the susceptibility of egg and four larval instars of Leptinotarsa decemlineata (Say) (Col.: Chrysomelidae) to Insect Growth Regulators (IGRs) compounds (lufenuron 25% EC and fenoxycarb 25% WP) was determined. Different larval instar groups were separated by measuring the head capsule width and were used in all bioassays. The data were analysed with log-probit transformation using the SPSS software. The LC₅₀ for egg was determined by dipping egg masses in different concentration of either compound for 10 s, and LC₅₀ values for each group of larvae was estimated by using treated potato plants. The LC₅₀ values of lufenuron on egg, first, second and third instars of larvae were 682.65, 40.58, 47.83 and 261.38 ppm, respectively, and for fenoxycarb, these were estimated as 897.50, 35.60, 57.91 and 355.23 ppm, respectively. The LD₅₀ values of lufenuron and fenoxycarb on second instar larvae were 139.56 and 228.42 ppm, respectively.     

Nucleopolyhedrovirus infection and/or parasitism by Microplitis pallidipes affected haemolymph titre of 20‐hydroxyecdysone in Spodoptera exigua larvae

This study examined the physiological effects of joint and separate parasitism and infection by the endoparasitoid Microplitis pallidipes Szépligeti and the nucleopolyhedrovirus (NPV), respectively, on haemolymph 20‐hydroxyecdysone (20‐E) titre in Spodoptera exigua (Hübner) larvae. The results indicated that in parasitized larvae, virus‐infected larvae (5.7 × 10³ and 5.7 × 10⁵ OB/ml) and parasitized larvae infected with virus at 5.7 × 10⁵ OB/ml, compared to healthy larvae, the 20‐E all declined during the first 3 days but began to increase from day 4 after treatment, while in jointly parasitized and infected larvae (5.7 × 10³ OB/ml), the 20‐E declined during the first 4 days but began to increase on day 5 after treatment. Meanwhile, compared to parasitized larvae, the 20‐E declined during the first 4 days but significantly increased on day 5 in jointly parasitized and infected larvae (5.7 × 10³ OB/ml), while significantly increased during the first 2 days but began to decrease from day 3 after treatment in jointly parasitized and infected larvae (5.7 × 10⁵ OB/ml). Finally, in larvae that were both parasitized and virus infected (5.7 × 10³ OB/ml), compared to just virus‐infected larvae (5.7 × 10³ OB/ml), the 20‐E was lower on days 3 and 4 but higher on other days after treatment; in larvae that were both parasitized and virus infected (5.7 × 10⁵ OB/ml), compared to just virus‐infected larvae (5.7 × 10⁵ OB/ml), the 20‐E was significantly higher at the first 2 days but lower from day 3 after treatment. Our results revealed that 2nd instar larval M. pallidipes in host bodies may release 20‐E into the haemolymph of S. exigua larvae and that NPV infection may stimulate S. exigua to release more 20‐E during its third to fourth instar larval moulting. We found that this stimulatory effect was greater with higher virus concentrations.     

The occurrence and distribution of resistance of codling moth to Cydia pomonella granulovirus in Europe

Thirty‐five codling moth (CM, Cydia pomonella L., Lepidoptera, Tortricidae) populations collected in different commercial orchards in six European countries were tested for their susceptibility to Cydia pomonella granulovirus (CpGV‐M). Including previously published data on CpGV‐M resistance, a total of 38 CM colonies showed considerably elevated LC₅₀ values, independent of the country origin. When only few test individuals are available, determination of mortality of neonate larvae at a discriminating concentration range of 10⁴ to 10⁶ OB/ml (>log4) as a direct measure of percentage susceptible individuals in a CM population is more advisable than calculation of LC₅₀ values. The >log4 mortality alone or in combination with the LC₅₀ value can be used for identification of resistance in a population. Results indicated a locally separated but widely spread occurrence of CM populations with low susceptibility to CpGV‐M. The most plausible hypothesis for the emergence of CpGV‐M resistance is its selection by repeated use of CpGV products.     

Potency of Bacillus thuringiensis and Bacillus subtilis against the cotton leafworm,Spodoptera littoralis (Bosid.) Larvae

The biological activities of two species of bacteria isolated from soil of cotton fields identified as Bacillus subtilis strain NRC313 (BS NRC313) and Bacillus thuringiensis strain NRC335 (BT NRC335) were evaluated against the third larval instar of the cotton leafworm, Spodoptera littoralis (Boisd.). The different entomopathogenic bacteria of BS NRC313and BT NRC335 contained 10 × 10⁸ cell/ml, and caused mortality of 100 and 97.3% for the above mentioned strains, respectively. Concentrations of 2.5 × 10⁸ to 10 × 10⁸ cell/ml of strains BS NRC313 and BT NRC335 were applied to the larvae: LC₅₀ were 3.3 × 10⁸ and 3.9 × 10⁸ cell/ml respectively. The influence of exposure to toxin concentrations manifested in terms of decreasing the adult emergence and prolongation of the generation period. The percentage of larvae that survived and succeeded to pupate increased by decreasing the concentration. The longevity of adult emergence that resulted from larvae treated with Bacillus subtilis were 6.0 ± 0.51 and 9.0 ± 0.63 days at 5 × 10⁸ and 2.5 × 10⁸ cell/ml, respectively compared with 9.8 ± 0.47 in control. The results indicated that Bacillus subtilis was more potent than Bacillus thuringiensis. Field applications of B. thuringiensis, B. subtilis and Reldan achieved 55.6, 67.4 and 89.4% reduction of the cotton leafworm larvae Spodoptera littoralis in clover plants under field conditions.