Diapause-induced shift in the content of major carbohydrates in Chilo partellus (Swinhoe).

Although several aspects like diapause determining factors, population structure, reproductive physiology, and genetics of diapause have been investigated, there is no clarity on carbohydrate energetics during larval diapause in Chilo partellus (Swinhoe). Present studies revealed significant variation between the nondiapausing and diapausing C. partellus for total carbohydrates, glycogen, sorbitol, and trehalose contents in different body parts, life stages, and for body parts × life stages interaction. Total carbohydrate content started declining, while sorbitol and trehalose increased in all the body parts as the C. partellus larvae progressed from prediapausing to diapausing state. However, glycogen content spiked in all the body parts at prediapausing stage, which then declined during diapause. Among the body parts, total carbohydrate content was significantly greater in the hemolymph as compared to other body parts of both larvae and pupae of C. partellus. Glycogen content was significantly greater in the larval fat bodies and pupal hemolymph as compared to their other body parts. In diapausing larvae, sorbitol and trehalose were greater in the integument than in other body parts. Furthermore, there was spike in trehalose and decrease in sorbitol in all the body parts of pupae from diapausing than those from nondiapausing larvae. These findings suggest that the diapause alterate and/or fluctuate major carbohydrates in different body parts of both larvae and pupae of C. partellus. This information will be helpful in better understanding the diapause energetics and overwintering metabolic cryoprotection in insects.

Nucleo-cytoplasmic interactions affecting biological performance of Lipaphis erysimi in Brassica juncea.

Hybrids have been successfully used to improve crop productivity, including Brassicas. Nucleo-cytoplasmic interactions have been reported to influence the expression of resistance to insect pests in several crops. We studied the effects of Cytoplasmic Male Sterility (CMS) in Brassica juncea carrying alien cytoplasms and their respective maintainer (B) lines on the antibiosis mechanism of resistance, involving development, survival, reproduction potential and population build-up of mustard aphid, Lipaphis erysimi, and the levels of defense phyto-chemicals. Present findings revealed that the numbers of aphids/plant, aphid multiplication rate and aphid resistance index were lower on ber CMS under natural, mori CMS under artificial infestation conditions, and juncea under both the test conditions indicating nucleo-cytoplasmic interactions for aphid reaction. Across cytoplasms, nymphal, reproductive and total developmental periods were significantly longer on SEJ 8, NPJ 161, LES 39, and NPJ 93, while the reproductive potential and survival were lower on PM 30, Pusa Tarak and SEJ 8 as compared to other nuclear backgrounds. Across nuclear backgrounds, nymphal, reproductive and total developmental periods were significantly longer on ber CMS, while reproductive potential and survival were lower on ber and mori CMS as compared to other cytoplasms. Total glucosinolates were significantly greater and myrosinase lower in Pusa Agrani, SEJ 8, LES 39, PM 30, NPJ 112, and Pusa Tarak as compared to the other nuclear backgrounds. Furthermore, total glucosinolates were significantly greater and myrosinase lower in ber CMS and juncea as compared to other cytoplasms. The studies suggest that CMS as well as cytoplasmic and nuclear gene interactions regulate the expression of defense compounds such as glucosinolates and determine the expression of resistance/susceptibility to L. erysimi. These findings shall help in identification of suitable L. erysimi tolerant nucleo-cytoplasmic combinations for their deployment in B. juncea hybrid breeding program.

Whole genome sequencing of spotted stem borer, Chilo partellus, reveals multiple genes encoding enzymes for detoxification of insecticides.

Spotted stem borer, Chilo partellus, is the most important constraint for increasing the production and productivity of maize and sorghum, the two major coarse cereals in Asia and Africa. The levels of resistance to this pest in the cultivated germplasm are low to moderate, and hence, farmers have to use insecticides for effective control of this pest. However, there is no information on the detoxification mechanisms in C. partellus, which is one of the constraints for deployment of appropriate insecticides to control this pest. The ability to detoxify insecticides varies across insect populations, and hence, we sequenced different populations of C. partellus to identify and understand detoxification mechanisms to devise appropriate strategies for deployment of different insecticides for controlling this pest. Larval samples were sequenced from three different cohorts of C. partellus using the Illumina HiSeq 2500 platform. The data were subjected to identify putative genes that are involved in detoxification on insecticides in our cohort insect species. These studies resulted in identification of 64 cytochrome P450 genes (CYP450s), and 36 glutathione S-transferases genes (GSTs) encoding metabolic detoxification enzymes, primarily responsible for xenobiotic metabolism in insects. A total of 183 circadian genes with > 80% homolog and 11 olfactory receptor genes that mediate chemical cues were found in the C. partellus genome. Also, target receptors related to insecticide action, 4 acetylcholinesterase (AChE), 14 γ-aminobutyric acid (GABA), and 15 nicotinic acetylcholine (nAChR) receptors were detected. This is the first report of whole genome sequencing of C. partellus useful for understanding mode of action of different insecticides, and mechanisms of detoxification and designing target-specific insecticides to develop appropriate strategies to control C. partellus for sustainable crop production.

The amino acid and lipophilic profiles of Chilo partellus (Swinhoe) larvae fluctuate with diapause.

The Chilo partellus (Crambidae: Lepidoptera) larvae undergoes both hibernation and estivation in India. Although, much has been done on reproductive physiological aspects, little is known about biochemical changes happening during hibernation and estivation in C. partellus. Thus, we mapped changes in amino acid and lipophilic profiles of C. partellus larvae while undergoing hibernation and estivation using high-performance liquid chromatography and gas chromatography mass spectroscopy. The studies revealed higher amounts of amino acids namely, serine, glycine, histidine, arginine, proline, tyrosine, and methionine in estivation, while lower in hibernation as compared with nondiapause larvae of C. partellus. Furthermore, the amounts of aspartic acid, glutamic acid, and alanine in hibernation, and threonine, valine, isoleucine, phenylalanine, and leucine in estivation were on par with nondiapause larvae. The lipophilic compounds namely, linoleic acid, stearic acid, eicosanoic acid, and n-pentadecanol were lower in hibernation than estivation and nondiapause larvae of C. partellus. Palmitoleic acid and methyl 3-methoxytetradecanoate contents were higher in hibernation than estivation and nondiapause, while myristic acid and lathosterol contents were higher in estivation than hibernation and nondiapause larvae of C. partellus. Cholesterol content was higher, while squalene and gamma-ergostenol were lower in hibernation and estivation as compared with nondiapause larvae of C. partellus. These findings suggest that certain amino acids may be constituents of heat-shock proteins and help C. partellus during estivation. However, the lipophilic compounds could be helpful in maintaining development during hibernation and estivation in C. partellus.

Biological and biochemical diversity in different biotypes of spotted stem borer, Chilo partellus (Swinhoe) in India.

Because of variation in incidence and severity of damage by Chilo partellus (Swinhoe) in different geographical regions, it is difficult to identify stable sources of resistance against this pest. Therefore, the present studies were undertaken on biological attributes (damage in resistant and susceptible genotypes, survival and development) and biochemical profiles (amino acids and lipophilic compound) of C. partellus populations from eight geographical regions to understand it's population structure in India. There was a significant variation in biological attributes and biochemical profiles of C. partellus populations from different geographical regions. Based on virulence and biological attributes, similarity index placed the C. partellus populations in five groups. Likewise, lipophilic and amino acid profiling also placed the C. partellus populations in five groups. However, the different clusters based on biological and biochemical attributes did not include populations from the same regions. Similarity index based on virulence, biological attributes, and amino acids and lipophilic profiles placed the C. partellus populations in six groups. The C. partellus populations from Hisar, Hyderabad, Parbhani and Coimbatore were distinct from each other, indicating that there are four biotypes of C. partellus in India. The results suggested that sorghum and maize genotypes need to be tested against these four populations to identify stable sources of resistance. However, there is a need for further studies to establish the restriction in gene flow through molecular approaches across geographical regions to establish the distinctiveness of different biotypes of C. partellus in India.

Ecotoxicity of neonicotinoids and diamides on population growth performance of Zygogramma bicolorata (Coleoptera: Chrysomelidae).

Relative ecotoxicity of approved neonicotinoids (i.e. imidacloprid, clothianidin, acetamiprid, thiacloprid, thiamethoxam and dinotefuran) and diamides (i.e. chlorantraniliprole, cyantraniliprole and flubendiamide) was examined on population growth parameters of Zygogramma bicolorata Pallister on parthenium under laboratory conditions at 27 ± 1 °C, 65 ± 5% relative humidity and 10 L : 14D photoperiod. The dose of all tested insecticides in the bioassay procedure was within a minimum range of their recommended field rate. In acute toxicity trial, imidacloprid caused highest rate of mortality in treated adults of Z. bicolorata, however, it was lowest in flubendiamide treatment followed by cyantraniliprole and chlorantraniliprole. Further, based on toxicity coefficient (E) value in acute toxicity trial, all were classified as harmful (H) and diamides were classified as moderately harmful (MH) as per IOBC classification. Moreover, chronic toxicity trials were carried out through life table response experiments (LTREs) in the F1 progeny of acute toxicity experienced group. Prolonged development with the highest mortality was evident in as compared to diamides. Furthermore, population growth parameters i.e. potential fecundity (Pf), natality rate (mx), intrinsic rate of increase (rm), net reproductive rate (R0) and finite rate of increase (λ) was greatly reduced in Z. bicolorata treated with neonicotinoids as compared with diamides. However, mean generation time (Tc), corrected generation time (τ) and the doubling time (DT) was prolonged in neonicotinoids followed by diamides. Furthermore, proportion of females was greatly reduced (0.43-0.48 females) in neonicotinoids as comparison to diamides (0.53-0.55 females) and control (0.67 females). On the basis of ecotoxicity trials, the tested neonicotinoids were highly toxic to Z. bicolorata than diamides. Therefore, diamide insecticides could be used with Z. bicolorata, however, for validation experimentation need to be done under natural field conditions.

Genetic regulation of diapause and associated traits in Chilo partellus (Swinhoe).

Diapause is an endocrine controlled arrested metabolic state to delay development or reproduction under unfavorable conditions. To gain an understanding of importance of diapause for ecological adaptation, it is important to study regulation of diapause in insects. We examined genetics of diapause in Chilo partellus by crossing the hibernating (HD), aestivating (AD), post-hibernating (PHD), post-aestivating (PAD), and nondiapause (ND) strains. Reciprocal crosses were also made to gain full understanding of diapause regulation and the maternal effects, if any. Data were recorded on fecundity, egg hatching, larval survival, diapause induction and termination, adult emergence, and morphometrics of larvae, pupae and adults in the parents (P1, P2), F1 hybrids, and the reciprocal crosses. Genetic analysis showed that AD strain is general combiner, which also improved egg hatching, larval survival, diapause termination, adult emergence and proportion of females in the progenies. Incidence of diapause was highest in HD × AD, whereas termination was greatest in PHD × AD. However, ND strain and its reciprocal crosses with other strains did not exhibit any noticeable developmental response associated with diapause. Specific combining ability analysis suggested that where PHD and AD strains exist together there will be likely reduction in diapause incidence, increased survival with greater fitness and faster multiplication of their progenies resulting in outbreak of C. partellus. Degree of dominance estimates revealed that diapause, developmental and morphometric traits in C. partellus are governed by over dominance gene effects, and mainly depend on parental diapause history.

Factors responsible for estivation in spotted stem borer, Chilo partellus (Swinhoe).

Diapause is an important adaptive trait in certain groups of insects at a particular stage, which contribute in controlling insect ecology, phenology, and physiology. Considering the importance of various factors in appropriate combinations, we studied effects of different temperature and photoperiod conditions on larval morphometrics, development, induction, and termination of estivation, and overall mortality due to estivation in Chilo partellus (Swinhoe). Weight, length, and head capsule width of C. partellus larvae significantly increased with increase in temperature and photoperiod treatments upto certain age and duration of exposure. According to Dyar's ratio, overall head capsule width progressed geometrically with minimum constant ratio of 1.20, and second-degree polynomial regression equations were best fitted to all the test treatment conditions. The mortality of diapausing C. partellus individuals varied between 7.0% and 37.3% under different constant and varying temperature and photoperiod regimes, which at 38°C + 14L:10D reached upto 100.0%. However, the exposure to 30°C + 12.5L:11.5D and 32°C + 13L:11D along with dry food accelerated the process of estivation, wherein the diapause larval survival was significantly higher and overall mortality of individuals due to diapause was lower suggesting these conditions appropriate for induction of estivation in C. partellus. These studies have implications for better understanding the bioecology and population regulation system under varying climatic conditions and devising novel strategies for management of C. partellus.

Fitness consequences of delayed mating on reproductive performance of Chilo partellus (Swinhoe).

We investigated the effects of mating among different-aged males and females on reproductive physiology, progeny production, and longevity of Chilo partellus (Swinhoe) adults under laboratory conditions. Present studies involved virgin males (M) and females (F) of three different ages namely, on the day of emergence (1D), next day (2D), and further next day (3D), thus comprising of a total of nine mating treatments (1DM × 1DF, 1DM × 2DF, 1DM × 3DF, 2DM × 1DF, 2DM × 2DF, 2DM × 3DF, 3DM × 1DF, 3DM × 2DF, and 3DM × 3DF). The observations were recorded on fecundity, fertility, hatchability, and longevity of male and female adults. We found that mating among old aged males and females significantly reduce fecundity, fertility, and egg hatchability. Further, the fecundity, fertility, and egg hatching were significantly higher when newly emerged females mated with either of the ages of males as compared with other mating treatments. Conversely, the 3-day-old females, when mated with either of the ages of males, laid more numbers of unfertilized eggs, thus reduction in egg hatchability. Mating among newly emerged adults significantly reduced the longevity of male and female adults. Kaplan-Meier estimation showed that none of the adults across mating treatments survived more than 6 days. The recovery of spermatophores from the reproductive tract of females from 3DM × 1DF were significantly higher than other mating treatments. However, no association was found between spermatophore recovery and longevity of male and female adults. Findings of present studies will help to devise appropriate techniques to disrupt mating and suppress C. partellus population under field conditions.

Temperature-dependent development of diapausing larvae of Chilo partellus (Swinhoe) (Lepidoptera: Crambidae).

Temperature-dependent development rate, percent diapause induction (hibernation at low temperature and aestivation at high temperature), and survival of diapausing larvae of Chilo partellus (Swinhoe, 1885) were examined on 13 constant temperatures ranging from 8 to 40°C. Development of hibernating and aestivating larvae occurred from 10 to 25°C and 27-38°C, respectively. However, no development occurred at 8°C and 40°C. To determine actual thermal conditions that affect development and trigger both kind of diapause (hibernation and aestivation), various thermal parameters were estimated by fitting the development rate data to two linear (Ordinary equation and Ikemoto & Takai) models and thirteen non-linear models. The lower thermal thresholds (Tmin) for development of diapausing larvae of C. partellus were calculated as 9.60°C and 10.29°C using the ordinary linear model and Ikemoto & Takai model, respectively. Similarly, the thermal constants (K) estimated using the ordinary linear model was 333.33 degree-days and that estimated with Ikemoto & Takai model was 338.92 degree-days. Among the non-linear models, Lactin-2 followed by Lactin-1 were found to be the best as these models estimated the critical temperatures (Tmin, Tmax and Topt) similar to those of observed values. Conclusively, the Ikemoto & Takai linear model and Lactin-2 followed by Lactin-1 non-linear models are useful and efficient for describing temperature-dependent development and estimating the temperature thresholds of diapausing larvae of C. partellus. Our findings provided fundamental information for estimation of thermal requirement and temperature based development models for diapausing larvae of C. partellus. This information will be highly useful for predicting the occurrence, seasonal emergence, number of generations and population dynamics of C. partellus.

Lipophilic metabolite profiling of maize and sorghum seeds and seedlings, and their pest spotted stem borer larvae: a standardized GC-MS based approach.

In order to better understand the biochemical interactions and to identify new biomarkers for plant resistance against insects, we proposed a suitable lipophilic profiling method for insects and their host plants. The critical components of GC-MS based analysis are: sample amount, extraction, derivatization, temperature gradient, run time, and identification of peaks. For lipophilic metabolite profiling of maize and sorghum, and their insect pest, spotted stem borer larvae, we recommend 100 mg sample weight for seeds and insect samples (whole insect body), and 200 mg for seedlings. Maize and sorghum seeds required less time for fat extraction in comparison to their seedlings and the pest fed on these seedlings. GC-MS was standardized for better separation and intensity of peaks using different temperature gradients in the range of 180-300 C. A total of 48 lipophilic compounds encompassing various classes based on their functional groups such as fatty acids, fatty alcohols, hydrocarbons, sterols and terpenoids, vitamin derivative, etc. were separated in the seedlings (30), seeds (14), and the pest (26) in the retention time range of 3.22 to 29.41 min. This method could be useful to study nutritional aspects of different field crops in relation to various stresses apart from the analysis of lipophilic compounds for better understanding of insect-plant interactions.

Influence of cytoplasmic male sterility on expression of different mechanisms of resistance in sorghum to Atherigona soccata (Diptera: Muscidae).

Atherigona soccata (Rondani) (Diptera: Muscidae) is one of the most important pests of sorghum, Sorghum bicolor (L.) Moench, in Asia, Africa, and the Mediterranean Europe. Exploitation of cytoplasmic male sterility (CMS) for hybrid production has resulted in considerable narrowing of the genetic base and may increase the vulnerability of this crop to insect pests. Therefore, we studied the expression of different mechanisms of resistance in sorghum to A. soccata in CMS (A) and maintainer (B) lines of 12 genotypes under field and greenhouse conditions. The CMS lines of A. soccata-resistant genotypes were preferred for oviposition (78.5 versus 71.5% plants with eggs) and suffered greater deadheart incidence (47.6 versus 41.6%) than the corresponding maintainer lines, whereas such differences were not apparent in CMS lines belonging to the susceptible genotypes (92.7 versus 92.3% plants with eggs and 75.6 versus 74.6% deadhearts) under multichoice field conditions. Similar differences also were observed under controlled conditions in the greenhouse. The larval period (9.0 versus 8.8 d) and pupal mortality (18.4 versus 13.4%) were greater on maintainer lines than that on the CMS lines in the resistant group. The male and female pupal weights, fecundity, and antibiosis index were greater on the CMS than on the maintainer lines. The maintainer lines showed better recovery resistance than the CMS lines, but no such differences were observed in tiller deadhearts. The differences in susceptibility to A. soccata were greater in the A. soccata resistant CMS and maintainer lines than in the CMS and maintainer lines belonging to susceptible genotypes. Conversion of A. soccata-resistant genotypes into alternate less susceptible cytoplasmic backgrounds may be undertaken for developing sorghum hybrids with stable resistance to A. soccata.

Detached leaf assay to screen for host plant resistance to Helicoverpa armigera.

The noctuid Helicoverpa armigera (Hübner) is a major insect pest of chickpea Cicer arietinum L., pigeonpea Cajanus cajan (L.) Millsp., peanut Arachis hypogaea L., and cotton Gossypium spp., and host plant resistance is an important component for managing this pest in different crops. Because of variations in insect density and staggered flowering of the test material, it is difficult to identify cultivars with stable resistance to H. armigera across seasons and locations. To overcome these problems, we standardized the detached leaf assay to screen for resistance to this pest in chickpea, pigeonpea, peanut, and cotton under uniform insect pressure under laboratory conditions. Terminal branch (three to four fully expanded leaves) of chickpea, first fully expanded leaf of cotton, trifoliate of pigeonpea, or quadrifoliate of peanut, embedded in 3% agar-agar in a plastic cup/jar of appropriate size (250-500-ml capacity) infested with 10-20 neonate larvae can be used to screen for resistance to H. armigera. This technique keeps the leaves in a turgid condition for approximately 1 wk. The experiments can be terminated when the larvae have caused > 80% leaf damage in the susceptible check or when differences in leaf feeding between the resistant and susceptible checks are maximum. Detached leaf assay can be used as a rapid screening technique to evaluate germplasm, segregating breeding materials, and mapping populations for resistance to H. armigera in a short span of time with minimal cost, and under uniform insect infestation. It also provides useful information on antibiosis component of resistance to the target insect pest.