General Esterase Research Articles

Susceptibilities of Candidatus Liberibacter asiaticus-infected and noninfected Diaphorinacitri to entomopathogenic fungi and their detoxification enzyme activities under different temperatures.

Some entomopathogenic fungi species, Isaria fumosorosea, and Hirsutella citriformis were found to be efficient against the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae). However, the susceptibility to these fungi increases when the psyllid infected with Candidatus Liberibacter asiaticus (Las), which is transmitted by D. citri and causes citrus greening disease. In this study, we examined the Las‐infected and Las‐uninfected D. citri susceptibility to entomopathogenic fungi at different temperature regimes (5–40°C). When D. citri adults exposed to cold temperature (5°C), they showed less susceptibility to entomopathogenic fungi as compared with control (27°C). Irrespective of infection with Las, a significantly positive correlation was observed between temperature and percentage mortality caused by different isolates of I. fumosorosea, 3A Ifr, 5F Ifr, PS Ifr, and H. citriformis isolates, HC3D and 2H. In contrast, a significantly negative correlation was found between temperature and percentage mortality for 3A Ifr for both Las‐infected and Las‐uninfected psyllids. Detoxification enzymes, Glutathione S‐transferase levels in D. citri showed a negative correlation, whereas cytochrome P450 and general esterase levels were not correlated with changes in temperature. These findings revealed that detoxification enzymes and general esterase levels are not correlated with altered susceptibility to entomopathogenic fungi at the different temperature regimes. Conclusively, temperature fluctuations tested appear to be a significant factor impacting the management strategies of D. citri using entomopathogenic fungi.

Role of host switching in the development of pesticide tolerance in Helopeltis theivora (Hemiptera: Miridae), the major pest of tea in India

SummaryThe polyphagous pest Helopeltis theivora Waterhouse (Heteroptera: Miridae) is one of the major pests of tea [Camellia sinensis, (Theaceae)]. Duranta repens (Verbenaceae), an ornamental plant producing various allelochemicals, is used in many tea estates as hedge plant. This plant was earlier reported to be an alternative host of H. theivora. To gain an insight into the pest and host–plant relationships, the present work reports studies on host-based life-cycle traits, pesticide tolerance status and levels of detoxification enzymes, such as general esterases (GEs), glutathione S-transferases (GSTs) and cytochrome P450 (CYP450). Host switching in the development of pesticide tolerance in H. theivora was evident. While comparing the biology of this pest on the primary host (tea) and the alternative host (D. repens) it was found that total developmental period was higher in the Duranta-reared population, along with significantly higher nymphal mortality and significantly less fecundity. However, there was no significant difference in incubation period, hatchability, sex ratio, pre-oviposition period and adult longevity. Susceptibility of the Duranta-reared population against quinalphos (a commonly used insecticide) was 1.41 times less than the tea-reared H. theivora population. The host-based variation in the relative susceptibility against insecticide was corroborated by differential activity of the three major xenobiotic detoxifying enzymes mentioned above. GE, GST and CYP450 activities were 2.67-fold, 1.43-fold and 1.37-fold higher in the Duranta-reared population than in the tea-reared population, possibly signifying that host switching can play a role in the development of tolerance of H. theivora against insecticides.

Change in activity of detoxifying enzymes in directionally selected population of tea mosquito bug (Helopeltis theivora) (Heteroptera: Miridae) by an organophosphate insecticide

The heteropteran bug, Helopeltis theivora is a polyphagous pest attacking the foliage crop tea and several other crops grown in the old world tropics. Application of synthetic insecticides, especially organophosphate was so far an effective and economic means of conventional management of the pest. Severe selection (bottlenecking) of H. theivora population by exposure to LC80 dose of an organophosphate insecticide, monocrotophos resulted in 105 fold increase in tolerance level in the third generation. The total activity of general esterases (GE) and cytochrome P450 mono-oxygenases (CYP450) analogously increased in insecticide-selected F2 generation by 16.4 and 9.5 fold, respectively. Such enhanced enzyme activity could be related to the higher tolerance levels of the selected bug. Electropherogram of GE and CYP450 of the insecticide-selected generations indicated a change in isozyme profile with their elevated expressions. Induced isozymes in insecticide selected generations were only partially inhibited, when blocked by the insecticide. These findings imply that different groups (zones) of isozymes of the detoxifying enzymes are involved in imparting higher tolerance in insecticide-selected generations. Isozyme profiles of defence enzymes can be used as indices for identifying tolerance level in the field population of H. theivora, enabling tea planters to carry out rapid monitoring of the tolerance status of the pest populations, thereby providing a ready clue to choose an effective insecticide for pest management.

Effects of the plant volatile trans‑2-hexenal on the dispersal ability, nutrient metabolism and enzymatic activities of Bursaphelenchus xylophilus

Bursaphelenchus xylophilus causes pine wilt disease (PWD), which severely damages pine species. The plant volatile trans‑2-hexenal has strong activity against nematodes, although the precise mechanism of this inhibitory action remains unclear. In this paper, the fumigant effects of the LC10 and LC30 of trans‑2-hexenal on B. xylophilus were demonstrated. The trans‑2-hexenal treatments significantly inhibited the dispersal ability of nematodes. The results also indicated that trans‑2-hexenal affects the metabolism of nutrients and the activity of digestive enzymes. Among detoxifying enzymes, after treatment with trans‑2-hexenal, glutathione S-transferase activity increased significantly and general esterase activity decreased significantly. Based on these results, trans‑2-hexenal disturbs the normal physiological and biochemical activities of this nematode. These results provide valuable insight into the nematicidal mechanisms of trans‑2-hexenal.

Evaluation of flubendiamide-induced mitochondrial dysfunction and metabolic changes in Helicoverpa armigera (Hubner).

Phthalic acid diamide insecticides are the most effective insecticides used against most of the lepidopteran pests including Helicoverpa armigera, a polyphagous pest posing threat to several crops worldwide. The present studies were undertaken to understand different target sites and their interaction with insect ryanodine receptors (RyR). Bioassays indicated that flubendiamide inhibited the larval growth in dose-dependent manner with LD50 value of 0.72 μM, and at 0.8 μM larval growth decreased by about 88%. Flubendiamide accelerated the Ca2+ -ATPase activity in dose-dependent trend, and at 0.8 μM, the activity was increased by 77.47%. Flubendiamide impedes mitochondrial function by interfering with complex I and F0 F1 -ATPase activity, and at 0.8 μM the inhibition was found to be about 92% and 50%, respectively. In vitro incubation of larval mitochondria with flubendiamide induced the efflux of cytochrome c, indicating the mitochondrial toxicity of the insecticide. Flubendiamide inhibited lactate dehydrogenase and the accumulation of H2 O2 , thereby preventing the cells from lipid peroxidation compared to control larvae. At 0.8 μM the LDH, H2 O2 content and lipid peroxidation was inhibited by 98.44, 70.81, and 70.81%, respectively. Cytochrome P450, general esterases, AChE, and antioxidant enzymes (catalase and superoxide dismutase) exhibited a dose-dependent increasing trend, whereas alkaline phosphatase and the midgut proteases, except amino peptidase, exhibited dose-dependent inhibition in insecticide-fed larvae. The results suggest that flubendiamide induced the harmful effects on the growth and development of H. armigera larvae by inducing mitochondrial dysfunction and inhibition of midgut proteases, along with its interaction with RyR.

Acaricide susceptibility of Oligonychus coffeae Nietner (Acari: Tetranychidae) with corresponding changes in detoxifying enzyme levels from tea plantations of sub-Himalayan Terai, India

Oligonychus coffeae Nietner is cosmopolitan in its distribution and is an important pest of a number of economically important tropical and sub-tropical crops including tea. It is the most damaging acarine pest of tea crops in the sub-Himalayan Terai region of India which is mostly controlled chemically in the conventionally managed tea plantations of the region. Objectives of the present study were to i) investigate the tolerance level of O. coffeae collected from bio-organically managed plantations (BMP) (with no synthetic acaricide application) and conventionally managed plantations (CMP) (with periodic application of synthetic acaricide) to the acaricides, ’ethion’ and ’fenpropathrin’, ii) Quantify the detoxifying enzymes, general esterases (GE) of phase I and glutathione S-transferases (GST) of phase II, in O. coffeae as these are deemed important in acquiring pesticide tolerance, iii) Establish the relation of GE and GST activity levels with acaricide tolerance levels in populations of O. coffeae. The study revealed that i) BMP populations of O. coffeae were susceptible to both of the acaricides whereas CMP populations were tolerant. CMP populations of the pest showed low to medium tolerance to the organophosphate acaricide ’ethion’ whereas tolerance to the synthetic pyrethroid, ’fenpropathrin’ was high; ii) Corresponding GE and GST levels were significantly higher in CMP populations compared to that of BMP population. Electrophoretic analysis of GE isozymes of CMP and BMP populations further corroborated the quantitative study; iii) activity of the detoxifying enzymes, GE and GST were positively correlated with the tolerance level of O. coffeae populations indicating involvement of these enzymes in the development of acaricide tolerance.

Activity of carboxylesterases, glutathione-S-transferase and monooxygenase on Rhipicephalus microplus exposed to fluazuron

The objective of this study was to assess the effect of the exposure to fluazuron on the activity of common pesticide detoxification enzyme groups in the cattle tick (Rhipicephalus microplus). Engorged females of a susceptible strain (POA) and a resistant strain (Jaguar) were exposed in vitro to fluazuron and their eggs and larvae were used to compare the activities of the general esterases, mixed-function oxidases (MFO) and glutathione-S-transferase (GST). The results showed significant elevation in MFO contents and esterases activity in the resistant strain when compared with the susceptible strain, in eggs and larvae respectively. In the POA strain, the MFO activity in eggs was down-regulated by fluazuron exposure. Based on these results, it can be concluded that different detoxification enzymes can act in distinct pathways depending on the tick's development stage, and may be related to fluazuron detoxification in resistant strains.

Effects of rosemary, thyme and lemongrass oils and their major constituents on detoxifying enzyme activity and insecticidal activity in Trichoplusia ni

Although there have been many reports on the synergistic interactions among the major constituents of plant essential oils regarding insecticidal activity, their underlying mechanism of synergy is poorly understood. In our previous studies, we found each of the two most abundant constituents of rosemary (Rosmarinus officinalis L.), thyme (Thymus vulgaris L.) and lemongrass (Cymbopogon citratus Stapf.) essential oils can be synergistic against the larvae of the cabbage looper, Trichoplusia ni at their natural proportion or equivalent blending ratios. In the present study, we investigated whether the enhanced toxicity between the major constituents could be the result of inhibited enzyme activity of cytochrome P450s, general esterases or glutathione S-transferases which are highly related to the development of insecticide resistance. Overall, although some combinations showed mild inhibitory activity, at least for these essential oils and their major constituents, inhibition of detoxication enzyme activity is unlikely to be a direct cause of increased toxicity in the cabbage looper. The results point to other factors, such as multiple modes-of-action or enhanced penetration through the cuticular layer, playing important roles in the elevated insecticidal activity. Moreover, application of enzyme inhibitors sometimes resulted in decreased activity when mixed with the target compounds, but these antagonistic interactions disappeared when they were applied separately, suggesting that the enzyme inhibitors can sometimes influence the penetrations of toxicants.

Synthesis, characterization, computational studies, antimicrobial activities and carbonic anhydrase inhibitor effects of 2-hydroxy acetophenone-N-methyl p-toluenesulfonylhydrazone and its Co(II), Pd(II), Pt(II) complexes

2-Hydroxyacetophenone-N-methyl p-toluenesulfonylhydrazone (afptsmh) derived from p-toluenesulfonicacid-1-methylhydrazide (ptsmh) and its Co(II), Pd(II), Pt(II) complexes were synthesized for the first time. Synthesized compounds were characterized by spectroscopic methods (FT-IR, 1H13C NMR, LC-MS, UV–vis), magnetic susceptibility and conductivity measurements. 1H and 13C shielding tensors for crystal structure of ligand were calculated with GIAO/DFT/B3LYP/6-311++G(d,p) methods in CDCl3. The vibrational band assignments were performed at B3LYP/6-311++G(d,p) theory level combined with scaled quantum mechanics force field (SQMFF) methodology. The antibacterial activities of synthesized compounds were studied against some Gram positive and Gram negative bacteria by using microdilution and disc diffusion methods. In vitro enzyme inhibitory effects of the compounds were measured by UV–vis spectrophotometer. The enzyme activities against human carbonic anhydrase II (hCA II) were evaluated as IC50 (the half maximal inhibitory concentration) values. It was found that afptsmh and its metal complexes have inhibitory effects on hCA II isoenzyme. General esterase activities were determined using alpha and beta naphtyl acetate substrates (α- and β-NAs) of Drosophila melanogaster (D. melanogaster). Activity results show that afptsmh does not strongly affect the bacteria strains and also shows poor inhibitory activity against hCAII isoenzyme whereas all complexes posses higher biological activities.

Host plant-based variation in fitness traits and major detoxifying enzymes activity in Scirtothrips dorsalis (Thysanoptera: Thripidae), an emerging sucking pest of tea

AbstractScirtothrips dorsalisHood is a polyphagous species and an important sucking pest of tea (Camellia sinensis) (Theaceae). The fitness traits ofS. dorsalison two alternative host plants:Capsicum annuumL. (chilli) (Solanaceae) andRicinus communis(castor oil plant) (Euphorbiaceae) and onC. sinensisand corresponding levels of defence enzymes was studied. The study revealed thatC. sinensisis the more suitable host ofS. dorsalisbased on faster development (13.6 days) compared to the alternative hosts,C. annuum(15.5 days) andR. communis(16.7 days), a higher fecundity (C. sinensis: 11.4 eggs;C. annuum: 9.7 eggs;R. communis: 8.6 eggs), and superior egg hatchability (C. sinensis, 92.6%;C. annuum: 82.5%; andR. communis: 74.6%). The host-based variation in the fitness traits ofS. dorsaliscorroborated in light of differential activity of three major detoxifying enzymes, such as the general esterases (GEs), glutathioneS-transferases (GSTs), and cytochrome P450 mediated monooxygenases (CYPs). Densitometric analysis of GEs showed five esterase isozymes (EST I–V) with retardation factor (Rf) values of 0.17, 0.22, 0.27, 0.35 and 0.52, respectively. The pixel density, and accordingly the profile height, varied in different host-specificS. dorsalis. A significant variation of the quantity of these enzymes was also apparent in the insect when reared on the three hosts. A 2.4 and 2.7, 1.6 and 2.0, and 2.0 and 2.3-fold higher GEs, GSTs and CYPs activity on the two non-tea hosts possibly signify a predisposition of the species for higher tolerance to insecticides, enabling the pest to switch to tea where synthetic insecticides are routinely used.

Biochemical basis of synergism between pathogenic fungus Metarhizium anisopliae and insecticide chlorantraniliprole in Locusta migratoria (Meyen).

We challenged Locusta migratoria (Meyen) grasshoppers with simultaneous doses of both the insecticide chlorantraniliprole and the fungal pathogen, Metarhizium anisopliae. Our results showed synergistic and antagonistic effects on host mortality and enzyme activities. To elucidate the biochemical mechanisms that underlie detoxification and pathogen-immune responses in insects, we monitored the activities of 10 enzymes. After administration of insecticide and fungus, activities of glutathione-S-transferase (GST), general esterases (ESTs) and phenol oxidase (PO) decreased in the insect during the initial time period, whereas those of aryl acylamidase (AA) and chitinase (CHI) increased during the initial period and that of acetylcholinesterase (AChE) increased during a later time period. Activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) decreased at a later time period post treatment. Interestingly, treatment with chlorantraniliprole and M. anisopliae relieved the convulsions that normally accompany M. anisopliae infection. We speculate that locust mortality increased as a result of synergism via a mechanism related to Ca2+ disruption in the host. Our study illuminates the biochemical mechanisms involved in insect immunity to xenobiotics and pathogens as well as the mechanisms by which these factors disrupt host homeostasis and induce death. We expect this knowledge to lead to more effective pest control.

Purification of a chymotrypsin-like enzyme present on adult Schistosoma mansoni worms from infected mice and its characterization as a host carboxylesterase.

A serine protease-like enzyme found in detergent extracts of Schistosoma mansoni adult worms perfused from infected mice has been purified from mouse blood and further characterized. The enzyme is approximately 85 kDa and hydrolyses N-acetyl-DL-phenylalanine β-naphthyl-ester, a chromogenic substrate for chymotrypsin-like enzymes. The enzyme from S. mansoni worms appears to be antigenically and enzymatically similar to a molecule that is present in normal mouse blood and so is seemingly host-derived. The enzyme was partially purified by depleting normal mouse serum of albumin using sodium chloride and cold ethanol, followed by repeated rounds of purification by one-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoresis. The purified material was subjected to tandem mass spectrometry and its derived peptides found to belong to mouse carboxylesterase 1C. Its ability to hydrolyse α- or β-naphthyl acetates, which are general esterase substrates, has been confirmed. A similar carboxylesterase was purified and characterized from rat blood. Additional evidence to support identification of the enzyme as a carboxylesterase has been provided. Possible roles of the enzyme in the mouse host-parasite relationship could be to ease the passage of worms through the host's blood vessels and/or in immune evasion.

First report of pyrethroid resistance in Rhipicephalus (Boophilus) annulatus larvae (Say, 1821) from Iran

Rhipicephalus (Boophilus) annulatus is one of the most important hard ticks parasitizing cattle in northern Iran. The aim of this study was to evaluate pyrethroid resistance levels of this species from Nur County, northern Iran. The hard ticks were collected through a multistage cluster randomized sampling method from the study area and fully engorged female R. (B.) annulatus were reared in a controlled insectary until they produced larvae for bioassay. Seventeen populations of the hard ticks were bioassayed with cypermethrin and 12 populations with lambda-cyhalothrin using a modified larval packet test (LPT). Biochemical assays to measure the contents/activity of different enzyme groups including mixed function oxidases (MFOs), glutathione S-transferases (GSTs) and general esterases were performed. Population 75 showed a resistance ratio of 4.05 with cypermethrin when compared with the most susceptible field population 66 at the LC50 level. With lambda-cyhalothrin the resistance ratio based on LC50 was 3.67 when compared with the susceptible population. The results of biochemical assays demonstrated significantly elevated levels of GSTs and esterases in populations tested compared with the heterozygous susceptible filed population and a correlation coefficient of these enzymes was found in association to lambda-cyhalothrin resistance. Based on the results, pyrethroid acaricides may operationally fail to control R. (B.) annulatus in North of Iran. This study is the first document of pyrethroid resistance in R. (B.) annulatus populations from Iran.

Effect of Artemisia annua L. essential oil on toxicity, enzyme activities, and energy reserves of cotton bollworm Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae)

AbstractThe essential oil of Artemisia annua L., a weed collected from northern Iran, was studied for its toxicity and physiological aspects on 4th instar larva of the cotton bollworm Helicoverpa armigera Hübner in controlled conditions (26±1°C, 65±10% RH and 16 L : : 8 D h). The artificial diet was used as a medium for investigating the toxicity and the effect of LC10, LC30, LC50, and LC90on the feeding efficiency of 4th instar larva. The essential oil in doses of LC10, LC30, LC50, and LC90were estimated to be 2.01%, 3.86%, 6.07%, and 18.34%, respectively. The activity of α-amylase, protease, lipase, general esterases, and glutathione S-transferase and protein, triglyceride, glucose for treated larva were measured. The results showed that all of these parameters were decreased compared with the control. Hence, A. annua essential oil is suggested as a botanical for controlling this important pest of field crops.

Effects of the microbial secondary metabolite benzothiazole on the nutritional physiology and enzyme activities of Bradysia odoriphaga (Diptera: Sciaridae)

Bradysia odoriphaga (Diptera: Sciaridae) is the major pest that damages Chinese chive production. As a volatile compound derived from microbial secondary metabolites, benzothiazole has been determined to possess fumigant activity against B. odoriphaga. However, the mechanism of action of benzothiazole is not well understood. In the present study, fourth-instar larvae of B. odoriphaga were exposed to LC10 and LC30 of benzothiazole. Sublethal concentrations (LC10 and LC30) of benzothiazole significantly reduced the food consumption of the larvae on the second day after treatment (2 DAT). However, there were no significant changes in pupal weight among the different treatments. We also measured the protein, lipid, carbohydrate, and trehalose contents and the digestive enzyme activities of the larvae, and the results suggest that benzothiazole reduced the nutrient accumulation and decreased the digestive enzyme activities of B. odoriphaga. In addition, the activity of glutathione S-transferase was significantly decreased at 6h after treatment with benzothiazole, whereas general esterase activities were significantly increased at 6 and 24h after treatment. The results of this study indicate that benzothiazole interferes in the normal food consumption and digestion process by decreasing the activities of digestive enzymes. These results provide valuable information for understanding the toxicity of benzothiazole and for exploring volatile compound for the control of this pest.

The Effect of Host Plant Species on the Detoxifying Enzymes of the Asian Citrus Psyllid,Diaphorina citri(Hemiptera: Liviidae)

Summary Diaphorina citri Kuwayama (Hemiptera: Liviidae) is a phytophagous insect and the vector of the bacterium ‘Candidatus' Liberibacter asiaticus. This is the likely causal pathogen of huanglongbing, which results in decline and possible death of citrus trees. It has been shown that host plants can affect the detoxification enzyme profile of arthropods. Here, we examined the effect of rearing D. citri on various host plant species with respect to activity of general esterases (ESTs), glutathione S-transferases (GST), and cytochrome P450 monooxygenases (P450s). These enzymes were selected because they are known to metabolize a wide diversity of insecticides and are known to directly contribute to resistance in D. citri. We reared D. citri on Citrus sinensis L., Bergera koenigii L. and Murraya paniculata (L.) Jacq. (Sapindales: Rutaceae). Following 12 generations of rearing, the activities of EST, GST, and P450 enzymes were compared between the colonies raised on the different host plants. The GST activi...

Virulence determination of Beauveria bassiana isolates on a predatory hemipteran, Andrallus spinidens Fabricius (Hemiptera: Pentatomidae)

Virulence of the two isolates of Beauveria bassiana, BB2 and AM-118, were evaluated on adults of a predatory hemipteran, Andrallus spinidens Fabricius by conidial bioassay and enzymatic activities. Results of the bioassay revealed LC50 of 37×104 and 15×103 spore/ml for isolates BB2 and AM-118, respectively. Activities of chitinase, lipase and ALP showed the higher activity in the media inoculated by AM-118 while no statistical differences were observed in activity of ACP. Although no statistical differences were found in general protease and Pr1 activities but activity of Pr2 in AM-118 was significantly higher than that of BB2. Activity of general esterases demonstrated no statistical differences when α- and β-naphtyl acetate were used as substrates but activity of glutathione S-transferase in AM-118 was higher than that of BB2 by using CDNB and DCNB as specific reagents. Results of the current study indicated higher virulence of isolate AM-118 against adults of A. spinidens by lower LC50 value and higher activities of the enzymes involved in pathogenicity. Recruiting of these isolates against C. suppressalis must be considered by their adaptability of A. spinidens. Moreover, AM-118 has been isolated from rice fields of northern Iran, so it may somehow indicate a type of host-microorganism interaction.

Immune and metabolic responses of Chilo suppressalis Walker (Lepidoptera: Crambidae) larvae to an insect growth regulator, hexaflumuron

Efficient control of Chilo suppressalis Walker is always controversial due to highly economic damage, resistance to insecticides and environmental pollutions. So, combination of safe pest controls e.g. biocontrol agents and insect growth regulators seems to be promising via integrated pest management program. Bioassay of hexaflumuron on 4th larval instars revealed concentrations of 44.34, 179.74 and 474.94µg/ml as LC10-50 values. Numbers of total hemocytes, plasmatocytes and granulocytes as well as phenoloxidase activity increased in the different time intervals following treatment by hexaflumuron. Combined effects of hexaflumuron and Beauveria bassiana Vuillemin also increased hemocyte numbers and phenoloxidase activity at different time intervals using all concentrations. Activities of general esterases assayed by α- and β-naphtyl acetate and glutathione S-transferase using CDNB and DCNB increased 1-12h post-treatment. Activities of aspartate aminotransferase, alanine aminotransferase, γ-glutamyl transferase and aldolase increased in the larvae treated by hexaflumuron. However enhanced activity of lactate dehydrogenase was only obtained by treating 180 and 470µg/ml concentrations of hexaflumuron. Activities of ACP and ALP were found to be higher than control for all time intervals even 1-12h post-treatment. The amounts of HDL and LDL increased in the highest concentrations of hexaflumuron after 12-24h of post-treatment. Amount of triglyceride was higher than that of control after 1 and 3h but it was lower in other time intervals. Amounts of glycogen and protein were lower than those of control for all time intervals except for 6 and 12h of post-treatment in case of protein. Results of the current study revealed negative effects of hexaflumuron on intermediary metabolism of Chilo suppressalis but it increased the number of hemocytes and activity of phenoloxidase which are responsible for spore removal from hemolymph. It can be concluded that hexaflumuron is able to decrease survival and biological performance of C. suppressalis via intervening in intermediary metabolism but the given results showed incompatibility of the IGR with possible microbial control.

Fitness traits of the tea defoliator, Hyposidra talaca (Walker 1860) (Lepidoptera: Geometridae) on natural and artificial diets in relation to gut enzymes and nutritional efficiencies

Summary. The looper pest Hyposidra talaca (Walker 1860) is known to cause heavy damage to tea plantations in North-East India, including Darjeeling Terai-Dooars, by defoliating the bushes. Laboratory rearing of this looper species resulted in production of a healthy population under controlled conditions (aseptic) for three consecutive generations on a newly formulated artificial diet. The performance and fitness traits of the looper species were compared on natural (tea-leaf) and artificial diets along with the quantity of its hydrolytic (amylase, protease, lipase) and detoxifying enzymes [general esterase (GE), glutathione S-transferase (GST)] when reared on these two diets. Activity of protease on the artificial diet was significantly higher than that on the natural diet; however, lipase showed only marginally increased activity on the former diet. When detoxifying enzymes such as general esterase (GE) and glutathione S-transferase (GST) were observed, they were found to be significantly higher on the natural diet. Such variations in the hydrolytic enzymes and detoxifying enzymes indicated a better capacity of the looper to utilize the artificial diet than the tea-leaf diet. The finding was further corroborated by the higher efficiencies (ECI and ECD) and production index (PI) that were recorded on the artificial diet, compared to higher maintenance cost on the tea-leaf diet. The artificial diet also supported a high survival rate of H. talaca in all generations along with higher adult emergence and fecundity. The newly designed artificial diet is a viable alternative for mass rearing and efficient production of control populations of H. talaca under sterile conditions.

Esterase and glutathione S-transferase levels associated with synthetic pyrethroid resistance in Hyalomma anatolicum and Rhipicephalus microplus ticks from Punjab, India

Larval packet test was used for assessment of resistance status against cypermethrin and deltamethrin in Hyalomma anatolicum and Rhipicephalus microplus from various districts of Punjab (India). Among the various field isolates of H. anatolicum susceptible status was recorded against cypermethrin in all isolates, whereas against deltamethrin resistance status (level I-III) was recorded. In R. microplus lower resistance levels (I-II) were recorded against cypermethrin in comparison to deltamethrin (level I-IV). Quantitative analysis of general esterase activity revealed a range of 4.21 ± 0.46 to 6.05 ± 0.55 and 2.23 ± 0.23 to 2.66 ± 0.24 µmol/min/mg protein for α- and β-esterase activity, respectively, in different field isolates of H. anatolicum and the increase in comparison to susceptible was not significant (P > 0.05). In contrast to H. anatolicum, the α- and β-esterase activity in all field isolates (except Jalandhar) of R. microplus was higher (range of 3.89 ± 0.26 to 10.85 ± 0.47 and 1.75 ± 0.08 to 5.87 ± 0.29 µmol/min/mg protein, respectively) (P < 0.001). The glutathione-S-transferase (GST) activity in field isolates of H. anatolicum and R. microplus was in the range of 0.01 ± 0.001 to 0.03 ± 0.001 and 0.02 ± 0.0003 to 0.03 ± 0.001 mM/mg/min. The enzyme ratios (α-and β-esterase and GST) and RR95 against deltamethrin of H. anatolicum isolates were correlated (P < 0.05), whereas in R. microplus only α-and β-esterase and RR50 against deltamethrin were correlated (P < 0.05).