The role of midgut nonspecific esterase in the susceptibility of Galleria mellonella larvae to Bacillus thuringiensis.

Some mechanisms of resistance to Bacillus thuringiensis, including esterase activity in the midgut, can appear in laboratory and field insect populations. We found that B. thuringiensis infection significantly reduced the esterase activity in the midgut of greater wax moth Galleria mellonella 48 h post infection. Esterase activity was inhibited 1.5-fold by triphenyl phosphate to test the impact of the enzyme in resistance to the bacteria. Bioassays demonstrated that pre-exposure of the insects to triphenyl phosphate followed by bacterial infection resulted in two-fold elevated susceptibility of the greater wax moth larvae to B. thuringiensis.

Entomopathogenic Fungi: New Insights into Host-Pathogen Interactions.

Although many insects successfully live in dangerous environments exposed to diverse communities of microbes, they are often exploited and killed by specialist pathogens. Studies of host-pathogen interactions (HPI) provide valuable insights into the dynamics of the highly aggressive coevolutionary arms race between entomopathogenic fungi (EPF) and their arthropod hosts. The host defenses are designed to exclude the pathogen or mitigate the damage inflicted while the pathogen responds with immune evasion and utilization of host resources. EPF neutralize their immediate surroundings on the insect integument and benefit from the physiochemical properties of the cuticle and its compounds that exclude competing microbes. EPF also exhibit adaptations aimed at minimizing trauma that can be deleterious to both host and pathogen (eg, melanization of hemolymph), form narrow penetration pegs that alleviate host dehydration and produce blastospores that lack immunogenic sugars/enzymes but facilitate rapid assimilation of hemolymph nutrients. In response, insects deploy an extensive armory of hemocytes and macromolecules, such as lectins and phenoloxidase, that repel, immobilize, and kill EPF. New evidence suggests that immune bioactives work synergistically (eg, lysozyme with antimicrobial peptides) to combat infections. Some proteins, including transferrin and apolipophorin III, also demonstrate multifunctional properties, participating in metabolism, homeostasis, and pathogen recognition. This review discusses the molecular intricacies of these HPI, highlighting the interplay between immunity, stress management, and metabolism. Increased knowledge in this area could enhance the efficacy of EPF, ensuring their future in integrated pest management programs.

[COMPARATIVE ANALYSIS OF THE IMMUNE REACTIONS IN COLORADO POTATO BEETLE LARVAE UNDER DEVELOPMENT OF MYCOSES CAUSED BY METARHIZIUM ROBERTSII, M. BRUNNEUM AND M. PEMPHIGI].

A comparative investigation of humoral and cellular immune response in larvae of Colorado potato beetle Leptinotarsa decemlineata was conducted under development of mycoses caused by entomopatho- genic fungi Metarhizium robertsii, M. brunneum and M. pemphigi. The larvae were found highly suscep- tible to M. robertsii, M. brunneum and less susceptible to M. pemphigi. The susceptibility to the fungi was not correlated with the rate of conidia germination in epicuticular extracts of larvae. A non-specific for Colorado beetle pathogen M. pemphigi did not cause significant changes in the immune response and did not result in colonization of larvae hemocoel. Infection with M. robertsi and M. brunneum led to an increase in total hemocyte count at the initial stages of mycoses (day 2) followed by a sharp decrease on day 3. The strongest decrease was observed for the immunocompetent cells - plasmatocytes and granu- locytes. Enhanced phenoloxidase activity in hemolymph and cuticle was found on days 2 and 3 after in- fection. These changes in immune reactions correlated with the level of virulence of the strains. Thus, the immune response in Colorado potato beetle larvae is an important factor determining differences in the development of mycoses caused by different Metarhizium species.

Intra- and trans-generational effects of larval diet on susceptibility to an entomopathogenic fungus, Beauveria bassiana, in the greater wax moth, Galleria mellonella.

In addition to nutritional conditions experienced by individuals themselves, those experienced by their parents can affect their immune function. Here, we studied the intra- and trans-generational effects of larval diet on susceptibility to an entomopathogenic fungus, Beauveria bassiana, in the greater wax moth, Galleria mellonella. In the first part of the study, a split-brood design was used to compare the susceptibility of full sibs raised either on low- or on high-nutrition larval diet. In the second part of the study, a similar experimental design was employed to investigate the effects of maternal and paternal diet as well as their interaction on offspring's susceptibility. In the first part of the study, we found that individuals fed with high-nutrition diet had higher mortality from infection than individuals fed with low-nutrition diet. However, diet did not affect post-infection survival time. Conversely, in the second part of the study, maternal diet was found to have no significant effect on final mortality rate of offspring, but it affected survival time: larvae with high-nutrition maternal diet survived fewer days after infection than larvae with low-nutrition maternal diet. Paternal diet had no significant effect on offspring's susceptibility to the fungus, indicating that paternal effects are not as important as maternal effects in influencing immune function in this species. Our findings provide further indication that maternal nutrition affects immune function in insects, and suggest that the direct effects of nutrition on immunity may be different, yet parallel, to those caused by parental nutrition.

Contributions of cellular and humoral immunity of Galleria mellonella larvae in defence against oral infection by Bacillus thuringiensis.

In this study the cellular and humoral immune reactions of the Greater wax moth Galleria mellonella have been investigated during bacterial infection caused by oral administration of Bacillus thuringiensis. Two different dose strengths were investigated to assess the contribution of immune parameters to induced Bt resistance. Low-dose (sublethal LC15) infection resulted in significantly elevated haemolymph phenoloxidase and lysozyme-like activity, enhanced phagocytic activity of haemocytes, and increased encapsulation responses in infected larvae at 48 and 72 h post infection. Higher doses of Bt (half-lethal LC50) also triggered significantly elevated haemolymph phenoloxidase and lysozyme-like activity, but decreased the coagulation index and activity of phenoloxidase in haemocytes of infected larvae. In both types of infection, the pool of circulating haemocytes became depleted. The importance of cellular and humoral immune reactions in induced insect resistance to intestinal bacterial infection Bt is herein discussed.

More than a colour change: insect melanism, disease resistance and fecundity.

A 'dark morph' melanic strain of the greater wax moth, Galleria mellonella, was studied for its atypical, heightened resistance to infection with the entomopathogenic fungus, Beauveria bassiana. We show that these insects exhibit multiple intraspecific immunity and physiological traits that distinguish them from a non-melanic, fungus-susceptible morph. The melanic and non-melanic morphs were geographical variants that had evolved different, independent defence strategies. Melanic morphs exhibit a thickened cuticle, higher basal expression of immunity- and stress-management-related genes, higher numbers of circulating haemocytes, upregulated cuticle phenoloxidase (PO) activity concomitant with conidial invasion, and an enhanced capacity to encapsulate fungal particles. These insects prioritize specific augmentations to those frontline defences that are most likely to encounter invading pathogens or to sustain damage. Other immune responses that target late-stage infection, such as haemolymph lysozyme and PO activities, do not contribute to fungal tolerance. The net effect is increased larval survival times, retarded cuticular fungal penetration and a lower propensity to develop haemolymph infections when challenged naturally (topically) and by injection. In the absence of fungal infection, however, the heavy defence investments made by melanic insects result in a lower biomass, decreased longevity and lower fecundity in comparison with their non-melanic counterparts. Although melanism is clearly correlated with increased fungal resistance, the costly mechanisms enabling this protective trait constitute more than just a colour change.

[An increase in the immune system activity of the wax moth Galleria mellonella and of the Colorado potato beetle Leptinotarsa decemlineata under effect of organophosphorus insecticide].

There has been performed evaluation of the effect of the organophosphorus insecticide (pirimifos-methyl) on some components of the insect immune response. The cellular (a change of the number of hemocytes and of intensity of incapsulation) and the humoral (a change of phenoloxidase activity) components of the immune response were studied in larvae of representatives of two orders--the Colorado potato beetle (Leptinotarsa decemlineata, Chrysomelidae, Coleoptera) and the wax moth (Galeriia mellonella, Pyralidae, Lepidoptera). The action of the insecticide has been found to lead to stimulation of immune reactions (an increase of phenoloxidase activities and of intensity of incapsulation, a rise of the number of hemocytes) at the contact treatment of both sublethal and the half-lethal doses of pirimifos-methyl.

The effects of dietary nickel on the detoxification enzymes, innate immunity and resistance to the fungus Beauveria bassiana in the larvae of the greater wax moth Galleria mellonella.

In this study, we tested the effects of dietary nickel on the activity of glutathione S-transferase (GST), esterases, phenoloxidase, and encapsulation in the haemolymph of larvae of the greater wax moth Galleria mellonella. We also explored the effects of dietary nickel on larval resistance to infection by the fungus Beauveria bassiana. Larvae fed a low dose of nickel (10 µg g(-1)) had significantly higher GST, phenoloxidase activity and encapsulation responses than controls fed on a nickel-free diet. We also found that larvae fed a sublethal dose of nickel (50 µg g(-1)) had increased GST, esterase activity and encapsulation rates but decreased phenoloxidase activity. Although, a sublethal dose of dietary nickel enhanced innate immunity, we found that this reduced resistance against the real pathogen. Our results suggest that enhanced immunity and detoxification enzyme activity of insects may not be beneficial to resistance to fungal infection. It appears that there is a trade off between different resistance mechanisms in insects under different metal treatments.

The effect of Habrobracon hebetor venom on the activity of the prophenoloxidase system, the generation of reactive oxygen species and encapsulation in the haemolymph of Galleria mellonella larvae.

The cellular and humoral immune reactions in haemolymph of the wax moth Galleria mellonella larvae naturally injected by venom of ectoparasitic wasp Habrobracon hebetor were analyzed. A strong decline of phenoloxidase (PO) activity in the haemolymph and the number of haemocytes with PO activity of envenomated wax moth was observed. In addition, it has been shown that the rate of capsule melanization in the envenomated larvae was half that of the control. Also production of reactive oxygen species (ROS) in the haemolymph of envenomated larvae decreased. The obtained data casts light on the suppression of the main immune reactions in G. mellonella larvae during natural envenomation by H. hebetor.

Effect of bacterial infection on antioxidant activity and lipid peroxidation in the midgut of Galleria mellonella L. larvae (Lepidoptera, Pyralidae).

Bacillus thuringiensis is one of the most widely used sources of biorational pesticides, as well as a key source of genes for transgenic expression to provide pest resistance in plants. In this study the effect of Bacillus thuringiensis ssp. galleriae (Bt) infection on the activity of superoxide dismutase (SOD), glutathione S-transferase (GST), catalase (CAT), concentrations of oxidated and reduced thiols (RSSR/RSH) and malondialdehyde (MDA) was tested in the midgut of Galleria mellonella larvae. We found that Bt infection resulted in increased activities of SOD, GST, malondialdehyde and RSSR/RSH ratio the first day after inoculation. However, catalase activity decreased on the first and following days after bacterial infection by Bt. Our results confirm the hypothesis that Bt infection increases the level of oxidative stress in the larval midgut. In light of this study, it seems possible that oxidative damage contributes to cell death in the midgut during bacteriosis.

Phagocytic activity and encapsulation rate of Galleria mellonella larval haemocytes during bacterial infection by Bacillus thuringiensis.

The bacterium Bacillus thuringiensis (Bt) is a pathogen of many insect species and is actively used in biocontrol. After the peroral inoculation of Galleria mellonella by the Bt in 5% sublethal concentration (LC(5)), a 1.5-fold increase in the phagocytic activity of infected larvae has been registered on the second and third days after the inoculation. With the increase of Bt-inoculum amount to 15% of sublethal concentration (LC(15)), a further increase of the phagocytic activity and enhanced encapsulation rates in the haemolymph of infected larvae has been observed. The enhanced cellular immunity during the bacteriosis seems to have resulted from the destruction of midgut epithelium cells followed by the subsequent exposure of gut content to lymph factors activating the immune system of haemocoel.