Wax Moth Research Articles

Immunomodulation and Protective Effects of Cordyceps militaris Extract Against Candida albicans Infection in Galleria mellonella Larvae

Cordyceps militaris-derived formulations are currently used for multiple purposes because of their medical properties, especially immune system modulation. This study analyzes the inhibitory effects of C. militaris aqueous extract on Candida albicans infections and the immune response in larvae of the greater wax moth Galleria mellonella (Lepidoptera: Pyralidae). Larvae exhibited melanization within 1 h of being infected with C. albicans inoculum at a concentration of 106 cells/larvae, and died within 24 h from a lethal dose. Aqueous extract of C. militaris proved to be nontoxic at concentrations of 0.25 and 0.125 mg/larvae, and had the greatest ability to prolong the survival of larvae infected with a sublethal dose of C. albicans at a concentration of 105 cells/larvae. Moreover, the number of hemocytes in the hemolymph of G. mellonella increased after infection with C. albicans and treatment with the aqueous extract of C. militaris at 1, 24, and 48 h by 1.21 × 107, 1.23 × 107, and 1.4 × 107 cells/100 µL, respectively. The highest number of hemocytes was recorded after treatment of infected G. mellonella with the extract for 48 h. Transcriptional upregulation of the immune system was observed in certain antimicrobial peptides (AMPs), showing that the relative expression of galiomicin, gallerimycin, and lysozyme genes were upregulated as early as 1 h after infection. Therefore, we conclude that C. militaris aqueous extract can modulate the immune system of G. mellonella and protect against infection from C. albicans.

Effect of a Low Cost Diet on the Quality Control of the Greater Wax Moth, Galleria mellonella for Improving the Sterile Insect Technique

Effect of a Low Cost Diet on the Quality Control of the Greater Wax Moth, Galleria mellonella for Improving the Sterile Insect Technique

Acquired blaCfxA-3 carried by a conjugative transposon or duplicated intrinsic blaCME-3 mediates cefiderocol resistance in Elizabethkingia anophelis clinical isolates

ObjectivesElizabethkingia spp. are resistant to multiple antibiotics. This study aimed to determine in vitro and in vivo activities of cefiderocol against Elizabethkingia spp. and to investigate resistance mechanisms. MethodsBloodstream isolates were collected from four hospitals. In vitro and in vivo activities were determined using broth microdilution and the wax moth model, respectively. Genome comparison and gene editing were used to confirm the contribution of target genes. Conjugation experiments and serial passage were used to determine transferability and stability, respectively. A MIC of ≤ 4 mg/L was designated as the susceptibility breakpoint. ResultsAmong 228 non-duplicated isolates, 226 exhibited a MIC of ≤ 4 mg/L with MIC50/90 of 1/2 mg/L. Two isolates had a MIC of 128 mg/L; both source patients had multiple comorbidities, were ventilator-dependent, and had not received cefiderocol previously. Resistance was attributable to acquisition of blaCfxA-3, carried by a conjugative transposon from Prevotella jejuni, and duplication of intrinsic blaCME-3, which led to its overexpression. tetQ coexisted with blaCfxA-3 in this conjugative transposon and minocycline facilitated its transfer among E. anophelis. Antibiotics prescribed for source patients did not induce blaCME-3 duplication. The stabilities of blaCfxA-3 and double blaCME-3 were 100% and > 90%, respectively, after 10-day serial passage. Cefiderocol failed to rescue moth larvae infected with resistant strains, but removal of resistance mechanisms restored in vivo efficacy. ConclusionsCefiderocol was in vitro and in vivo active against Elizabethkingia spp. but resistance may emerge due to the availability, transferability, and/or stability of resistance mechanisms.

Effect of Benzyl Alcohol on Main Defense System Components of Galleria mellonella (Lepidoptera).

Benzyl alcohol (E1519) is an aromatic alcohol used in the pharmaceutical and food industry. It is used to protect food products against microorganisms during storage, as a flavoring in the production of chocolate and confectionery products, as an important ingredient in fragrance, and as a preservative in medical products. However, little is known of its effect on insects. The main aim of this study was to determine the influence of benzyl alcohol on the defense systems of the wax moth Galleria mellonella, i.e., its cuticular lipid composition and critical elements of its immune system. A gas chromatography/mass spectrometry (GC/MS) analysis found benzyl alcohol treatment to elicit significant quantitative and qualitative differences in cuticular free fatty acid (FFA) profiles. Our findings indicate that benzyl alcohol treatment increased the levels of HSP70 and HSP90 and decreased those of HSF1, histamine, and cysteinyl leukotriene. Benzyl alcohol application also increased dismutase level in the hemolymph and lowered those of catalase and 8-OHdG. The treatment also had negative effects on G. mellonella hemocytes and a Sf9 cell line in vitro: 48-h treatment resulted in morphological changes, with the remaining cells being clearly spindle-shaped with numerous granules. The high insecticidal activity of compound and its lack of toxicity towards vertebrates suggest it could be an effective insecticide.

Galleria mellonella as an Antimicrobial Screening Model.

To combat the rising global issue of antibiotic resistance, the accelerated development of novel antibiotics is essential. Current preclinical antimicrobial development yields a significant number of leads that prove unsuitable either prior to or during clinical trials. To increase the efficiency of preclinical development, relevant, standardized, accessible, and cost-effective models must be developed. Galleria mellonella (greater wax moth) larvae are widely used as an infection model to assess microbial virulence, conduct drug toxicity testing, and serve as a preliminary means of evaluating the in vivo efficacy of novel antimicrobial compounds. These infection models have greater biological relevance than many in vitro screens of comparable throughput and decrease reliance on mammalian models when used as a pre-screen for antimicrobial testing. This protocol describes a standardized methodology for the optimization of G. mellonella infection models, which can be applied to bacterial species and antimicrobial therapeutics of choice. Using the WHO priority pathogen Pseudomonas aeruginosa as an exemplar, we outline steps that can be undertaken to develop a reproducible model of infection and therapeutic testing. This includes recommendations on experimental setup, sample preparation, and infection and treatment protocols. Integration of this model within preclinical antimicrobial development pipelines would decrease reliance on mammalian models, reduce the number of ineffective compounds reaching clinical trials, and ultimately increase the efficiency of preclinical antimicrobial development.

Toxicity and Physiological Effect of Lemongrass Essential Oil Nano-capsules on the Greater Wax Moth Larvae Galleria mellonella L. (Lepidoptera: Pyralidae)

Toxicity and Physiological Effect of Lemongrass Essential Oil Nano-capsules on the Greater Wax Moth Larvae Galleria mellonella L. (Lepidoptera: Pyralidae)

Structural characterization and bioactivity profiling of the fungal peptaibiotic tolypin reveal protective effects against influenza viruses.

In a bioprospection for new antivirals, we tested nonribosomally biosynthesized polypeptide antibiotics in MDCK II cells for their actions on influenza A and B viruses (IAV/IBV). Only tolypin, a mixture of closely related 16-residue peptaibiotics from the fungus Tolypocladium inflatum IE 1897, showed promising activity. It was selected for further investigation and structural characterization byultrahigh performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HR-MS/MS) and ultrahigh performance liquid chromatography coupled to in-source collision-induced dissociation tandem mass spectrometry (UHPLC-isCID-HR-MS/MS), revealing 12 partially co-eluting individual peptides that were fully sequenced. Since tolypin-related efrapeptins are potent inhibitors of F1/Fo-ATPase, we screened tolypin for its toxicity against MDCK II cells and larvae of the greater wax moth Galleria mellonella. We found that a nontoxic concentration of tolypin (1 µg/mL) reduced the titer of two IBV strains by 4-5 log values, and that of an H3N2 strain by 1-2 log values, but the H1N1pdm strain was not affected. The higher concentrations of tolypin were cytostatic to MDCK II cells, shifted their metabolism from oxidative phosphorylation to glycolysis, and induced paralysis in G. mellonella, supporting the inhibition of F1/Fo-ATPase as the mode of action. Our results lay the foundations for future work to investigate the interplay between viral replication and cellular energy metabolism, as well as the development of drugs that target host factors.

Transcriptional responses of detoxification genes to coumaphos in a nontarget species, Galleria mellonella (greater wax moth) (Lepidoptera: Pyralidae), in the beehive environment

The greater wax moth Galleria mellonella is a cosmopolitan pest of hives of the western honey bee Apis mellifera, where it remains exposed to varroicides applied by beekeepers in past decades as pest management chemicals for control of Varroa destructor, a devastating ectoparasite of bees. The prolonged presence of coumaphos residues, an organophosphate varroicide, in beeswax may be responsible for current levels of tolerance exhibited by G. mellonella, a non-target species that infests beehives. In this study, a field-collected strain of waxworms exhibited a higher LC50 value for coumaphos than that of a laboratory strain that had not been continuously exposed to coumaphos residues at field concentrations. Despite its higher tolerance for coumaphos, the field strain experienced growth inhibition at ecologically relevant concentration of coumaphos. Moreover, at low environmental concentrations that did not alter growth, detoxification gene expression levels were substantially altered. RNA-Seq analysis revealed 1181 and 658 differentially expressed genes in fat body and midgut, respectively, with 378 and 186 of those genes upregulated. This large-scale upregulation encompassed 21 genes encoding cytochrome P450 monooxygenases (CYPs), 13 encoding UDP-glycosyltransferases (UGTs), 5 encoding glutathione-S-transferases (GSTs), 2 encoding carboxylesterases (COEs), and 2 encoding ABC transporters (ABCs) in either tissue. Expression analysis of 13 selected candidate detoxification genes by RT-qPCR was consistent with their expression from RNA-Seq data. In sum, our results indicate that long-lasting pesticide residues in beeswax from past Varroa mite management may continue to act as selective agents on detoxification systems of hive residents other than the initial target species and that multiple resistance mechanisms to these chemicals may coexist within the beehive fauna.

Green synthesis of silver nanoparticles using carob leaf extract: Characterization and analysis of toxic effects in model organism Galleria mellonella L. (The greater wax moth).

Silver nanoparticles (Ag NPs) have been used in many studies due to their inhibitory properties on microorganisms such as bacteria and viruses. In recent years, due to global problems such as environmental pollution, the green synthesis (biosynthesis) method is frequently preferred because it is simple and low cost and does not require the use of toxic substances. The aim of this study is to synthesize silver nanoparticles (Ag NPs) from Ceratonia siliqua L. leaves and investigate their antioxidant and immunotoxic properties using Galleria mellonella last instar larvae. The UV spectrophotometer, TEM, XRD and FTIR measurements were used to characterize the Ag NPs. In this study, it was determined that the effects on antioxidant enzyme activities (SOD, CAT, GPx, GST), acetylcholinesterase (AChE) and total hemocyte count (THC) as well as phenoloxidase activity determine their effect on antioxidant defence and the immune system in model organism G. mellonella larvae. We observed that green synthesized Ag NPs accumulate in the midgut of the larvae and led to the increasing of CAT and SOD activities. GST and AChE activities were increased in the fat body of the larvae; otherwise, it was decreased in the midgut. Moreover, increases were found in THC and phenoloxidase activity. Consequently, green synthesized silver nanoparticles led to oxidative stress and immunotoxic effects on G. mellonella larvae.

Effect of Co-Diet Supplementation on Biodegradation of Polyethylene by Galleria mellonella L. (Lepidoptera: Pyralidae).

Pollution coming from plastic polymers, particularly polyethylene (PE), poses a serious threat to both humans and animals. The biodegradation of plastics facilitated by insects is a crucial and eco-friendly approach that can be employed to combat this global concern. Recently, the larvae of the greater wax moth Galleria mellonella (L.) have been recognized as avid 'plastivores'. The current study was aimed at evaluating the feeding efficiency of G. mellonella larvae on PEs of various densities with a co-diet supplementation of wheat germ + honey and beeswax. The results reveal that maximum PE consumption (9.98 ± 1.25 mg) was recorded in the case of 1.0 mm thick PE after a 24 h interval; however, the same scenario also achieved the greatest reduction in larval weight (27.79 ± 2.02 mg). A significant reduction in PE mass (5.87 ± 1.44 mg) was also observed in 1.0 mm PE when fed beeswax; however, the larvae experienced minimal weight loss (9.59 ± 3.81 mg). The larvae exhibited a higher PE consumption in 1.0 mm PE, indicating that the lower the density of PE, the greater the consumed area. Moreover, the biodegradation levels were notably higher within the 24 h interval. In conclusion, these findings suggest that the density of PEs and the supplementation of the co-diet have an impact on PE biodegradation. Additionally, the utilization of G. mellonella for the biodegradation of PE proves effective when combined with beeswax, resulting in minimal weight loss of the larvae. Our findings offer initial insights into how Galleria mellonella larvae biodegrade polyethylene (PE) of four different densities, along with co-diet supplementation. This approach helps us evaluate how varying densities affect degradation rates and provides a better understanding of the larvae's capabilities. Additionally, our observations at three specific time intervals (24, 48, and 72 h) allow us to identify the time required for achieving degradation rates. Through examining these time points, our method offers valuable insights into the initial phases of plastic consumption and biodegradation.

Development of Galleria mellonella (wax moth) larvae as a model for snakebite envenoming

Development of Galleria mellonella (wax moth) larvae as a model for snakebite envenoming

The venom of Habrobracon hebetor induces alterations in host metabolism.

The ability of parasitic wasps to manipulate a host's metabolism is under active investigation. Components of venom play a major role in this process. In the present work, we studied the effect of the venom of the ectoparasitic wasp Habrobracon hebetor on the metabolism of the greater wax moth host (Galleria mellonella). We identified and quantified 45 metabolites in the lymph (cell-free hemolymph) of wax moth larvae on the second day after H. hebetor venom injection, using NMR spectroscopy and liquid chromatography coupled with mass spectrometry. These metabolites included 22 amino acids, nine products of lipid metabolism (sugars, amines and alcohols) and four metabolic intermediates related to nitrogenous bases, nucleotides and nucleosides. An analysis of the larvae metabolome suggested that the venom causes suppression of the tricarboxylic acid cycle, an increase in the number of free amino acids in the lymph, an increase in the concentration of trehalose in the lymph simultaneously with a decrease in the amount of glucose, and destructive processes in the fat body tissue. Thus, this parasitoid venom not only immobilizes the prey but also modulates its metabolism, thereby providing optimal conditions for the development of larvae.

Transcriptome analysis and functional study of phospholipase A2 in Galleria mellonella larvae lipid metabolism in response to envenomation by an ectoparasitoid, Iseropus kuwanae.

There is abundant evidence that parasitoids manipulate their hosts by envenomation to support the development and survival of their progeny before oviposition. However, the specific mechanism underlying host nutritional manipulation remains largely unclear. To gain a more comprehensive insight into the effects induced by the gregarious ectoparasitoid Iseropus kuwanae (Hymenoptera: Ichneumonidae) on the greater wax moth Galleria mellonella (Lepidoptera: Pyralidae) larvae, we sequenced the transcriptome of both non-envenomed and envenomed G. mellonella larvae, specifically targeting genes related to lipid metabolism. The present study revealed that 202 differentially expressed genes (DEGs) were identified and 9 DEGs were involved in lipid metabolism. The expression levels of these 9 DEGs relied on envenomation and the duration post-envenomation. Further, envenomation by I. kuwanae induced an increase in triglyceride (TG) level in the hemolymph of G. mellonella larvae. Furthermore, silencing GmPLA2 in G. mellonella larvae 24h post-envenomation significantly decreased the content of 4 unsaturated fatty acids and TG levels in the hemolymph. The content of linoleic acid and α-linoleic acid were significantly decreased and the content of oleic acid was significantly increased by exogenous supplement of arachidonic acid. Meanwhile, the reduction in host lipid levels impairs the growth and development of wasp offspring. The present study provides valuable knowledge about the molecular mechanism of the nutritional interaction between parasitoids and their hosts and sheds light on the coevolution between parasitoids and host insects.

Bioefficacy and molecular characterization of Bacillus thuringiensis strain NBAIR BtGa against greater wax moth, Galleria mellonella L.

Galleria mellonella, the greater wax moth has always been an important pest against honeybees and has remained a nightmare for beekeeping farmers. Management of G. mellonella in live honeybee colonies is very difficult because most current management practices can destroy whole honeybee colonies. In the present study, experiments were conducted to isolate and characterize Bacillus thuringiensis from infected greater wax moth cadavers and to evaluate their biocontrol ability against G. mellonella. The bioefficacy of these isolates has been evaluated against greater wax moth along with the standard strain HD-1. Among all the strains tested, NBAIR BtGa demonstrated higher efficacy compared to other strains, with an LC50 value of 125.17µg/ml, whereas HD-1 exhibited a significantly higher LC50 value of 946.61µg/ml. Considering the economic importance of NBAIR BtGa we performed whole genome sequencing of this strain resulting in the identification of a genome size of 5.96Mb consisting of 6888 protein-coding genes. Gene ontology analysis categorized these genes into three groups based on their roles, i.e., biological functions (2169 genes), cellular components (1900 genes), and molecular functions (2774 genes). Through insecticidal toxicity-related genes (ITRG) profiling of our strain across the genome by Bt toxin scanner and cry processor resulted in the identification of several Cry proteins namely Cry1Ab11, Cry1Ia44, Cry1Aa2, Cry2Af1, Cry1Da2, Cry1Eb1, Cry1Ab5, Cry1Cb2, Cry1Ac2. Besides Cry proteins, other ITRG genes, viz. Vip3Bb2, Zwittermicin A resistance proteins, Chitinase C, Mpp46Ab1, immune inhibitor A, Bmp1, Vpb4Ca1, and Spp1Aa1 were also reported, which show toxicity against lepidopteran pests. The studies were also conducted to test the biosafety of Bt toxins against honeybee larvae and adults, which showed strain NBAIR BtGa was more than 99% safer for honeybee larvae as well as adults. Thus, the data generated ascertains its effectiveness as a biocontrol agent and it can be used further for the development of bio formulation for the management of G. mellonella in honeybee colonies.

Telling Ecopoetic Stories: Wax Worms, Care, and the Cultivation of Other Sensibilities.

Recently, a beekeeper discovered the metabolic wizardry of wax worms, their ability to decompose polyethylene. While this organism has usually been perceived as a model organism in science or a pest to beekeepers, it acquired a new mode of being as potentially probiotic, inviting us to dream of a future without plastic waste. In this paper, I explore how wax worms are entangled with material practices of care and narratives that give meaning to these practices. These stories, however, are marked by manipulation, exploitation, and extermination, and call for a questioning of our modes of caring. Consequently, I offer a counter-narrative that questions our anthropocentric practices of caring and the stories we attach to them. Borrowing Puig de la Bellacasa's notion of ecopoetics, I tell another story based on my participation in the making of an art installation hosting wax worms. The installation creates an opening of a world of curiosity and cultivates a sensibility for wax worms expanding their modes of being and our capabilities of appreciation. In the end, I argue that by mattering and storying differently, we have the opportunity to challenge anthropocentric interests and make a different world of caring and co-existence possible.

Bioactivity of silverleaf nightshade (Solanum elaeagnifolium Cav.) berries parts against Galleria mellonella and Erwinia carotovora and LC-MS chemical profile of its potential extract

Natural products received much attention as an environmentally beneficial solution for pest management. Therefore, the extracts of invasive silverleaf nightshade (Solanum elaeagnifolium Cav.) weeds using their berries parts (seeds, peels and mucilage) supported by bioassay-guided fractionation were tested against both the greater wax moth (Galleria mellonella) and Erwinia carotovora pv. carotovora causes of the blackleg of potatoes. The seeds and peels of S. elaeagnifolium were successively extracted by maceration using dichloromethane (DCM), ethyl acetate (EtOAc), and ethanol (EtOH), respectively. While, its mucilage was extracted using EtOAc. The successive EtOH extract of the plant seeds had promising inhibition efficacy and the best minimal inhibition concentration (MIC) of 50 µg/ml against E. Carotovora amongst other extracts (DCM and EtOAc of the plant berries parts). Depending on dose response activity, EtOH extract had G. mellonella larval mortality and pupal duration rates (LC50; 198.30 and LC95; 1294.73 µg/ml), respectively. Additionally, this EtOH extract of seeds was fractionated using preparative TLC to three characteristic bands. The insecticidal and bacterial activities of these isolated bands (SEA, SEB, and SEC) were evaluated at a dose of 100 µg/ml, causing mortality by 48.48, 62.63 and 92.93% (G. mellonella larvae) and inhibition by 15.22, 0.00 and 31.66 mm (E. carotovora), respectively. Moreover, the separated major three bands were tentatively identified using LC-ESI-MS analysis revealing the presence of two phenolic acids; chlorogenic acid (SEA) and dicaffeoyl quinic acid (SEB) in addition to one steroidal saponin (SEC) annotated as borassoside E or yamoscin. Finally, the plant seeds’ successive EtOH extract as well as its active constituents, exhibited potential broad-spectrum activity and the ability to participate in future pest management initiatives. A field study is also recommended to validate its bio-efficacy against selected pests and to develop its formulations.

Efficient CRISPR/Cas9-mediated ebony gene editing in the greater wax moth Galleria mellonella.

The greater wax moth, Galleria mellonella (Lepidoptera, Pyralidae), is a major bee pest that inflicts considerable harm on beehives, leading to economic losses. It also serves as a valuable resource insect and a model organism. The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system plays a crucial role in improving economic insect breeding and developing efficient agricultural pest management systems in Lepidoptera. However, the CRISPR/Cas9 protocols have not been developed for G. mellonella. Here, the Gmebony knockout (KO) strain was established using the CRISPR/Cas9 genome editing system. We obtained Gmebony KO strain in the G4 generation, which took approximately 10 months. When compared with wild-type, the head, notum, and the terminal abdominal surface of 1st to 4th instar larvae in the KO strain changed from yellow to brown, and these regions of the KO strain gradually transformed into a black color from the 5th instar larvae, and the body color of the adult moth in the KO strain changed to black. The developmental period of the early larval and the following larval instars extended. The embryonic hatchability of the Gmebony KO strain was significantly decreased. The pupal body weight of the Gmebony KO strain was not affected. The feasibility of the CRISPR/Cas9 methodology was validated by single-target editing of Gmebony. Our findings provide the first evidence that the ebony gene can serve as a pigmentation reference gene for genetic modifications of G. mellonella. Meanwhile, it can be utilized in the development of genome editing control strategies and for gene function analyses in G. mellonella.

Toxicological assessment of a novel spiro‐derivative, 4‐amino‐1‐azaspiro[4.5]dec‐3‐en‐2‐one for the greater wax moth, Galleria mellonella

AbstractThe greater wax moth, Galleria mellonella L., poses a substantial threat to apiculture, adversely impacting wax quality and potentially acting as a disease vector. There is an urgent need to develop innovative and effective control strategies to address the challenges posed by this pest and ensure the sustainability of the apiculture industry. Spiro‐derivative compounds represent a novel class of environmentally safe compounds with high insecticidal properties. For example, they have demonstrated high efficacy against sap‐sucking insects such as aphids and whiteflies. However, their effect on chewing insects has not yet been well explored. In this study, we hypothesized that 4‐amino‐1‐azaspiro[4.5]dec‐3‐en‐2‐one (4A1AD), a spiro‐derivative compound, would exhibit efficacy against G. mellonella larvae. To test this hypothesis, we conducted dipping and feeding bioassays, incorporating varying concentrations of 4A1AD on fourth instar G. mellonella larvae, and recorded mortality, biochemical parameters including total soluble protein content and enzyme activities, and anatomical abnormalities resulting from treatment. Results revealed a significant increase in larval mortality with increased concentrations and treatment durations across both bioassays. Additionally, a significant decrease in protein content and an increase in phenol‐oxidase activity were observed with alterations in α and β‐esterase activities. Scanning electron microscope findings revealed abnormalities in larval cuticle, spiracular openings and legs. These findings demonstrated the efficacy of 4A1AD, indicating a potential alternative to conventional insecticides in promoting sustainable apiculture practices. Moreover, they provide valuable contributions to the development of novel strategies for effective pest management in apicultural settings.

Diagnostic study on major honeybee disease, pests and predators in North Western Ethiopia.

The study was conducted in Pawe district from Benishangul-Gumuz and Jawi and Fagita Lekoma districts from the Amhara region to investigate major honeybee pests, predators and diseases. Using a purposive sampling technique, 183 households were interviewed, and 240 samples were collected for laboratory analysis of bee disease; data were analysed using descriptive statistics. The share of hive types owned by sampled respondents was 88.6%; overall, 1.1% and 10.3% were traditional, transitional and modern beehives, respectively. About 92% of the sample respondents acquired their base colonies by catching swarm bees on the apex of trees. The majority of beekeepers executed external inspections of their colony, whereas only 50% carried out internal inspections. Based on the responses of beekeepers, around 48.9%, 56.3% and 23.1% of colonies absconded every year from Pawe, Jawi and Fagita Lekoma districts, respectively. Ants, wax moths, bee lice, beetles, spiders, birds, monkeys and honey badgers were the major honeybee pests and predators discovered in study areas in decreasing order. Concerning the incidence of Varroa mites, Nosema apis and amoeba disease, 27.5%, 60% and 71.6% of samples showed positive results in study locations, respectively. From this result, we observed that ants, wax moths, bee lice, beetles, spiders, birds, monkeys and honey badgers were the major honeybee pests and predators. The prevalence of amoeba disease was comparatively higher in highland areas and in the summer season. This finding suggests the need for the alertness of beekeepers in controlling bee disease and pests and strengthening bee colonies through seasonal colony management. There should be a strict quarantine, and check-up undertaken when a new colony is purchased from one region to another is essential.

Impacts of Combining Steinernema carpocapsae and Bracon hebetor Parasitism on Galleria mellonella Larvae.

The greater wax moth, Galleria mellonella, is a significant pest in apiculture and a well-established model organism for immunological and ecotoxicological studies. This investigation explores the individual and combined effects of the ectoparasite Bracon hebetor (B.h.) and the entomopathogenic nematode Steinernema carpocapsae (S.c.) on G. mellonella larvae. We evaluated the activity of oxidative stress enzymes, including superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione S-transferase (GST), malondialdehyde (MDA) levels, cytochrome P450 activity, cell viability using Annexin V-FITC, DNA damage via comet assay, and larval morphology through scanning electron microscopy (SEM). Control larvae exhibited higher GPx and GST activities compared to those treated with B.h., S.c., or the B.h. + S.c. combination. Conversely, MDA levels displayed the opposite trend. SOD activity was reduced in the B.h. and S.c. groups but significantly higher in the combined treatment. Cytochrome P450 activity increased in response to parasitism by B. hebetor. The Annexin V-FITC assay revealed decreased cell viability in parasitized groups (B.h. 79.4%, S.c. 77.3%, B.h. + S.c. 70.1%) compared to controls. DNA damage analysis demonstrated significant differences between groups, and SEM observations confirmed severe cuticle abnormalities or malformations in G. mellonella larvae. These findings highlight the complex interactions between B. hebetor, S. carpocapsae, and their host, G. mellonella. Additionally, they illuminate the intricate physiological responses triggered within the host larvae.