Massimo Iorizzo, Gianluca Albanese, Cosimo Tedino, Mariantonietta Succi, Licia Maria Di Donato, Antonio de Nigris, Dalila Di Criscio, Sonia Ganassi, Luigi Ambrosone, Antonio De Cristofaro
Honey bee gut microbiota play important roles in host nutrition, immune regulation, and health, and their composition can vary in response to environmental conditions, diet, seasonality, and beekeeping management. In this two-year field study, we investigated seasonal variation in the gut bacterial communities of Apis mellifera ligustica colonies maintained under stationary or short-distance migratory beekeeping management. Six colonies originating from the same experimental apiary were assigned to stationary or migratory management, with three migratory colonies seasonally transferred to Mount Taburno during summer and three stationary colonies remaining at Castelpoto. Bacterial communities were characterized by high-throughput sequencing of the V1–V3 region of the 16S rRNA gene. The gut microbiota was consistently dominated by characteristic honey bee-associated taxa, particularly Gilliamella and Snodgrassella, although their relative abundances varied substantially across seasons and years. Alpha-diversity analyses based on harmonized genus-level relative abundance profiles revealed significant seasonal variation in both years, whereas no significant overall effect of management system was detected. Similarly, PERMANOVA of Bray–Curtis dissimilarities revealed significant seasonal structuring of bacterial community composition in 2022 and 2023, while stationary versus migratory management did not significantly explain overall community variation. The persistence of dominant gut-associated taxa across sampling periods indicates compositional continuity of major bacterial community members despite temporal variation, but does not by itself demonstrate functional stability or microbial resilience. Colony-performance measurements showed significantly higher annual honey production in migratory than in stationary colonies in 2022, whereas no significant differences in the investigated performance parameters were detected between management systems in 2023. Thus, the productive response was not consistent between years. Although temporal variation in gut bacterial communities occurred alongside differences in colony performance, the present study does not establish a causal or functional relationship between microbiota composition and productivity. Overall, these findings identify seasonality as the predominant factor associated with variation in the gut bacterial communities of the investigated colonies and provide no evidence for a consistent overall effect of short-distance migratory management on bacterial community diversity or structure. Further studies involving larger numbers of independent colonies and apiaries, together with direct functional analyses of the microbiome and characterization of dietary and environmental factors, are required to clarify the consequences of migratory beekeeping for honey bee microbiota and colony performance.
Anuja Shrestha, Yva Eline, Kenzie Sundstrom, James T. Van Leuven
Honeybee (Apis mellifera L.) larvae are susceptible to Paenibacillus larvae, a spore-forming, Gram-positive bacterium that causes American Foulbrood (AFB), a destructive honeybee brood diseases. Current antibiotic treatments are inadequate because of rising pathogen resistance and residue accumulation in hive products, driving interest in biological and other natural strategies for AFB control. Most candidate biocontrol agents, however, have been evaluated only for their ability to inhibit P. larvae or protect larvae, with less attention to their effects on adult bees. Here we assessed the biocontrol potential of hive-associated Bacillus species against P. larvae infection. Bacteria isolated from AFB-infected combs were sequenced and screened for antagonistic activity, and active isolates, their lysates, and lysate-derived protein fractions were tested for their ability to improve larval survival during P. larvae challenge. To evaluate effects on adult bees, newly emerged workers were caged and fed the B. licheniformis lysate (ASx), and their survival and gut microbiome were compared with untreated controls and with bees fed Terra-Pro, a Terramycin-based AFB treatment. Several Bacillus species – including B. zhangzhouensis, B. subtilis, B. amyloliquefaciens, B. licheniformis, and B. mojavensis – inhibited one or more P. larvae strains. The B. licheniformis lysate (ASx) and its 30 -50 kDa protein fraction improved larval survival during P. larvae challenge. In caged adults, survival of lysate-fed bees differed from untreated controls but was comparable to that of Terra-Pro-fed bees, and their gut microbiome remained similar to that of untreated controls. Together, these results indicate that hive-associated Bacillus species – and B. licheniformis -derived proteins in particular – are promising candidates for AFB management that improve larval survival without harming adult bees more than an existing antibiotic treatment.
Comstockaspis perniciosa (Comstock, 1881) is one of the most destructive quarantine pests of fruit crops, posing a serious threat to commercial orchards and natural populations of wild apple trees. Despite its widespread distribution, information on the molecular genetic characteristics of this species in Central Asia remains limited. The aim of this study was to characterize C. perniciosa collected from commercial orchards in southeastern Kazakhstan based on external morphological observations and molecular data, determine the phylogenetic position of the examined specimens using 28S rRNA gene sequences, and detect associated parasitoids using high-throughput sequencing technologies. During the growing season in 2025, surveys were conducted in ten commercial orchards, including sites located near natural populations of wild apple trees. Species-level identification of the collected specimens was confirmed by analysis of the 28S rRNA gene. All sequences clustered with reference C. perniciosa sequences, confirming species identity. Analysis of the high-throughput sequencing data additionally revealed sequences of parasitoids belonging to the genus Aphytis (Hymenoptera: Aphelinidae) in ten samples, indicating the presence of natural enemies associated with C. perniciosa. These findings expand current knowledge of the molecular characteristics of C. perniciosa in Central Asia and demonstrate the potential of NGS (Next Generation Sequencing) technologies for the simultaneous detection of quarantine pests and their associated entomophagous natural enemies within phytosanitary monitoring programs.
Growing interest in insects capable of developing in the presence of synthetic polymers has created new opportunities to investigate host responses alongside cultivable gut microorganisms. This study evaluated cumulative adult production in Galleria mellonella maintained on a control diet or diets containing polyethylene terephthalate (PET), polystyrene (PS), or polyethylene (PE), together with the recovery of bacteria from the larval gut. Four dietary treatments control, PET, PS, and PE were established, each with five replicates of 15 initial larvae. Each experimental colony remained on its assigned diet until the initial cohort completed its life cycle and produced a second generation. Larvae from this offspring generation were selected for bacterial isolation, while the remaining individuals continued development for measurement of cumulative adult production. The isolates were characterized by sequencing the V4 region of the 16S rRNA gene and comparison using BLASTn, followed by genus-level taxonomic assignment. Cumulative adult production in the second generation differed among treatments (P = 0.0194). The control produced the highest mean number of adults (122.0 ± 56.3), whereas PET produced the lowest (41.8 ± 19.1) and was the only treatment significantly lower than the control. Fourteen bacterial isolates, selected as representatives of visually distinct colony morphotypes, were characterized; these isolates were not assumed to represent 14 distinct taxa or strains. Thirteen were assigned to Bacillus and one to Psychrobacillus. Overall, the dietary formulations evaluated were associated with variation in cumulative adult production and yielded an initial collection of cultivable gut-associated bacteria for subsequent bioprospecting studies.
Gut microbiota can influence insect physiology and ecological interactions, but how botanically distinct host plants affect gut bacterial communities in polyphagous insects remains poorly understood. We investigated the midgut bacterial communities of Loxostege sticticalis larvae fed three host plants from different genera or an artificial diet using 16S rRNA gene sequencing. Bacterial diversity, community composition, differential taxa, and predicted functional profiles were compared among dietary treatments. Dietary treatments were associated with differences in gut bacterial community structure. The MS group exhibited higher bacterial richness and diversity, whereas the CA group showed a relatively simplified bacterial community. Bray–Curtis-based PCoA and PERMANOVA indicated significant overall variation in community composition among dietary treatments, although exact pairwise PERMANOVA tests were not significant. Bacillota and Pseudomonadota were dominant across treatments, while the MS group showed increased representation of Actinomycetota-associated taxa. LEfSe identified Actinomycetota-related taxa as characteristic features of the MS group and Bacillota-related taxa as features of the AD group. PICRUSt2 suggested differences in predicted functional potential among dietary treatments, including pathways related to amino acid metabolism, metabolism of cofactors and vitamins, and xenobiotics biodegradation and metabolism. The results demonstrate diet-associated variation in the gut bacterial communities of L. sticticalis and provide a descriptive basis for further investigation of diet–microbiota interactions. Because the study included only three pooled biological replicates per treatment and did not characterize plant-associated microbiota or plant chemistry, the observed associations and predicted functions should not be interpreted as evidence of specific microbial mechanisms or host adaptation.
