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Lepidoptera (butterflies and moths) is a major insect order including important pollinators and agricultural pests, however their microbiomes are little studied. Here, using next-generation sequencing (NGS)-based shotgun metagenomics, we characterize both the biodiversity and functional potential of gut microbiota of a lepidopteran model insect, the silkworm Bombyx mori. Two metagenomes, including the standard inbred strain Dazao (P50) and an improved hybrid strain Qiufeng × Baiyu (QB) widely used in commercial silk production, were generated, containing 45,505,084 and 69,127,002 raw reads, respectively. Taxonomic analysis revealed that a total of 663 bacterial species were identified in P50 silkworms, while 322 unique species in QB silkworms. Notably, Enterobacter, Acinetobacter and Enterococcus were dominated in both strains. The further functional annotation was performed by both BlastP and MG-RAST against various databases including Nr, COG, KEGG, CAZy and SignalP, which revealed >5 × 106 protein-coding genes. These datasets not only provide first insights into all bacterial genes in silkworm guts, but also help to generate hypotheses for subsequently testing functional traits of gut microbiota in an important insect group.

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BACKGROUND Many insect pests rely on microbial symbionts to obtain nutrients or for defence, thereby allowing them to exploit novel food sources and degrade environmental xenobiotics, including pesticides. Although Lepidoptera is one of the most diverse insect taxa and includes important agricultural pests, lepidopteran microbiotas, particularly functional traits, have not been studied widely. Here, we provide a comprehensive characterization of the gut microbiota across multiple mulberry‐feeding lepidopteran species, resolving both community structure and metabolic potential. RESULTS Our results indicate abundant bacteria inside the gut of larval Lepidoptera. However, even though they were fed the same diet, the structures of the bacterial communities differed in four major mulberry pest species, suggesting host‐specific effects on microbial associations. Community‐level metabolic reconstructions further showed that although taxonomic composition varied greatly, carbohydrate and amino acid metabolism and membrane transporter were key functional capabilities of the gut bacteria in all samples, which may play basic roles in the larval gut. In addition, principal coordinate analysis (PCoA) of gut bacterial‐predicted gene ontologies revealed specialized features of the microbiota associated with these mulberry pests, which were divided into two distinct clusters (macrolepidopterans and microlepidopterans). This pattern became even more prominent when further Lepidoptera species were involved. CONCLUSIONS A suite of gut microbiota metabolic functions significantly correlated with larval size; the metabolism of terpenoids and polyketides, xenobiotics biodegradation and metabolism were specifically enriched in large species, while small larvae had enhanced nucleotide metabolism. Our report paves the way for uncovering the correlation between host phenotype and microbial symbiosis in this notorious insect pest group. © 2019 Society of Chemical Industry

Mulberry (Morus) is an economically important woody tree that is suitable for use in sericulture as forage and in medicine. However, this broad-leaved tree is facing multiple threats ranging from phytopathogens to insect pests. Here, a Gram-positive, endospore-forming bacterium (ZJU1) was frequently isolated from healthy mulberry plants by screening for foliar endophytes showing antagonism against pathogens and pests. Whole-genome sequencing and annotation resulted in a genome size of 4.06 Mb and classified the bacterium as a novel strain of Bacillus amyloliquefaciens that has rarely been identified from tree leaves. An integrative approach combining traditional natural product chemistry, activity bioassays, and high-resolution mass spectrometry confirmed that strain ZJU1 uses a blend of antimicrobials including peptides and volatile organic compounds to oppose Botrytis cinerea, a major phytopathogenic fungus causing mulberry gray mold disease. We showed that the inoculation of endophyte-free plants with ZJU1 significantly decreased both leaf necrosis and mortality under field conditions. In addition to the direct interactions of endophytes with foliar pathogens, in planta studies suggested that the inoculation of endophytes also induced plant systemic defense, including high expression levels of mulberry disease resistance genes. Moreover, when applied to the generalist herbivore Spodoptera litura, ZJU1 was sufficient to reduce the pest survival rate below 50%. A previously undiscovered crystal toxin (Cry10Aa) could contribute to this insecticidal effect against notorious lepidopteran pests. These unique traits clearly demonstrate that B. amyloliquefaciens ZJU1 is promising for the development of successful strategies for biocontrol applications. The search for new plant-beneficial microbes and engineering microbiomes is therefore of great significance for sustainably improving plant performance.