Short-term triphenyltin exposure alters microbial homeostasis in the silkworm (Bombyx mori) midgut.

Triphenyltin (TPT) is a widespread synthetic chemical used in many fields and its potential risk to organisms has been comprehensively investigated using different animal models and species. Currently, little is known about the effects of TPT exposure on microbial midgut diversity, therefore we explored these effects in the lepidopterous silkworm model using 16S rDNA sequencing. In total, 5273 and 5065 operational taxonomic units (OTUs) were identified in control and TPT-exposure group samples, ranging from 424 to 728 OTUs/sample. Alpha-diversity analyses revealed that TPT exposure induced the fluctuations of gut microbial diversity and abundance while beta-diversity analyses identified a distinct impact on major gut microbiota components. In our microbiome analyses, 23 phyla and 353 genera were recognized in the control group, while 20 phyla and 358 genera were recognized in the TPT exposure group. At the genus level, midgut microbiota were composed of several predominant bacterial genera, including Muribaculaceae, Lactobacillus, and UCG-010. In the TPT exposure group, o__Bacillales, f__Bacillaceae, and f__Caldicoprobacteraceae abundance was relatively high, while f__Oscillospiraceae, f__Fusobacteriaceae, and f__SC_I_84 abundance was relatively high in the control group. Gene function analyses in silkworm microbiota after TPT exposure showed that biosynthesis of ansamycins, fructose and mannose metabolism, glycerolipid metabolism, type II diabetes mellitus, glycolysis/gluconeogenesis, lipid metabolism, translation proteins, atrazine degradation, DNA repair and recombination proteins, nicotinate and nicotinamide metabolism were significantly increased. Collectively, our silkworm model identified gut microbial diversity risks and the adverse effects from TPT exposure, which were similar to other aquatic animals. Therefore, TPT levels in environmental samples must be monitored to prevent ecological harm.

The effects of polylactic acid bioplastic exposure on midgut microbiota and metabolite profiles in silkworm (Bombyx mori): An integrated multi-omics analysis.

Polylactic acid (PLA) is a highly common biodegradable plastic and a potential threat to health and the environment. However, limited data are available on the effects of PLA exposure in the silkworm (Bombyx mori), a model organism used in toxicity studies. In this study, silkworms with or without PLA exposure (P1: 1 mg/L, P5: 5 mg/L, P25: 25 mg/L, and P0: 0 mg/L) for the entire 5th instar period were used to investigate the impact of PLA exposure on midgut morphology, larvae growth, and survival. Mitochondrial damage was observed in the P5 and P25 groups. The weights of the P25 posterior silk gland (5th day in the 5th instar), mature larvae and pupae were all significantly lower than those of the controls (P < 0.05). Dead worm cocoon rates and larva-pupa to 5th instar larvae ratios showed a positive and negative dose-dependent manner with respect to PLA concentrations, respectively. Additionally, reactive oxygen species levels and superoxide dismutase activity of the P25 midgut were significantly higher and lower when compared with controls, respectively (P < 0.05). The molecular mechanisms underlying the effects of PLA and associated physiological responses were also investigated. In the midgut metabolome, 127 significantly different metabolites (variable importance projection >1 and P < 0.05) were identified between the P0 and P25 groups and were mainly enriched for amino acid metabolism and energy supply pathways. The 16 S rDNA data showed that PLA altered microbial richness and structural composition. Microbiota, classified into 34 genera and 63 species, were significantly altered after 25 mg/L PLA exposure (P < 0.05). Spearman's correlation results showed that Bifidobacterium catenulatum and Schaalia odontolytica played potentially vital roles during exposure, as they demonstrated stronger correlations with the significantly different metabolites than other bacterial species. In sum, PLA induced toxic effects on silkworms, especially on energy- and protein-relevant metabolism, but at high concentrations (25 mg/L). This prospective mechanistic investigation on the effects of PLA on larval toxicity provides novel insight regarding the ecological risks of biodegradable plastics in the environment.

Dietary administration with hydrolyzed silk sericin improves the intestinal health of diabetic rats.

Type II diabetes (T2D) is a global epidemic disease with an increased incidence and prevalence. Gut microbiota plays an important role in controlling T2D development. Dietary administration of prebiotics, probiotics, and drugs, including metformin, showed the regulatory impact on the change of gut microbiota, which is associated with the improvement of glucose tolerance. In this study, silk sericin was manufactured into hydrolyzed sericin peptide (HSP) powders as a dietary additive to investigate the effect on the gut microbiota of T2D model rats. The results indicated that the HSP-augmented dietary administration lowers the fast glucose level of diabetic rats, and HSP augmentation induces a change in the gut microbiota composition of T2D model rats toward the normal rats. Some key taxa, including Lactobacillus gasseri, were suggested to be involved in controlling T2D development. This finding provides new insight into developing sericin as functional food or therapeutic prebiotics against T2D in clinical practice.

Toxic effects of triphenyltin on the silkworm Bombyx mori as a lepidopterous insect model.

Triphenyltin (TPT) is a widely used reagent in various industries and agriculture, but is also known to accumulate in natural ecosystems and animal tissues. Hence, the aim of this study was to comprehensively assess the toxicity of TPT in the silkworm Bombyx mori as a model insect. The results showed that TPT exposure for the entire 5th instar larval stage significantly reduced the weight of silkworm pupa and inhibited development of the silkworm midgut. Following exposure to 2 µg/kg of TPT for 4 days, differentially expressed genes in midgut were associated with enriched pathways involved in the metabolism of carbohydrates, lipids, and amino acids, as determined by RNA sequencing. Furthermore, the metabolic profiles of the intestinal content of silkworms exposed to 2 µg/kg of TPT for 4 days were markedly altered and differential metabolites produced by metabolism of carbohydrates, lipids, and amino acids were enriched as determined by non-targeted GC-MS/MS metabolomics. This study provides novel insights into the mechanisms underlying the toxicity of TPT and emphasizes the risks posed by such pollutants released into the environment.

Multi-tissue metabolomic profiling reveals potential mechanisms of cocoon yield in silkworms (Bombyx mori) fed formula feed versus mulberry leaves.

Use of formula feed (FF) for silkworms for all instars, has promoted transformation and progress in traditional sericulture. However, the cocoon yield of FF silkworms has failed to reach that of silkworms fed mulberry leaves (ML). The biological mechanisms underlying this phenomenon have not been well described. This study aimed to identify metabolic mechanisms and potential biomarkers relating to the poor cocoon yield of FF silkworms. In this study, silkworms received treatments of either ML (ML group) or FF (FF group) for all instars. At the 3rd day of the 5th instar, the midgut (MG), hemolymph (HL) and posterior silk gland (PSG) were collected for the metabolome profiles detection. The remaining silkworms were fed ML or FF until cocooning for investigation. The whole cocoon yield (WCY) was significantly higher in the FF group than the ML group (p < 0.05), whereas the cocoon shell weight (CSW) and cocoon shell rate (CSR) were significantly lower in the FF group (p < 0.05). A total of 845, 867 and 831 metabolites were qualified and quantified in the MG, HL and PSG of the FF silkworms, respectively. Correspondingly, 789, 833 and 730 metabolites were quantified in above three tissues of the ML group. Further, 230, 249 and 304 significantly different metabolites (SDMs) were identified in the MG, HL and PSG between the FF and ML group, respectively. Eleven metabolic pathways enriched by the SDMs were mutual among the three tissues. Among them, cysteine and methionine metabolism, arginine biosynthesis, and arginine and proline metabolism were the top three pathways with the highest impact value in the PSG. Six biomarkers were obtained through biomarker analysis and Pearson correlation calculation. Among them, homocitrulline, glycitein, valyl-threonine, propyl gallate and 3-amino-2,3-dihydrobenzoic acid were positively correlated with WCY, but negatively correlated with CSW and CSR (p < 0.05). An opposite correlation pattern was observed between 3-dimethylallyl-4-hydroxyphenylpyruvate and the three cocoon performance traits. Overall, three key metabolic pathways and six biomarkers associated with cocoon yield were interpreted, and should provide directions for formula feed optimization in factory-raised silkworms.

Combined analysis of silk synthesis and hemolymph amino acid metabolism reveal key roles for glycine in increasing silkworm silk yields.

Rearing silkworms (Bombyx mori) using formula feed has revolutionized traditional mulberry feed strategies. However, low silk production efficiencies persist and have caused bottlenecks, hindering the industrial application of formula feed sericulture. Here, we investigated the effects of formula feed amino acid composition on silk yields. We showed that imbalanced amino acids reduced DNA proliferation, decreased Fib-H, Fib-L, and P25 gene expression, and caused mild autophagy in the posterior silk gland, reducing cocoon shell weight and ratio. When compared with mulberry leaves, Gly, Ala, Ser, and Tyr percentages of total amino acids in formula feed were decreased by 5.26%, while Glu and Arg percentages increased by 9.56%. These changes increased uric acid and several amino acids levels in the hemolymph of silkworms on formula feed. Further analyses showed that Gly and Thr (important synthetic Gly sources) increased silk yields, with Gly increasing amino acid conversion efficiencies to silk protein, and reducing urea levels in hemolymph. Also, Gly promoted endomitotic DNA synthesis in silk gland cells via phosphoinositide 3-kinase (PI3K)/Akt/target of rapamycin (TOR) signaling. In this study, we highlighted the important role of Gly in regulating silk yields in silkworms.

Germline transformation of the silkworm Bombyx mori L. by sperm-mediated gene transfer.

The domesticated silkworm Bombyx mori L. has important roles in basic biological research and applied science. To explore the practical use of transgenic technology in agricultural silkworm varieties, we fused the neomycin-resistance gene (Neo(R)) and the green fluorescent protein gene (gfp) into the piggyBac-based transposon vector and transduced it into silkworms by sperm-mediated gene transfer (SMGT). Fluorescence observation indicated the positive rate of G0 egg-batches is 72.7%. After screening against the antibiotic G418, development of individual larvae in the same brood showed significant size differences. PCR detection indicated the existence of gfp and Neo(R) and confirmed the positive rate of transgenesis as 0.47%. Southern blot analysis confirmed the presence of the exogenous genes in the genome of G7 larvae. These results show that our strategy is practical and markedly improves the efficiency of SMGT.