Effects Of Mulberry Leaf Surface Moisture On Growth And Gut Microbiota Of Bombyx Mori

Silkworm,Bombyx mori Linnaeus,is a phytophagous insect.Its growth and development are closely related to environmental humidity.At present,antibiotics are used to prevent the disease of silkworm during the rainy season in the large-scale breeding of silkworm.The excessive use of antibiotics will cause microbiota imbalance and environmental pollution.In addition,B.mori is a model insect in Lepidoptera.Studying the effects of water factors on B.mori can also provide new ideas for the prevention and control of Lepidoptera pests.In this study,we found the mortality rate and growth development have been significantly affected by feeding wet mulberry leaves.The relationships between water factors and insect development were analyzed by tissue sections,16 S r DNA and ITS sequencing of intestinal microbiota,and activity determination of major digestive enzymes.1.The physiological indexes of silkworms feeding wet mulberry leaves were recorded and analyzed.The survival rate of silkworm decreased rapidly on the fourth day after feeding wet mulberry leaves,and the fourth day was the third molting period.Until the thirteenth day,the average survival rate was only 13.33%.Secondly,the developmental period of the third and fourth instar larvae was significantly delayed.The development period of the third instar was delayed from 3.13±0.01 d to 3.72±0.12 d;the fourth instar was delayed from 4.92±0.05 d to 5.53±0.09 d.The body weight of the fifth instar silkworm significantly decreased after feeding wet mulberry leaves,but the pupa weight was not significantly less than control group.After feeding wet mulberry leaves,the cocoon weight and fecundity of B.mori were significantly decreased,the average cocoon weight decreased from 0.05±0.00 g to 0.04±0.00 g,and the average fecundity decreased from278.21±11.07 eggs to 224.80±18.29 eggs.The above results show that moisture factors can affect the biological characteristics of silkworm.2.According to the results of the silkworm midgut paraffin section,it was found that the peritrophic membrane becomes small in space,loosely structured,and not close to the epithelial cells after feeding wet mulberry leaves at B.mori.These results indicate that feeding wet mulberry leaves could change the morphological structure of the main digestive organ in the silkworm.3.There were significant differences in the abundance of gut microbiota after silkworm feeding wet mulberry leaves.In the composition of gut bacteria,the abundance of Proteobacteria was significantly increased,and the abundance of Firmicutes and Cyanobacteria was significantly decreased.The abundance changes increased or decreased with the increase of larval instars.At the genus level,the abundance of Staphylococcus significantly decreased,and the abundance change decreased with the increase of larval instars.Staphylococcus is the most distributed biomarker taxa in the third and fourth instar larvae of B.mori.The abundance of Acinetobacter was only significantly increased in the fifth instar larvae,with an abundance of 10.24%,and the abundance of Bacillus was only significantly increased in the third instar,with an abundance of 7.55%.In the composition of gut fungi,the abundances of Ascomycota and Basidiomycota were significantly changed,and the abundance changes between them were opposite during the larval instar.At the genus level,the abundances of Malassezia,Aspergillus,Sympodiomycopsis and Jaminaea significantly changed during different instars of B.mori.In addition,Buckleyzyma was the biomarker taxa in the gut fungi of the third and fourth instar larvae of silkworm.Feeding wet mulberry leaves had the most significant effect on the intestinal microbiota of the fourth instar,significantly down-regulated the functions related to material metabolism in intestinal bacteria,and significantly up-regulated the functions related to nucleobase biosynthesis in intestinal fungi.4.By measuring the activity of digestive enzymes in the midgut of silkworm at different instars.The results showed that there was no significant difference in the activities of lipase and α-amylase in the third instar silkworm after feeding wet mulberry leaves,but the enzyme activities were significantly decreased in the midgut of the fourth and fifth instar larvae.The activity of trypsin was significantly decreased,and the activity of trehalase was significantly increased during different instars of B.mori.It is concluded that the digestive enzyme activity of silkworm feeding on wet mulberry leaves is basically consistent with the change in enzyme-producing bacteria abundance.This provides a basis for exploring the internal mechanism of the relationship between water factors and insect development.