Diversity Of Intestinal Bacteria Of Black Soldier Fly Larvae And Their Effects On The Growth And Development Of Black Soldier Fly

Hermetia illucens L.,also known as black soldier fly(BSF),is native to America and widely distributed in many countries over the world.The living environment of BSF in nature often contains a large number of microorganisms.Researches show that BSF larvae are used to treat food waste and livestock manure.Treating food waste and livestock manure with BSF can significantly improve the nutritional structure and microbial community structure of substrate and inhibit the growth of pathogenic bacteria.BSF larvae are also high-quality protein and fat sources,which can be used as feed additives to feed poultry and livestock.BSF can also produce antimicrobial peptides.Intestinal microorganisms can affect the growth and development of insects by decomposing food,stimulating host immune system,regulating intestinal metabolism and host gene expression and so on.So far,the researches of BSF mainly focused on agricultural application,the effects of intestinal microorganisms on its growth and development are still unclear.In this study,16S rDNA pyrosequencing technology was used to analyze the intestinal microorganisms of different instars of BSF larvae,intestinal bacteria of BSF were isolated and cultured.The effects of six intestinal bacteria on the growth and development of BSF were studied with germfree BSF larvae system.The main results are as follows.(1)Diversity of intestinal bacteria in different instars of BSF larvae16S rDNA pyrosequencing was used to investigate the composition of intestinal microbial community and its dynamic change of 1～6 instar BSF larvae.The results showed that:Proteobacteria,Firmicutes,Actinobacteria and Bacteroidetes were dominant in the intestine of each instar larvae at the phylum level.Dysgononadaceae,Enterobacteria,Enterococceae,Moraxellaceae,Lactobacillus,Weeksellaceae were dominant in the intestinal tract of each instar larvae at the family level.Providencia,Dysgonomonas,Klebsiella,Enterococcus existed in the intestinal tract of each instar larvae with high abundance at the genus level.The abundance and proportion of Klebsiella in the intestinal tract decreased with the increase of instar,while Providencia in the sixth instar larvae accounted for 47.22%(2)Isolation,purification and preliminary analysis of intestinal bacteria of BSFIn total,78 bacterial strains from 16 genus were isolated by using EG,1/3 TSB and LB media under anaerobic or aerobic conditions.Six strains of bacteria,BSF1(Providencia),BSF2(Citrobacter),BSF3(Klebsiella),BSF4(Proteus),BSF5(Ochrobactrum)and BSF6(Dysgonomonas),were selected for the further experiment.The growth curves of the six bacteria at the fit growth temperature(30℃)were evaluated.The extracellular enzyme activities and carbohydrate fermentation ability of the six strains were systematically analyzed by means of API-ZYM and API-50CH strip identification system.(3)Effects of intestinal bacteria on the growth and development of BSFGermfree larvae of BSF were obtained by disinfecting the surface of eggs.The growth and development data of germfree BSF larvae,single strain supplemented larvae and larvae without disinfection treatment were collected to analyze the effects of these six bacteria on the growth and development of BSF.The results showed that these strains could stably exist in the intestines of BSF larvae after disinfection.Compared with the sterile BSF larvae,the control group with egg surface bacteria could significantly shorten the growth cycle of BSF,gain weight of larvae and pupae,increase the percentage of eclosion.Among them,BSF2 and BSF3 could significantly promote the weight of the larvae,BSF1 could increase the percentage of eclosion,BSF4 had no significant effect on the growth and development of BSF larvae,while the proportion of the larvae entering the pupal stage was significantly reduced.