Study On The Microbial Diversity And Enzymatic Activity Of Omphisa Fuscidentalis Hampson

Cellulosic biomass is the most abundant natural resource in the biosphere,the conversion of these resources into energy has attracted more and more attention in recent year.However,the application and production of cellulase are restricted by the limitation of cellulase activity and its high cost.More and more attention from reseachers has been focused on finding novel cellulose degrading microorganism,obtaining resources of cellulase with high yield and improving the efficiency of cellulase.From the studies on the degrading cellulose by gut microorganisms of insects,it was found that the microorganisms in the gut of insects could be good resources of isolating cellulose degrading microorganisms,genes and enzyme.In this paper,we study on the gut microbes of Omphisa fuscidentalis Hampson by High-throughput sequencing,and isolated cellulose degrading strains from Hampson gut.Main conclusions were as follows:(1)The high-throughput sequencing results showed that bacteria at Phylum level were Proteobacteria,Bacteroidetes and Firmicutes,at genus level are Commensalibacter,genus Dysgonomonas,Acinetobacter and so on.Fungus at Phylum level was dominated by Ascomycota,at genus level were Phaeoacremonium,Ophiosphaerella,Acremonium and Myrothecium.It was found that there are a lot of potential microbial resources for cellulose degradation.(2)The metagenome sequencing of Omphisa fuscidentalis Hampson gut microbes were carried out.By annoted with the database KEGG,eggNOG and CAZy,we got the data of metabolic enzymes encoding genes and carbohydrate enzymes of Hampson gut microbes.The analysis showed that 344glycoside hydrolase encoding genes were from 56 different families,compared with the giant panda,tammar wallabies,wood feeding termite gut microbes and higher rumen microbial metagenomic sequencing samples,there are 13 specific GH family genes in the gut of Omphisa fuscidentalis Hampson,and it is abundant in genes ecoding enzymes of the degradation of hemicellulose and oligosaccharides.(3)54 strains were isolated from Hampson gut,and determined endo-1,4-?-D-glucanohydrolase,exo-1,4-?-D-glucannase and?-1,4-glucosidase of 10 strains,named C-41,C-25,C-24,C-17,C-13,C-5,W-26,W-17,W-9,W-3 by DNS method.It was found that these isolates were mainly from Proteobacteria,Bacteroides,and Firmicutes by the NJ phylogenetic trees.(4)Strain BRTC-1 belongs to the genus Acinetobacter,the 16S rRNA,rpoB and gyrB sequences of GenBank accession number were KU560468.2,KU837228,KU981067.Strain BRTC-1 was identified as novel Acinetobacter species by polyphase taxonomy methods which was designated as Acinetobacter larvae.The type strain is available in Agricultural Culture Collection of China(ACCC19936)and Japan Collection of Microorganisms(JCM 31367).(5)The whole genome sequencing of the strain Acinetobacter larvae BRTC-1~T(NCBI accession number:CP016895.1)was completed by PacBio sequencing,the plasmids pRW1 and pRW2accession numbers were CP018141.1 and CP018142.1,respectively.The genome harbors 3,741,098bp,with an average G+C content of 41.64%and contains 3,325 predicted CDSs and 102 ncRNA genes.The plasmids pRW1 and pRW2 sequences were 12,170 bp and 8,455 bp,with an average GC content of 33.86%and 34.54%,respectively.From the analysis of genome we found that multiple genes for encoding carbohydrate-active enzymes.The genome information will pave the way for further elucidation of cellulose degrading mechanism and wider exploitation of functional genes.(6)The Native-Page of crude enzymes showed that cellulase in Hampson gut were endo-1,4-?-D-glucano hydrolase,exo-1,4-?-D-glucannase and xylanase.The filter paper degradation experiments verify the ability of cellulose degradation by Hampson gut microbes independently.High-throughput sequencing showed there were lots of cellulose degrading microbes in the MMP medium.This study will help us understanding microorganism in Hampson,and given a basis for exploiting cellulose degrading microorganism form Hampson.