Dataset Information

High Genetic Diversity of Microbial Cellulase and Hemicellulase Genes in the Hindgut of Holotrichia parallela Larvae.

ABSTRACT: In this study, we used a culture-independent method based on library construction and sequencing to analyze the genetic diversity of the cellulase and hemicellulase genes of the bacterial community resident in the hindgut of Holotrichia parallela larvae. The results indicate that there is a large, diverse set of bacterial genes encoding lignocellulose hydrolysis enzymes in the hindgut of H. parallela. The total of 101 distinct gene fragments (similarity <95%) of glycosyl hydrolase families including GH2 (24 genes), GH8 (27 genes), GH10 (19 genes), GH11 (14 genes) and GH36 (17 genes) families was retrieved, and certain sequences of GH2 (10.61%), GH8 (3.33%), and GH11 (18.42%) families had <60% identities with known sequences in GenBank, indicating their novelty. Based on phylogenetic analysis, sequences from hemicellulase families were related to enzymes from Bacteroidetes and Firmicutes. Fragments from cellulase family were most associated with the phylum of Proteobacteria. Furthermore, a full-length endo-xylanase gene was obtained, and the enzyme exhibited activity over a broad range of pH levels. Our results indicate that there are large number of cellulolytic and xylanolytic bacteria in the hindgut of H. parallela larvae, and these symbiotic bacteria play an important role in the degradation of roots and other organic matter for the host insect.

SUBMITTER: Sheng P

PROVIDER: S-EPMC4519965 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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High Genetic Diversity of Microbial Cellulase and Hemicellulase Genes in the Hindgut of Holotrichia parallela Larvae.

Sheng Ping P Li Yushan Y Marshall Sean D G SD Zhang Hongyu H

International journal of molecular sciences 20150721 7

In this study, we used a culture-independent method based on library construction and sequencing to analyze the genetic diversity of the cellulase and hemicellulase genes of the bacterial community resident in the hindgut of Holotrichia parallela larvae. The results indicate that there is a large, diverse set of bacterial genes encoding lignocellulose hydrolysis enzymes in the hindgut of H. parallela. The total of 101 distinct gene fragments (similarity <95%) of glycosyl hydrolase families inclu ...[more]

PMID: 26197317

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Project description:BackgroundGoat rumen microbial communities are perceived as one of the most potential biochemical reservoirs of multi-functional enzymes, which are applicable to enhance wide array of bioprocesses such as the hydrolysis of cellulose and hemi-cellulose into fermentable sugar for biofuel and other value-added biochemical production. Even though, the limited understanding of rumen microbial genetic diversity and the absence of effective screening culture methods have impeded the full utilization of these potential enzymes. In this study, we applied culture independent metagenomics sequencing approach to isolate, and identify microbial communities in goat rumen, meanwhile, clone and functionally characterize novel cellulase and xylanase genes in goat rumen bacterial communities.ResultsBacterial DNA samples were extracted from goat rumen fluid. Three genomic libraries were sequenced using Illumina HiSeq 2000 for paired-end 100-bp (PE100) and Illumina HiSeq 2500 for paired-end 125-bp (PE125). A total of 435gb raw reads were generated. Taxonomic analysis using Graphlan revealed that Fibrobacter, Prevotella, and Ruminococcus are the most abundant genera of bacteria in goat rumen. SPAdes assembly and prodigal annotation were performed. The contigs were also annotated using the DOE-JGI pipeline. In total, 117,502 CAZymes, comprising endoglucanases, exoglucanases, beta-glucosidases, xylosidases, and xylanases, were detected in all three samples. Two genes with predicted cellulolytic/xylanolytic activities were cloned and expressed in E. coli BL21(DE3). The endoglucanases and xylanase enzymatic activities of the recombinant proteins were confirmed using substrate plate assay and dinitrosalicylic acid (DNS) analysis. The 3D structures of endoglucanase A and endo-1,4-beta xylanase was predicted using the Swiss Model. Based on the 3D structure analysis, the two enzymes isolated from goat's rumen metagenome are unique with only 56-59% similarities to those homologous proteins in protein data bank (PDB) meanwhile, the structures of the enzymes also displayed greater stability, and higher catalytic activity.ConclusionsIn summary, this study provided the database resources of bacterial metagenomes from goat's rumen fluid, including gene sequences with annotated functions and methods for gene isolation and over-expression of cellulolytic enzymes; and a wealth of genes in the metabolic pathways affecting food and nutrition of ruminant animals.

Dataset Information

High Genetic Diversity of Microbial Cellulase and Hemicellulase Genes in the Hindgut of Holotrichia parallela Larvae.

Publications

High Genetic Diversity of Microbial Cellulase and Hemicellulase Genes in the Hindgut of Holotrichia parallela Larvae.

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