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

Ping Sheng,Yushan Li,Hongyu Zhang,Sean Marshall

doi:10.3390/ijms160716545

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.

Highlights

The family Scarabaeidae is currently defined as comprising over 30,000 species of beetles, which are almost exclusively herbivorous or saprophagous [1]
We report on a phylogenetic analysis of bacterial cellulase and hemicellulase genes cloned from the hindgut of H. parallela larvae using degenerate primers
Cellulase and hemicellulase genes originating from symbiotic bacteria residing in the phytophagous H. parallela larval hindgut were targeted for diversity analysis because of their synergistic roles in lignocelluloses degradation

Summary

Introduction

The family Scarabaeidae is currently defined as comprising over 30,000 species of beetles, which are almost exclusively herbivorous or saprophagous [1]. Many scarab larvae live in the soil and feed on peanut, plant roots and other organic matters [2]. In China, the phytophagous scarab H. parallela larva is a severe peanut and sweet potato crops pest [3]. The hindgut of scarab larvae is similar to that of the wooding-feed termite: It is enlarged and houses diverse microbes [2,4]. Previous studies have shown that 25%–65% of the ingested pure cellulose or neutral detergent fibers in scarab larvae’s diet are degraded by scarab larvae and their intestinal bacteria [4,5]. Cellulolytic and hemicellulolytic bacteria have been isolated from the hindgut of some scarab species [6,7]. The scarab gut may be a potential source of bioreactor activities for bio-fuel production [1]

Methods

Results

Conclusion