Analysis of intestinal bacterial community diversity of adult Dastarcus helophoroides

Z Q Zhang,M L Li,C He,Allen Cohen

doi:10.1093/jis/14.1.114

Abstract

Polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE), and a culture-dependent technique were used to study the diversity of the intestinal bacterial community in adult Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae). Universal bacterial primers targeting 200 bp regions of the 16S rDNA gene were used in the PCR-DGGE assay, and 14 bright bands were obtained. The intestinal bacteria detected by PCR-DGGE were classified to Enterococcus (Lactobacillales: Enterococcaceae), Bacillus (Bacillales: Bacillaceae), Cellvibrio (Pseudomonadales: Pseudomonadaceae), Caulobacter (Caulobacterales: Caulobacteraceae), and uncultured bacteria, whereas those isolated by the culture-dependent technique belonged to Staphylococcus (Bacillales: Staphylococcaceae), Pectobacterium Enterobacteriales: Enterobacteriaceae), and Enterobacter (Enterobacteriales: Enterobacteriaceae). These intestinal bacteria represented the groups Lactobacillales ( Enterococcus ), Pseudomonadales ( Cellvibrio ), Caulobacterales ( Caulobacter ), Bacilli ( Bacillus and Staphylococcus ), and Gammaproteobacteria ( Pectobacterium and Enterobacter ). Our results demonstrated that PCR-DGGE analysis and the culture-dependent technique were useful in determining the intestinal bacteria of D. helophoroides and the two methods should be integrated to characterize the microbial community and diversity.

Highlights

Many insects are inhabited by diverse communities of microorganisms
The DGGE bands of D. helophoroides intestinal bacteria are shown in Fig. 1, and the results of the phylogenetic analysis are shown in Fig. 2, revealing the presence of a variety of different genera
Table 1. 16S rDNA sequences of Dastarcus helophoroides intestinal bacteria identified by the PCR-DGGE method

Summary

Introduction

Many insects are inhabited by diverse communities of microorganisms. It is possible that the number of microbes in most insects is larger than the number of somatic cells (Campbell 1990). The intestinal microbial bacteria in insects have been shown to play important roles, such as providing vitamins, aiding in fat and carbohydrate metabolism, preventing the invasion of external bacteria, and promoting the function of the immune system (Eutick et al 1978, Abe et al 2000, Suchodolski and Ruaux 2004). The analysis of intestinal bacteria was performed by culturedependent and molecular methods. The culture-dependent technique defined the gut microbes by phenotypic characterization (morphology, immunology, and physiological-biochemical reaction), but unculturable bacteria were largely ignored (Lysenko 1985). Molecular biology methods allowed extracting the total genomic DNA of bacteria directly from samples and sequencing and analyzing the DNA to characterize the bacteria species composition and abundance (Yu et al 2008)

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