Use of tuf as a target for sequence-based identification of Gram-positive cocci of the genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus

Xuerui Li,Baoyu Li,Juan Xing,Pu Wang,Jixing Liu

doi:10.1186/1476-0711-11-31

Abstract

BackgroundAccurate identification of isolates belonging to genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations,especially when large blind samples must be analyzed. It is useful to simultaneously identify species in different genera using a single primer pair.MethodsWe developed a primer pair based on the tuf gene (encoding elongation factor) sequence to identify 56 Gram-positive cocci isolates.ResultsThe target sequences were amplified from all 56 samples. The sequencing results and the phylogenetic tree derived from the partial tuf gene sequences identified the isolates as three enterococcal species, two lactococcal species, two staphylococcal species, and six streptococcal species, as well as eight isolates that were novel species of the genus Streptococcus. Partial gene sequence analysis of the sodA, dnaK, and 16S RNA genes confirmed the results obtained by tuf gene sequencing.ConclusionBased on the uniform amplification of the tuf gene from all samples and the ability to identify all isolates at both the genus and species levels, we conclude that the primer pair developed in this research provides a powerful tool for identifying these organisms in clinical laboratories where large blind samples are used.

Highlights

Accurate identification of isolates belonging to genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations,especially when large blind samples must be analyzed
We developed a simple tuf gene based polymerase chain reaction (PCR) and sequencing assay to identify isolates belonging to the genera Enterococcus, Staphylococcus, Streptococcus, and Lactococcus
Bacterial strains The strains examined in this study included Streptococcus suis (S.suis)serotype 2 strain 9801, S. suis 05ZYH33, one Lactococcus lactis subsp. lactis isolate, 48 cocci isolates that was isolated by our lab between 2006 and 2011, and four cocci strains purchased from the Chinese General Microbiology Culture Collection Center (CGMCC), which included the following: Streptococcus pernyi (S.peryi) CGMCC1.1010, Streptococcus salivarius CGMCC 1.2498, Streptococcus mutans CGMCC 1.2499, and Streptococcus bovis CGMCC1.2502

Summary

Introduction

Accurate identification of isolates belonging to genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations,especially when large blind samples must be analyzed. Lactococcus is a genus of lactic acid bacteria that were formerly included in the Accurate identification of isolates belonging to these genus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations. RpoB gene-based PCR sequencing has been used for accurate detection and identification of species in the genera Streptococcus, Enterococcus, Gemella, Abiotrophia, Granulicatella [11], and Staphylococcus [16]. The groES and groEL genes have been used as a target for identification of Abiotrophia, Granulicatella, and Gemella species [12]

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