Advantages of multilocus sequence analysis for taxonomic studies: a case study using 10 housekeeping genes in the genus Ensifer (including former Sinorhizobium)

Abstract

There is a need for easy, practical, reliable and robust techniques for the identification and classification of bacterial isolates to the species level as alternatives to 16S rRNA gene sequence analysis and DNA-DNA hybridization. Here, we demonstrate that multilocus sequence analysis (MLSA) of housekeeping genes is a valuable alternative technique. An MLSA study of 10 housekeeping genes (atpD, dnaK, gap, glnA, gltA, gyrB, pnp, recA, rpoB and thrC) was performed on 34 representatives of the genus Ensifer. Genetic analysis and comparison with 16S and 23S rRNA gene sequences demonstrated clear species boundaries and a higher discrimination potential for all housekeeping genes. Comparison of housekeeping gene sequence data with DNA-DNA reassociation data revealed good correlation at the intraspecies level, but indicated that housekeeping gene sequencing is superior to DNA-DNA hybridization for the assessment of genetic relatedness between Ensifer species. Our MLSA data, confirmed by DNA-DNA hybridizations, support the suggestion that Ensifer xinjiangensis is a later heterotypic synonym of Ensifer fredii.

DNA barcoding of the fungal genus Neonectria and the discovery of two new species

“…The aligned sequences of each gene and the combined EF-1α and RPB2 genes were analyzed using DNAstar 7.1.0 (Lasergene, USA) to calculate the similarity matrices and then illustrate the intra-and inter-specific variations of the candidate barcode loci for each of the 19 species tested in this study, using a visualization analysis tool, TaxonGap 2.4.1 [42]. As suggested by Martens et al [43], Nectria pseudotrichia was designated as the outgroup in the analysis.…”

Vibrio inhibens sp. nov., a novel bacterium with inhibitory activity against Vibrio species

“…Previous studies have demonstrated that multilocus sequencing analysis has a higher discriminatory power than DNA-DNA hybridization. 27,28 In fact, Thompson et al 29 have reported that when two Vibrio strains share more than 95% similarity by multilocus sequencing analysis, they are the same species. These results demonstrate that strain BFLP-10 T is distinct from any recognized species of the genus Vibrio.…”

Ribosomal protein biomarkers provide root nodule bacterial identification by MALDI-TOF MS

“…Sequence analysis of the 16S ribosomal RNA (rRNA) gene allows the construction of valid bacterial phylogenies (Schleifer 2009), but lacks resolving power at and below the species level, due to the extent of gene conservation (Willems 2006). Multilocus sequence analysis (MLSA) of conserved core genes was proposed as a robust and reliable alternative method for classifying bacteria to species level (Gevers et al 2005;Hanage et al 2006) and has been successfully used to delineate species within a number of genera including Ensifer, formerly Sinorhizobium (Martens et al 2008), and Bradyrhizobium (Rivas et al 2009). However, incongruent phylogenies have been reported among several genes commonly used in MLSA, along with evidence of lateral gene transfer or intergenic recombination, resulting in unclear taxonomic resolution Zhang et al 2012).…”