Species identification and phylogenetic relationships based on partial HSP60 gene sequences within the genus Staphylococcus

Abstract

The phylogenetic relationships among 36 validly described species or subspecies within the genus Staphylococcus were investigated by cloning and sequencing their 60 kDa heat-shock protein (HSP6O) genes using a set of universal degenerate HSP6O PCR primers. The cloned partial HSP60 DNA sequences f rom nine Staphylococcus aureus strains were highly conserved (97-100% DNA sequence similarity; mean 98O/0), indicating that the HSP60 gene of multiple isolates within the same species have little microheterogeneity. A t the subspecies level, DNA sequence similarity among members of 5. aureus, Staphylococcus schleiferi, Staphylococcus cohnii and Staphylococcus capitis ranged from 91 to 98%. A t the interspecies level, sequence similarity among 23 distinct species of staphylococci ranged from 74 to 93 O/ O (mean 82 YO). By comparison, the highest sequence similarity of Bacillus subtilis and Escherichia coli with members within the genus Staphylococcus was only 70 and 59 %, respectively. Importantly, phylogenetic analysis based on the neighbour-joining distance method revealed remarkable concordance between the tree derived from partial HSP60 gene sequences and that based on genomic DNA-DNA hybridization, while 165 rRNA gene sequences correlated less well. The results demonstrate that DNA sequences from the highly conserved and ubiquitous HSP6O gene offer a convenient and accurate tool for species-specif ic identification and phylogenetic analysis of staphylococci.

Identification of Coagulase-Negative Staphylococci by SodA Gene Sequence Analysis

“…The 16S rDNA sequence similarity has been shown to be very high at 90-99% in 29 Staphylococcus species studied (Kwok et al, 1999). A sequence similarity of 95 and 99% was seen between S. caprae and S. capitis using the 16S rRNA and hsp60 gene, respectively (Ghebremedhin et al, 2008) while the partial rpoB sequences of these two species were lower at 84.5% (Drancourt and Raoult, 2002) as compared to 94% similarity observed in the present study.…”

“…A sequence similarity of 95 and 99% was seen between S. caprae and S. capitis using the 16S rRNA and hsp60 gene, respectively (Ghebremedhin et al, 2008) while the partial rpoB sequences of these two species were lower at 84.5% (Drancourt and Raoult, 2002) as compared to 94% similarity observed in the present study. Staphylococcus sciuri and S. vitulinus species showed 90.1-98.6% similarity in their hsp60 gene sequences (Kwok and Chow, 2003;Kwok et al, 1999) while a similarity of 94% between these species were detected in the present study. Similarly, the sodA sequence revealed a similarity of 89% between S. simulans and S. carnosus while the sequence similarity was 87, 95 and 96% with rpoB, gap and 16s rRNA gene sequence, respectively (Ghebremedhin et al, 2008).…”

“…In general, the results suggest that apart from the subspecies level, the sodA internal sequence is able to show more species discrimination as compared to the sequence of other conserved genes such as the 16S rRNA with mean similarity of 90% (Shah et al, 2007), rpoB with mean similarity of 86% (Drancourt and Raoult, 2002;Mellmann et al, 2006) and hsp60 with mean similarity of 82% (Kwok et al, 1999). Following general convention, a 95% identity threshold was adopted as the limit for a positive identification and unambiguous assignment to a particular species.…”

“…For example, dnaJ gene sequence displayed a similarity of 90.9% between S. condimenti and S. carnosus as compared to 97% similarity observed in the present study. At subspecies level, with 7% divergence, dnaJ (Shah et al, 2007) and hsp60 (Kwok et al, 1999) gene sequences can be used to discriminate S. cohnii subsp. cohnii from S. cohnii subsp.…”

“…However, the use of this gene is limited due to high degree of sequence similarity between closely related species (Becker et al, 2004;Goh et al, 1996;Gribaldo et al, 1997;Kwok et al, 1999). Hence, other highly conserved sequence has been exploited which includes sodA (Poyart et al, 2001), gap (Bergeron et al, 2011;Ghebremedhin et al, 2008), hsp60 (Ghebremedhin et al, 2008;Kwok et al, 1999), rpoB (Drancourt and Raoult, 2002;Mellmann et al, 2006), tuf (Heikens et al, 2005) and dnaJ (Shah et al, 2007) gene sequencing methods. However, the sequences of some genes are not sufficient to discriminate between closely related Staphylococcus at both species and subspecies level and the databases only includes a limited number of species.…”