Multiple-Locus Variable-Number Tandem-Repeat Analysis of Salmonella enterica Serovar Typhi

Abstract

Multilocus variable-number tandem repeats (VNTRs) are widely used as molecular markers to differentiate isolates of homogenous pathogenic clones. We explored the genomes of Salmonella enterica serovar Typhi strains CT18 and Ty2 for potential VNTRs. Among the 43 potential VNTRs screened, 2 were found to be polymorphic. Together with seven polymorphic VNTRs from previous studies, they were used to type 73 global serovar Typhi isolates. A total of 70 multilocus VNTR analysis (MLVA) profiles were found, distinguishing all except three pairs of isolates into individual profiles. The discriminatory power was 0.999. Phylogenetic analysis showed that the MLVA profiles can be divided into seven clusters. However, except for the closely related isolates, the relationships derived were in conflict with those inferred from single nucleotide polymorphism (SNP) typing using 38 SNPs done previously. We concluded that MLVA can resolve the relationships only among closely related isolates. A combination of SNP typing and MLVA typing offers the best approach for local and global epidemiology and the evolutionary analysis of serovar Typhi. We suggest that seven of the nine most polymorphic VNTRs be used as a standardized typing scheme for epidemiological typing.

Use of multilocus variable-number tandem repeat analysis in molecular subtyping of Salmonella enterica serovar Typhi isolates

“…Four loci (Sty2, Sty3, Sty39 and Sty42) had low diversity values. The allelic diversity of all VNTRs, with the exception of Sty2, Sty3 and Sty42, were previously evaluated using 73 global S. Typhi isolates (Octavia & Lan, 2009) and the diversity levels observed for the VNTRs were similar to those described in this study.…”

“…Based on the analyses of a small set of highly variable loci, MLVA is sufficient to replace PFGE in resolving closely (Liu et al, 2003); STTR5 (Lindstedt et al, 2003); Sal02, Sal06 and Sal20 (Ramisse et al, 2004); TR4500 and TR4699 (Octavia & Lan, 2009). DThe range of repeat units for allele 173 of Sty20, which has a 7 bp deletion at the 59-flanking region immediately adjacent to the repeats, is reported instead of that for the fragment size (bp).…”

“…The primer screening process selected 34 TRs that were easily amplified by PCR and had only one amplicon in the Y.-Y. Tien and others (Liu et al, 2003); STTR5 (Sty44) (Lindstedt et al, 2003), Sal02 (Sty37), Sal06 (Sty39), Sal20 (Sty40) and Sal10 (Sty43) (Ramisse et al, 2004); and TR4500 (Sty20) and TR4699 (Sty25) (Octavia & Lan, 2009). Two novel VNTRs, Sty2 and Sty3, were discovered in the present study.…”

Core genome conservation of Staphylococcus haemolyticus limits sequence based population structure analysis

“…Reports of low polymorphism in housekeeping genes resulting in limited discriminatory power of MLST have previously been reported for species such as Salmonella enterica, serovar Typhi, Mycoplasma pneumonia and Escherichia coli (Degrange et al, 2009;Dumke et al, 2003;Fakhr et al, 2005;Noller et al, 2003aNoller et al, , 2003b. Molecular typing by MLVF has shown to effectively discriminate homogenous bacterial populations (Noller et al, 2003a(Noller et al, , 2003bOctavia and Lan, 2009). The application of MLVF for epidemiologic studies of S. aureus and S. epidermidis has previously shown a resolution comparable to PFGE and MLST (Francois et al, 2008;Holmes et al, 2010;Pourcel et al, 2009).…”

A novel broadly applicable PCR-RFLP method for rapid identification and subtyping of H58 Salmonella Typhi

“…The fact that S. Typhi is restricted to humans has resulted in a very low genetic variability in this serovar, and the genomes of distinct isolates are extremely conserved ( Achtman, 2008 andHolt et al, 2010). For this reason, the most widely used molecular typing methods, including Multilocus Sequence Typing (MLST) and Pulsed-Field Gel Electrophoresis (PFGE), are not sufficiently discriminative for phylogenetic and epidemiological analysis of this pathogen ( Achtman, 2008, Octavia and Lan, 2009and Thanh et al, 2013. On the contrary, single nucleotide polymorphisms (SNPs) assay has proved to be the most reliable method for S. Typhi genotyping, and over 2000 SNPs were discovered by sequencing different S. Typhi genomes.…”