Discrimination of d -Tartrate-Fermenting and -Nonfermenting Salmonella enterica subsp. enterica Isolates by Genotypic and Phenotypic Methods

Abstract

ABSTRACT A multiplex PCR and an improved lead acetate test were developed to discriminate d -tartrate-fermenting and -nonfermenting Salmonella enterica subsp. enterica strains. Both methods showed an accuracy of 100% when 125 Salmonella strains belonging to 15 serovars were tested. Special emphasis was given to S. enterica subsp. enterica serovar Paratyphi B isolates because of the clinical importance of its d -tartrate-nonfermenting variant and the recently increasing numbers of cases of human outbreaks caused by its fermenting variant (formerly Salmonella serovar Java). The lead acetate test described previously (G. A. Alfredsson, R. M. Barker, D. C. Old, and J. P. Duguid, J. Hyg. 70:651-666, 1972) was modified in the inoculation and incubation procedure. The PCR assay was based on the genotypic difference of the presence ( d -tartrate-fermenting strains) or absence ( d -tartrate-nonfermenting strains) of the ATG start codon for the gene STM 3356, which encodes a putative cation transporter. Sequence data revealed a nucleotide exchange from G to A within the ATG start codon of gene STM 3356 in the d -tartrate-nonfermenting strains. In order to increase the reliability of the PCR assay, a positive control based on a Salmonella genus-specific primer set for the detection of Salmonella DNA was included. The PCR-based discrimination needs only several hours compared to 6 days needed by the improved lead acetate test to obtain reliable results. Consequently, the PCR d -tartrate assay should be the method of choice for the discrimination of d -tartrate-fermenting and -nonfermenting Salmonella strains in the future.