Characterization of Salmonella enterica serovar Typhimurium and monophasic Salmonella serovar 1,4,[5],12:i:- isolates in Thailand

Abstract

Duplex PCR was developed to screen Salmonella enterica serovar Typhimurium phage type DT104 and related strains in Thailand because a phage typing laboratory of serovar Typhimurium is not available. Of 46 isolates of serovar Typhimurium and 32 isolates of S. enterica serovar 1,4,[5],12:i:-, 15 (33%) and 30 (94%) were duplex PCR positive, respectively. All isolates were submitted for phage typing to analyze the specificity of the PCR assay. Among serovar Typhimurium isolates that yielded positive duplex PCRs, only seven isolates were phage types DT104 or U302, and eight isolates were undefined types, whereas the negative PCR isolates were either other phage types, including DT7, DT12, DT66, DT79, DT166, DT170, DT193, and DT208 or an undefined type. The serovar Typhimurium and serovar 1,4,[5],12:i:- isolates that were duplex PCR positive were further subtyped by using XbaI PFGE to reveal their genetic relatedness. All serovar Typhimurium phage type DT104 strains had indistinguishable chromosomal patterns. The isolates of phage type U302 and most of the serovar 1,4,[5],12:i:- isolates that were duplex PCR positive yielded similar pulsed-field gel electrophoresis patterns. The patterns of PCR-negative isolates distinctly differed from the patterns of PCR-positive isolates. A total of 26% of all isolates had a dominant R-type ACSSuTG that was not found in the isolates of phage type DT104.

FIG. 1.

Duplex PCR of Salmonella isolates. The 162-bp fragment is specific to phage type DT104. The 261-bp fragment is specific to serovar Typhimurium. Lane M, molecular marker pUC18 digested with HaeIII; lane 1, serovar Typhimurium phage type DT104; lane 2, serovar Typhimurium phage type DT12; lanes 3, 6, and 8, serovar 1,4,[5],12:i:-; lane 4, serovar Typhimurium RDNC; lane 5, Salmonella serovar Mbandaka.

FIG. 2.

PFGE XbaI digestion patterns of Salmonella Typhimurium phage types DT104 and U302 and Salmonella serovar 1,4,[5],12:i:-. Lane M, lambda ladder molecular marker; lane λ, 23 kb of λHindIII; lane 2, pattern U; lanes 1, 3, 6, and 8, patterns U6, U7, U5, and U10, respectively; lanes 4 and 5, patterns of serovar Typhimurium that yielded only one band of duplex PCR; lane 7, PFGE pattern of Salmonella serovar Mgulani.