MOLECULAR EPIDEMIOLOGY AND EVOLUTIONARY GENETIC RELATIONSHIPS OF SALMONELLA TYPHI

Abstract

Characterization of strains in outbreak investigations depends largely on the utility of highly precise typing tools. The genomic DNAs of 39 Salmonella typhi strains isolated from local residents, patients who had visited neighbouring countries and tourists from the Asian region were analysed for restriction fragment length polymorphisms (RFLP). Analysis was carried out by pulsed-field gel electrophoresis (PFGE) of Xbal and Spel restriction fragments. IS 200 internal fragment and ribosomal genes (rRNA) were also used as probes to detect chromosomal restriction fragment length polymorphisms. Pulsed-field gel electrophoresis analysis established 22 PFGE types whereas phage typing differentiated the 39 isolates into 9 distinct phage types. Ribotyping analysis of genomic DNA digested with Clal and Pst1 produced 15 combined ribotypes, while IS 200 analysis with Hincll and Pst1 generated four combined IS 200 types. All the three molecular typing methods had 100% typeability and reproducibility, but PFGE with either Spel and Xbal was established to be the most discriminatory method (Simpson's Index for diversity [01]=0.95). Ribotyping with two enzymes was more discriminatory (01=0.89) than IS 200 profiling with Hincll and Pst1 (DI=O.71 ). The discriminatory power of ribotyping (D1=0.89) could only approach that of phage typing (D1=0.90) whilst IS 200 profiling was inferior. The three molecular methods demonstrated the multiclonal nature of S.typhi isolates recovered from Asia. RAPD-PCR analysis which had a discriminatory index value of 0.76 was able to delineate the 39 S.typhi isolates into 3 profiles, but it's discriminatory power was inferior to that of PFGE, phage typing and ribotyping. Three other PeR-methodologies comprising of PCR-ribotyping, PCR-RFLP and rep-PCR were found to be of limited value in the epidemiological studies of S.typhi because these methods were unable to differentiate the 39 S.typhi isolates. Genetic relatedness among the isolates was determined based on PFGE studies. Seven major clusters were identified at SABs of >0.80 and the remaining 13 isolates were distributed into minor clusters which were related at SABs of less than 0.80. PFGE analysis in conjunction with distance matrix analysis served as a useful tool for delineating common S.typhi phage types of diverse origins from different geographical locales and separated in time, and thus making PFGE analysis a powerful genotyping method for establishing clonal and epidemiological relationships among S.typhi isolates.