43 AsPac J. Mol. Biol. Biotechnol., Vol. 11 (1), 2003 AFLP and PFGE of MDR S.typhi Asia Pacific Journal of Molecular Biology and Biotechnology, 2003 Vol. 11 (1) : 43-50 *Author for Correspondence. Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603, Kuala Lumpur, Malaysia. Tel: 603-79674437; Fax: 603-79675908; Email: thongkl@um.edu.my Fluorescent Amplified Fragment Length Polymorphism and Pulsed-Field Gel Electrophoresis Analyses of Multidrug Resistant Salmonella enterica serotype Typhi from different geographical endemic regions in Asia Thong Kwai Lin 1 *, Sleman Elgar 1 , Zulfikar Bhutta 2 , Rohani Yasin 3 and Edgar Schreiber 4 1 Institute of Biological Sciences, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia 2 Aga Khan Uniersity, Medical Centre, Karachi, Pakistan 3 Institute for Medical Research, Malaysia, 4 PE Biosystem, Foster City, USA Received 22 November 2002 / Accepted 2 June 2003 Abstract. Sixty-three isolates of Salmonella enterica serotype Typhi (S.typhi) from sporadic cases of typhoid fever obtained from Malaysia (n=6), Vietnam (n=13), India (n=8) and Pakistan (n=36) were characterized by phage typing, drug-susceptibility test- ing, amplified fragment length polymorphism (AFLP) and pulsed-field gel electrophoresis (PFGE) with restriction endonucle- ase, XbaI. The strains analyzed were mostly resistant to ampicillin, co-trimoxazole, tetracycline and chloramphenicol. These strains were represented by phage types, E1, E9, 46, J1, VNS and UVS. PFGE analysis showed that multidrug resistant (MDR) strains isolated in different countries and at different periods were genetically very homogenous where 90% of the strains analyzed had very closely related XbaI patterns. AFLP was able to subtype 42 clonally related MDR strains (represented by 8 PFGE patterns) into 16 profiles. Cluster analysis based on the AFLP and PFGE data using unweighted pair group mean averages could differentiate MDR strains from different geographical region and that the drug-sensitive strains were in a distinct cluster. The clustering of MDR strains from each country and the presence of a dominant XbaI–PFGE pattern indicated that the MDR S.typhi had probably been spread clonally in these countries. AFLP is clearly more discriminative than PFGE in differentiating the MDR S.typhi, hence providing an alternative, sensitive method for detailed analysis of the multidrug resistant strains. Keywords. AFLP, PFGE, S. typhi INTRODUCTION With regard to emerging infectious diseases, typhoid fever caused by multidrug resistant (MDR) strains of Salmonella enterica serotype Typhi (herein named as S. typhi) present a serious health problem, especially in the Asian and Southeast Asian countries. Multidrug resistant (MDR) S. typhi strains were defined as those resistant to ampicillin, chloramphenicol and trimethoprim-sulphamethoxazole with additional resistance to streptomycin, sulfonamides and tetracycline. Drug-resistant S.typhi began to appear in the late 1980’s in the South Asian region and have now spread widely to the Middle East, Africa and Asia (Rowe et. al ., 1997). The problem is particularly acute on the Indian subcontinent where approximately 60-65 % of strains isolated are MDR strains. In addition, the disease associated with MDR tends to be more severe with patients registering a higher morbidity score and a higher mortality rate, often with unusual complications (Bhuttta, 1996). The impact of this MDR further complicates the antibiotic treatment by limiting treatment options, prolonged hospitalization and the increasing costs of therapy. In many tropical countries, typhoid fever is endemic and has the potential for epidemic spread as drug resistance is often encoded by plasmids and are highly transmissible (Connerton et al., 2000). Studies have shown these plasmids encoding resistance to chloramphenicol, ampicillin, trimethoprim, sulfonamides, and tetracycline belonged to the IncHI incompatibililty group and are conjugative (Hermans et al., 1996, Hampton et al., 1998). The ability to characterize and subtype S. typhi isolates is important from an epidemiological perspective in order to trace and control the dissemination of this infectious agent. Recently, many DNA-based approaches have been utilized in the molecular subtyping of S. typhi, including ribotyping (Ling et al., 2000), pulsed-field gel electrophoresis (PFGE)