title: Molecular characterization and PCR-based replicon typing of multidrug resistant Shigella sonnei isolates from an outbreak in Thimphu, Bhutan.
abstract: BACKGROUND
  Shigella species are an important cause of diarrhea in developing countries. These bacteria normally acquire their antibiotic resistance via several different mobile genetic elements including plasmids, transposons, and integrons involving gene cassettes. During a diarrhea surveillance study in Thimphu, Bhutan in June and July, 2011, Shigella sonnei were isolated more frequently than expected. This study describes the antibiotic resistance of these S. sonnei isolates.
  
  
  METHODS
  A total of 29 S. sonnei isolates from Thimphu, Bhutan was characterized for antimicrobial susceptibility by disc diffusion assay and minimum inhibitory concentration (MIC) assay. All isolates were tested by pulsed-field gel electrophoresis (PFGE) with restriction enzyme XbaI and were tested for plasmid. The plasmid patterns and the PFGE patterns were analyzed by Bionumerics software. DNA sequencing was performed on amplified products for gyraseA gene and class 1 and class 2 integrons. S. sonnei isolates were classified for incompatibility of plasmids by PCR-based replicon typing (PBRT).
  
  
  RESULTS
  These S. sonnei were resistant to multiple drugs like ciprofloxacin, nalidixic acid, trimethoprim-sulfamethoxazole, streptomycin, and tetracycline but susceptible to azithromycin. All isolates had class 2 integrons dfrA1, sat1 and aadA1 genes. Two point mutations in Gyrase A subunit at position Ser83Leu and Asp87Gly were detected in these quinolone resistant isolates. The plasmid and PFGE patterns of S. sonnei isolates suggested a clonal relationship of the isolates. All isolates carried common ColE plasmid. ColE plasmid co-resided with B/O plasmid (nine isolates) or I1 plasmid (one isolate).
  
  
  CONCLUSIONS
  The characteristics of 29 S. sonnei isolates from Thimphu, Bhutan in June and July, 2011 are identical in PFGE, plasmid and resistance pattern. This study suggests that these recent S. sonnei isolates are clonally related and multidrug-resistant.
authors.affiliation: Department of Enteric Diseases, Armed Forces Research Institute of Medical Sciences (AFRIMS), Bangkok, Thailand. sirigader@afrims.org.
authors.forename: Sirigade
authors.initials: S
authors.lastname: Ruekit
authors.forename: Sonam
authors.initials: S
authors.lastname: Wangchuk
authors.forename: Tshering
authors.initials: T
authors.lastname: Dorji
authors.forename: Kinzang Pem
authors.initials: KP
authors.lastname: Tshering
authors.forename: Piyarat
authors.initials: P
authors.lastname: Pootong
authors.forename: Panida
authors.initials: P
authors.lastname: Nobthai
authors.forename: Oralak
authors.initials: O
authors.lastname: Serichantalergs
authors.forename: Kamonporn
authors.initials: K
authors.lastname: Poramathikul
authors.forename: Ladaporn
authors.initials: L
authors.lastname: Bodhidatta
authors.forename: Carl Jeffries
authors.initials: CJ
authors.lastname: Mason
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001631:Bhutan / Q000453:epidemiology; D002675:Child, Preschool; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004196:Disease Outbreaks / Q000517:prevention & control; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004405:Dysentery, Bacillary / Q000453:epidemiology / Q000382:microbiology; D016521:Electrophoresis, Gel, Pulsed-Field; D054884:Host-Pathogen Interactions; D006801:Humans; D007223:Infant; D032023:Integrons / Q000235:genetics; D008826:Microbial Sensitivity Tests; D058889:Molecular Typing / Q000379:methods*; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D012093:Replicon / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012764:Shigella sonnei / Q000145:classification / Q000235:genetics* / Q000502:physiology; D013045:Species Specificity
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; C025580:endodeoxyribonuclease XBAI; D015252:Deoxyribonucleases, Type II Site-Specific