VIRULENCE GENE AND CRISPR MULTILOCUS SEQUENCE TYPING SCHEME FOR SUBTYPING THE MAJOR SEROVARS OF SALMONELLA ENTERICA SUBSPECIES ENTERIC

Open Access
Author:
Liu, Fenyun
Graduate Program:
Food Science
Degree:
Master of Science
Document Type:
Master Thesis
Date of Defense:
October 04, 2010
Committee Members:
  • Dr Steve Knabel And Dr Edward Dudley, Thesis Advisor
  • Stephen John Knabel, Thesis Advisor
  • Edward G Dudley, Thesis Advisor
Keywords:
  • subtyping
  • MLST
  • Salmonella
Abstract:
Salmonella enterica subsp. enterica is the leading cause of bacterial foodborne disease in the United States. Molecular subtyping methods are powerful tools for tracking the farm-to-fork spread of foodborne pathogens during outbreaks. In order to develop a novel multilocus sequence typing (MLST) scheme for subtyping the most prevalent serovars of Salmonella, the virulence genes fimH and sseL and Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) regions were sequenced from 171 clinical isolates from serovars Typhimurium, Enteritidis, Newport, Heidelberg, Javiana, I 4, [5], 12; i: -, Montevideo, Muenchen and Saintpaul. Another 63 environmental isolates and 70 poultry isolates of S. Enteritidis from poultry industries in PA were also analyzed. The MLST scheme using only virulence genes was insufficient to separate all unrelated outbreak clones. However, the addition of CRISPRs sequences dramatically improved discriminatory power of this MLST method. Moreover, the present MLST scheme provided better discrimination of S. Enteritidis strains than PFGE. Cluster analyses revealed the current MLST scheme is highly congruent with serotyping and epidemiological data. For the analyses with S. Enteritidis isolates, the current MLST scheme identified three persistent and predominate sequence types circulating among humans in the U.S. and poultry and hen house environments in PA. It also identified an environment-specific sequence type. In conclusion, the novel MLST scheme described in the present study accurately differentiated outbreak clones of the most common serovars of Salmonella, and therefore maybe an excellent tool for subtyping this important foodborne pathogen during outbreak investigations. Furthermore, the MLST scheme may provide information about the ecological origin of S. Enteriditis isolates, potentially identifying strains that differ in virulence capacity.