Vibrio vulnificus is a marine bacterium that causes septicemia with high mortality via wound infection or seafood ingestion. Subspecies typing of V. vulnificus can be performed by pulsed-field gel electrophoresis (PFGE). However, this method is hampered by the degradation of chromosomal DNA in about 12% of strains. This study presents a modified PFGE procedure. The outer membrane of V. vulnificus cells was lysed utilizing a sucrose-EDTA method and components in the periplasmic space were removed prior to making plugs. Cytoplasmic digestive enzymes were inactivated by an extended proteinase K reaction. Experimental results indicated that five out of eight strains exhibiting smeared DNA in a previous study (Appl. Environ. Microbiol. 70: 5153-5158. 2004) achieved clear banding patterns. This modified PFGE procedure can be applied to improve PFGE typing of untypeable strains of V. vulnificus.
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