title: Multipurpose assessment for the quantification of Vibrio spp. and total bacteria in fish and seawater using multiplex real-time polymerase chain reaction.
abstract: BACKGROUND
  This study describes the first multiplex real-time polymerase chain reaction assay developed, as a multipurpose assessment, for the simultaneous quantification of total bacteria and three Vibrio spp. (V. parahaemolyticus, V. vulnificus and V. anguillarum) in fish and seawater. The consumption of raw finfish as sushi or sashimi has been increasing the chance of Vibrio outbreaks in consumers. Freshness and quality of fishery products also depend on the total bacterial populations present.
  
  
  RESULTS
  The detection sensitivity of the specific targets for the multiplex assay was 1 CFU mL⁻¹ in pure culture and seawater, and 10 CFU g⁻¹ in fish. While total bacterial counts by the multiplex assay were similar to those obtained by cultural methods, the levels of Vibrio detected by the multiplex assay were generally higher than by cultural methods of the same populations. Among the natural samples without Vibrio spp. inoculation, eight out of 10 seawater and three out of 20 fish samples were determined to contain Vibrio spp.
  
  
  CONCLUSIONS
  Our data demonstrate that this multiplex assay could be useful for the rapid detection and quantification of Vibrio spp. and total bacteria as a multipurpose tool for surveillance of fish and water quality as well as diagnostic method.
authors.affiliation: Department of Human Ecology, Food Science, College of Agriculture and Related Sciences, Delaware State University, Dover, DE 19901, USA.
authors.forename: Ji Yeun
authors.initials: JY
authors.lastname: Kim
authors.forename: Jung-Lim
authors.initials: JL
authors.lastname: Lee
mesh_terms: D000818:Animals; D017756:Aquaculture; D001267:Atlantic Ocean; D015169:Colony Count, Microbial; D017124:Conserved Sequence; D004269:DNA, Bacterial / Q000378:metabolism; D004275:DNA, Ribosomal / Q000378:metabolism; D030561:Databases, Nucleic Acid; D003691:Delaware; D005399:Fishes / Q000382:microbiology*; D005513:Food Inspection / Q000379:methods*; D006090:Gram-Negative Bacteria / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D058889:Molecular Typing; D060885:Multiplex Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D017747:Seafood / Q000191:economics / Q000382:microbiology*; D012623:Seawater / Q000382:microbiology*; D014733:Vibrio / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D014736:Vibrio parahaemolyticus / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification / Q000378:metabolism; D041261:Vibrio vulnificus / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S
keywords: Vibrio spp.; fish; multiplex real-time PCR; seafood safety; seawater; total bacteria
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