Rapid and sensitive PCR detection of Vibrio penaeicida, the putative etiological agent of syndrome 93 in New Caledonia.
Experimental infections of Penaeus (Litopenaeus) stylirostris were performed with a Vibrio penaeicida strain (AM101) isolated in New Caledonia from Syndrome 93 diseased shrimp. Cumulative mortalities resulting from intramuscular injection or immersion of shrimp in bacterial suspensions demonstrated high virulence for this bacterial strain and suggested that V. penaeicida could be the etiological agent of Syndrome 93. The median lethal dose (LD50) for AM101 was 1.3 x 10(4) CFU (colony forming units) ml-1 by immersion and less than 5 CFU shrimp-1 by intramuscular challenge, with mortality outbreaks at 48 and 22 h after challenge, respectively. A polymerase chain reaction (PCR) detection assay using a primer set designed from the 16S ribosomal RNA gene of V. penaeicida was developed. It gave an expected amplicon of approximately 310 bp in ethidium bromide-stained agarose gels. The specificity of these primers was assessed with different Vibrio species. Furthermore, DNA extracted by the Chelex method could be used to detect fewer than 20 cultured Vibrio cells in sea-water or shrimp hemolymph by this assay. It appears to be a reliable screening method for detecting V. penaeicida in shrimp and from the aquatic environment.
Saulnier
D
D
Laboratoire d'Aquaculture Tropicale, Centre Océanologique du Pacifique, IFREMER, Taravao, Tahiti, French Polynesia. dsaulnie@ifremer.fr
Avarre
J C
JC
Le Moullac
G
G
Ansquer
D
D
Levy
P
P
Vonau
V
V
D000818:Animals; D001483:Base Sequence; D033364:Decapoda / Q000382:microbiology*; D008969:Molecular Sequence Data; D009510:New Caledonia; D016133:Polymerase Chain Reaction / Q000662:veterinary*; D012329:RNA, Bacterial / Q000737:chemistry; D012336:RNA, Ribosomal, 16S / Q000737:chemistry; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D014733:Vibrio / Q000302:isolation & purification*
D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S