RESUMEN
Two Vibrio strains (VPAP36 and VPAP40) were isolated from moribund-settled larvae of the Chilean scallop Argopecten purpuratus during vibriosis outbreaks that occurred in two commercial scallop larvae hatcheries located in the Inglesa and Tongoy bays in Northern Chile. The strains were identified as Vibrio chagasii using phenotypic characterization and whole genome sequence analysis. Both strains exhibited the phenotypic properties associated with virulence, gelatin hydrolysis and ß-hemolysis, whereas only VPAP36 produced phospholipase and only VPAP40 produced caseinase. The whole genome analysis showed that the strains harbored genes encoding for the virulence factors, the EPS type II secretion system, and Quorum Sensing (auto-inductor 1 and auto-inductor 2), whereas genes encoding a metalloproteinase and a capsular polysaccharide were detected only in the VPAP40 genome. When challenge bioassays using healthy 11-day-old scallop larvae were performed, the V. chagasii VPAP36 and VPAP40 strains exhibited significant (p < 0.05) differences in their larval lethal activity, producing, after 48 h, larval mortalities of 65.51 ± 4.40% and 28.56 ± 5.35%, respectively. Otherwise, the cell-free extracellular products of the VPAP36 and VPAP40 strains produced larval mortalities of 20.86 ± 2.40% and 18.37 ± 2.40%, respectively, after 48 h of exposure. This study reports for the first time the isolation of V. chagasii from the massive larval mortalities of the farmed scallop (Argopecten purpuratus) in Chile, and demonstrates the pathogenic activity of V. chagasii towards the Chilean scallop, the second most important species for Chilean mariculture.
RESUMEN
Vibrio europaeus is an emergent pathogen affecting the most important bivalve species reared in Spanish and French hatcheries. Using a genomic approach, we identified V. europaeus outside Europe for the first time from massive larval mortalities of scallop (Argopecten purpuratus) in Chile and from seawater near a shellfish hatchery in the US West Coast. Results show the worldwide spreading and potential impact of V. europaeus for aquaculture; these four countries are among the 10 major producers of mollusks. Pathogenicity of V. europaeus was demonstrated for the first time towards scallop, the second most important species for Chilean mariculture.
Asunto(s)
Pectinidae/microbiología , Vibrio/aislamiento & purificación , Animales , Acuicultura , Chile , Filogenia , Estados Unidos , Vibrio/clasificaciónRESUMEN
The VPAP30 strain was isolated as the highly predominant bacteria from an episode of massive larval mortality occurring in a commercial culture of the Chilean scallop Argopecten purpuratus. The main aims of this study were, to characterize and identify the pathogenic strain using biochemical and molecular methods, to demonstrate its pathogenic activity on scallop larvae, to characterize its pathogenic properties and to describe the chronology of the pathology. The pathogenic strain was identified as Vibrio bivalvicida based on its phenotypic properties, the multilocus sequence analysis (MLSA) of eight housekeeping genes (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA, and topA) and different in silico genome-to-genome comparisons. When triplicate cultures of healthy 10 days old scallop larvae were challenged with 1 × 105 colony forming units (CFU) mL-1 of the VPAP30 strain, percentages of larval survival of 78.9 ± 3.3%, 34.3 ± 4.9%, and 0% were observed at 12, 2,4 and 36 h, respectively, whereas uninfected larval cultures showed survival rates of 97.4 ± 1.2% after of 48 h. Clinical symptoms exhibited by the scallop larvae infected with the VPAP30 strain include the accumulation of bacteria around the scallop larvae, velum disruption and necrosis of digestive gland. The 50% lethal dose (LD50) of VPAP30 strain at 24 and 48 h was 1.3 × 104 and 1.2 × 103 CFU mL-1, respectively. The invasive pathogenic activity of the VPAP30 strain was investigated with staining of the bacterial pathogen with 5-DTAF and analyzing bacterial invasion using epifluorescence, and a complete bacterial dissemination inside the larvae at 24 h post-infection was observed. When scallop larvae were inoculated with cell-free extracellular products (ECPs) of VPAP30, the larval survival rate was 59.5 ± 1.7%, significantly (P < 0.001) lower than the control group (97.4 ± 1.2%) whereas larvae treated with heat-treated ECPs exhibited a survival rate of 61.6 ± 1.8% after 48 h of exposure. V. bivalvicida VPAP30 exhibits high pathogenic activity on scallop larvae, mediated both by bacterial invasion and the production of toxigenic heat-stable compounds. This report constitutes the first isolation of V. bivalvicida out of Europe and extends the host range of this species, having demonstrated its pathogenic activity on the Chilean scallop larvae (A. purpuratus). These results supporting the pathogenic potential of V. bivalvicida to kill the larvae of a broad range of bivalve species reared in hatcheries located in the Atlantic and the Pacific coasts.
RESUMEN
Strain CAIM 1076T was isolated from a cultured oyster Crassostrea gigas in Puerto Peñasco, Sonora state, México. The strain was taxonomically characterised by means of a genomic approach, comprising 16S rRNA gene sequence analysis, multilocus sequence analysis (MLSA), the DNA G+C content and whole genome analyses (ANI and GGDC), and by phenotypic characterisation. Strain CAIM 1076T was found to be catalase and oxidase positive, and cells were observed to be motile and facultative anaerobic. Analysis of the almost-complete 16S rRNA gene sequence placed this strain within the genus Vibrio; closely related species were Vibrio maritimus, Vibrio variabilis, Vibrio proteolyticus, and Vibrio nigripulchritudo with similarity values of 98.9, 98.5, 98.1, and 98.0 %, respectively. MLSA of six housekeeping genes (ftsZ, gapA, gyrB, recA, rpoA and topA) was performed with the closely related species. A draft genome sequence of strain CAIM 1076T was obtained. The DNA G+C content of this strain was determined to be 44.5 mol %. The genomic similarity values with V. maritimus were 71.6 % (ANIb), 85.1 % (ANIm) and a GGDC value of 20.3 ± 2.3 %; with V. variabilis the genomic similarities were 71.8 % (ANIb), 85.4 % (ANIm) and 20.0 ± 2.3 % (GGDC); with V. proteolyticus, 71.6 % (ANIb), 84.1 % (ANIm) and 18.8 ± 2.2 % (GGDC); and with V. nigripulchritudo, 70.8 % (ANIb), 84.9 % (ANIm) and 20.5 ± 2.3 % (GGDC). These ANI and GGDC values are below the thresholds for the delimitation of prokaryotic species, i.e., 95-96 and 70 %, respectively. Phenotypic characters also showed differences with the closely related species analysed. The results presented here support the description of a novel species, for which the name Vibrio sonorensis sp. nov. is proposed, with strain CAIM 1076T (=CECT 9100T, =DSM 102190T) as the type strain.