Vibrio tapetis-like strain isolated from introduced Manila clams Ruditapes philippinarum showing symptoms of brown ring disease in Norway

Abstract

The Manila clam Ruditapes philippinarum was introduced to Norway in 1987 and was produced in 2 hatcheries until 1991. Clam seed was planted at 6 sites. Two sites were on the Island of Tysnes, south of Bergen. Surviving adult Manila clams were recovered in 1995 and 1996. In the present study, Manila clams from the original seeding that displayed morphological signs of brown ring disease (BRD) were recovered in June 2003 (n = 7) and in June 2004 (n = 17). Samples from extrapallial fluid, tissues and haemolymph were inoculated on marine agar. Replicate subcultures on selective media were used to select potential Vibrio tapetis strains, and in total, 190 bacterial strains were isolated. One of these strains clustered within the V. tapetis clade and was named NRP 45. DNA:DNA hybridisation with the type strain CECT4600 showed 52.7 and 57.3% DNA:DNA similarity. Hybridisation of NRP 45 and the V. tapetis LP2 strain, isolated from corkwing wrasse Symphodus melops, produced 46.6 and 44.4% re-association. Partial gene segments encoding 16S rRNA, gyrase B protein (GyrB) and chaperonin 60 protein (Cpn60) were characterised and compared to CECT 4600. NRP 45 showed 5 differences in the 1416 nucleotides (nt) of the 16S rRNA encoding gene (99.6% similarity), while the GyrB encoding gene had 62 substitutions of 1181 nt compared (94.8% similarity) and the Cpn60 encoding gene had 22 substitutions out of 548 nt compared (96% similarity). This is the first finding of BRD and the first isolation of a V. tapetis-like bacterial strain from a bivalve in Norway. KEY WORDS: Brown ring disease · Vibrio tapetis · Bivalve · Manila clams · Bacteria · NorwayResale or republication not permitted without written consent of the publisher Dis Aquat Org 81: [153][154][155][156][157][158][159][160][161] 2008 disease has subsequently spread along the East Atlantic coast from England to North Africa (Paillard 2004). BRD was first observed in Spain in 1994, in England in 1997 and in Ireland in 1998 (Paillard & Maes 1994, Castro et al. 1996, Novoa et al. 1998. The highest prevalence is encountered in the northernmost countries, and BRD is generally classified as a coldwater disease (Paillard et al. 1994). In Norway, the Manila clam was introduced in 1987 (Mortensen & Strand 2000). Approximately 500 broodstock specimens were imported from Scottish Seafarms Ltd to 2 Norwegian hatcheries, located in Espevik on the Island of Tysnes, south of Bergen, and in Vallersund in South Trøndelag County (Fig. 1A). From 1987 to 1991, more than 200 million seed were produced, with ongrowth done in nurseries at Espevik, Rong, Vallersund and Vågstanda. Most of the Manila clam seed were exported to Spain and Ireland. In Espevik, the native clam R. decussatus was also produced by the same methods and during the same period as the Manila clams. Clam production was terminated in 1991. Between 1988 and 1991, Manila clam seed were released on beaches at 6 sites. At Seløy (Fig.1B), approximately 2000 Manila clam spat were released in 1988, and a total of 28 speci...

Mitochondrial thioredoxin-2 from Manila clam (Ruditapes philippinarum) is a potent antioxidant enzyme involved in antibacterial response

“…We investigated the transcriptional response of RpTRx-2 in gill and hemocytes to immune challenges, by using LPS endotoxin and a potent Manila clam pathogen V. tapetis [36]. RpTRx-2 was significantly up-regulated in gills of immune-challenged animals (P < 0.05), with the highest induced expression (2-fold increases) occurring at 6 h post-LPS challenge and at 24 and 96 h postbacterial challenge (Fig.…”

A survey of the health status of the Manila clam Ruditapes philippinarum in Ireland with specific reference to brown ring disease

“…In Europe, R. philippinarum was first introduced to France in 1972, England in 1980, Ireland in 1982, and later in Spain and Italy in 1983and Norway in 1987(Breber 1985Flassch and Leborgne 1992;Clarke 1999;Paillard et al 2008). Due to its fast growth and high productivity, it has proved to be suitable for cultivation and is now grown commercially in Italy, France, Spain, the UK and Ireland (FAO 2002).…”

Evaluation of different culture media for the isolation and growth of the fastidious Vibrio tapetis, the causative agent of brown ring disease

“…In addition, Nutrient Blood Agar (NBA), Brain Heart Broth Agar (BHBA) and Trypticase Soy Agar (TSA) supplemented 1.5% (w/v) with sea salts and citrated horse blood (TSSBA) were employed when the isolation was made from fish (Bergh and Samuelsen, 2007;Jensen et al, 2003;Reid et al, 2003). Incubation conditions in the different studies ranged from 18°C to 22°C and from 24 h to 7 days (Bergh and Samuelsen, 2007;Castro et al, 1997;Jensen et al, 2003;Maes and Paillard, 1992;Novoa et al, 1998;Paillard et al, 2008;Reid et al, 2003). It has been often described that classical methods may fail in the detection of typical V. tapetis or that, although being detected by indirect procedures (i.e.…”

“…For the isolation of this pathogen from shellfish samples several media were used (Castro et al, 1997;Maes and Paillard, 1992;Novoa et al, 1998;Paillard et al, 2008), including Marine Agar (MA), Thiosulfate-Citrate-Bile salt-Sucrose agar (TCBS), and Mannitol Marine Agar (MMA). In addition, Nutrient Blood Agar (NBA), Brain Heart Broth Agar (BHBA) and Trypticase Soy Agar (TSA) supplemented 1.5% (w/v) with sea salts and citrated horse blood (TSSBA) were employed when the isolation was made from fish (Bergh and Samuelsen, 2007;Jensen et al, 2003;Reid et al, 2003).…”