Value of recN sequences for species identification and as a phylogenetic marker within the family “Leuconostocaceae”

The genera Leuconostoc, Oenococcus, and Weissella (family “Leuconostocaceae”) constitute a group of lactic acid bacteria of great interest in food microbiology. From the taxonomic point of view, they are considered phylogenetically coherent according to their 16S rRNA gene sequences and other macromolecules. These three genera were the focus of the present study; specifically, the resolution and discriminatory power of recN (encoding a DNA repair and genetic recombination protein) as a molecular marker at the species level were investigated. For this purpose, partial sequences (about 1200 nt) were obtained from 23 type strains and from several additional strains following direct amplification of recN and subsequent sequencing. Phylogeny was evaluated according to different treeing methods (neighbor joining, maximum likelihood, and maximum parsimony) and the inclusion of variability filters. The results showed that recN, used either alone or in combination with 16S rRNA data, can serve as a phylogenetic marker as well as a tool for species identification.Macian Rovira, M. Carmen, [email protected] ; Garay Auban, Esperanza, [email protected]

Aerobic and facultative anaerobic heterotrophic bacteria associated to Mediterranean oysters and seawater

A comparative study on the composition and seasonal fluctuations of the main heterotrophic bacterial groups and species isolated from Mediterranean oysters and their growing-seawater was carried out. For the study we used 574 strains isolated from Marine Agar (MA) and submitted to numerical analysis of phenotypic traits in previous studies, plus 323 isolates recovered on Thiosulphate Citrate Bile Sucrose (TCBS) agar from the same samples and identified in this study. Oyster samples were dominated by halophilic fermentative bacteria during most of the year with predominance of two Vibrio species, V. splendidus (at temperatures lower than 20°C), and V. harveyi (at higher temperatures). On the contrary, Vibrio spp. was not the predominant microbiota of seawater, where most isolates had remained unidentified but corresponded to α-Proteobacteria, as shown by rDNA hybridization with phylogenetic probes in this study. Among the strict aerobes that could be identified, none of them showed a clear dominance, and many different groups were represented in very low percentages, in contrast with the major species from oyster samples. Shannon-Weaver diversity index revealed significant differences between both types of samples. No apparent seasonality was found in the distribution of seawater species, in sharp contrast with oyster-associated [email protected]; [email protected]; [email protected]

Carriage of potentially fish-pathogenic bacteria in Sparus aurata cultured in Mediterranean fish farms

A bacteriological survey of gilthead sea bream Sparus aurata from different fish farms and culture systems on the Spanish Mediterranean coast was conducted. Three different studies were performed. Study A included hatchery-reared larvae; Study B, periodic examination of randomly sampled growing fish; and Study C, growing fish sampled only during mortality/morbidity events. In Studies B and C, sea cages, earth ponds and indoor tanks were surveyed, and in both cases diseased (showing clinical signs) and non-diseased fish were included. In Study A, a shift from Vibrio spp. (30 d after hatching) to oxidative species (60 d after hatching) was detected, and no mortality events were registered. The percentage of fish yielding bacterial growth were similar in Studies B and C, reaching 57.4 and 61.3%, respectively. A statistically significant relationship between the bacterial carriage and the type of facility was only found in Study B, showing that fish from sea cages had a higher bacterial occurrence than fish from other facilities. A statistically significant relationship between bacterial carriage and signs of disease was found, although the pattern differed in each study. Thus, in Study B only 36.2% of fish yielding abundant bacterial growth were diseased, versus 68.0% in Study C. In total, 25.0% of the fish examined were diseased. Bacterial species composition was similar in asymptomatic and diseased fish, except for a group of V. ichthyoenteri-like isolates that occurred almost exclusively in asymptomatic fish. Dominant bacterial species were V. harveyi and V. splendidus, followed by V. ichthyoenteri-like isolates, Photobacterium damselae ssp. damselae and V. fisheri. Non-fermenters were less frequent but, among them, unidentified halophilic Cytophaga-Flavobacterium isolates and Pseudoalteromonas haloplanktis were the most abundant. An association of individual species with disease was not clear, which suggests the involvement of mixed infections.Pujalte Domarco, M Jesus, [email protected] ; Garay Auban, Esperanza, [email protected]

Identification of Vibrio spp. (other than V. vulnificus) recovered on CPC agar from marine natural samples

Two hundred and eighty four presumptive but not confirmed Vibrio vulnificus isolates grown on cellobiose-polymixin B-colistin agar (CPC) at 40°C, recovered from sea water samples from Valencia, Spain, during a microbiological survey for V. vulnificus, were phenotypically identified. Most of the isolates (91%) corresponded to Vibrio species. V. harveyi (24%) and V. splendidus(19%) were the most abundant species identified, followed by V. navarrensis (13%), V. alginolyticus (8%) and V. parahaemolyticus (5%). The ability to grow on CPC agar and ferment cellobiose of several V. vulnificus strains from different origins and serovars, including reference strains, was tested. Most serovar E isolates and 25% of non-serovar E isolates could not grow on CPC [email protected]; [email protected]; [email protected]; [email protected]; [email protected]