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Background: Vibriosis is one of the most ubiquitous fish diseases caused by bacteria belonging to the genus Vibrio such as Vibrio (Listonella) anguillarum. Despite a lot of research efforts, the virulence factors and mechanism of V. anguillarum are still insufficiently known, in part because of the lack of standardized virulence assays. Methodology/Principal Findings: We investigated and compared the virulence of 15 V. anguillarum strains obtained from different hosts or non-host niches using a standardized gnotobiotic bioassay with European sea bass (Dicentrarchus labrax L.) larvae as model hosts. In addition, to assess potential relationships between virulence and genotypic and phenotypic characteristics, the strains were characterized by random amplified polymorphic DNA (RAPD) and repetitive extragenic palindromic PCR (rep-PCR) analyses, as well as by phenotypic analyses using Biolog’s Phenotype MicroArray™ technology and some virulence factor assays. Conclusions/Significance: Virulence testing revealed ten virulent and five avirulent strains. While some relation could be established between serotype, genotype and phenotype, no relation was found between virulence and genotypic or phenotypic characteristics, illustrating the complexity of V. anguillarum virulence. Moreover, the standardized gnotobiotic system used in this study has proven its strength as a model to assess and compare the virulence of different V. anguillarum strains in vivo. In this way, the bioassay contributes to the study of mechanisms underlying virulence in V. anguillarum. [ABSTRACT FROM AUTHOR]

Hatching of resting eggs of Streptocephalus torvicornis (Waga, 1942) and Branchipus schaefferi Fischer, 1834, two sympatric fairy shrimps, was tested using medium in which these two species and Chirocephalus diaphanus (Prévost, 1803) had been raised. The medium of adults inhibited the hatching of conspecific nauplii, even when diluted by 50% using distilled water. Streptocephalus torvicornis resting eggs hatch in the medium of C. diaphanus but not in the medium of B. schaefferi. Resting eggs of B. schaefferi hatched in low numbers in S. torvicornis medium, and in high numbers in that of C. diaphanus. There appears to be one or more substances associated with the presence of adults of conspecifics and sympatrics which exceed the influences of abiotic factors, such as light, pH, conductivity and osmotic pressure in fresh media. We propose that this substance(s) functions as a pheromone to inhibit further hatching once a population of adults is present. [ABSTRACT FROM AUTHOR]

Abstract: Axenic rotifers (Brachionus plicatilis sensu strictu, clone 10) were obtained by treating amictic eggs with glutaraldehyde. Depending of the batch of rotifers, total disinfection could be obtained by exposure to 50–100 ppm from 1 to 2 h at 28 °C. The hatched axenic neonates were used to test the effect of microbial communities (MCs) which were isolated from either normal-performing or crashed rotifer cultures. These MCs were either used directly or were first regrown on Marine Agar. MCs were introduced to gnotobiotic Brachionus cultures in combination with three different food types, i.e. Chlorella, wild-type yeast and the mnn9 yeast mutant, which is deficient in cell wall-bound mannoprotein. In the absence of MCs or when heat-killed MCs were added, Chlorella was always the best food, while lower growth rates were observed with wild-type yeast and the mnn9 mutant as food. In the presence of live MCs and when rotifers were fed with Chlorella, the added MCs had no effect on rotifer performance. When yeasts were used as major food, all the tested MCs were able to increase the rotifer growth rate. The experiments with heat-killed MCs yielded no increase in rotifer growth rate, suggesting that the observed enhancement in rotifer growth rate was truly a probiotic effect rather than a nutritional effect. The results of this study demonstrate that gnotobiotic rotifer cultures obtained from axenic amictic eggs can be used as a test system for studying microbial-attributed as well as nutritional functions in the aquatic food chain. In addition, since the MCs originating from the crashed rotifer cultures did not decrease the growth rate in the tested rotifer cultures, it is likely that the observed crashes were not due to the presence of a standing deleterious MC. [Copyright &y& Elsevier]

Gastrointestinal microbiota have an important impact on fish health and disease, stimulating interest in a better understanding of how these gastrointestinal microbial communities are composed and consequently affect host fitness. In this respect, probiotic microorganisms have been extensively used in recent aquaculture production. To study the use of probiotics in the treatment of infectious diseases, the establishment of a method of experimental infection to obtain consistent results for mortality and infection in challenge tests is important. In pathogen-screening tests, 4 candidate pathogenic bacteria strains (Edwardsiella ictaluri gly09, E. ictaluri gly10, E. tarda LMG2793 and Streptococcus agalactiae LMG15977) were individually tested on xenic Nile tilapia larvae. Only Edwardsiella strains delivered via Artemia nauplii, with or without additional pathogen delivery via the culture water, led to increased mortality in fish larvae. A gnotobiotic Nile tilapia larvae model system was developed to provide a research tool to investigate the effects and modes-of-action of probiotics under controlled conditions. A double disinfection procedure using hydrogen peroxide and sodium hypochlorite solution was applied to the fish eggs, which were subsequently incubated in a cocktail of antibiotic and antifungal agents. In the gnotobiotic challenge test, E. ictaluri gly09R was added to the model system via Artemia nauplii and culture water, resulting in a significant mortality of the gnotobiotic fish larvae. The developed gnotobiotic Nile tilapia model can be used as a tool to extend understanding of the mechanisms involved in host-microbe interactions and to evaluate new methods of disease control.

Vibrio anguillarum is the major cause of haemorrhagic septicaemia, vibriosis, which is a severe disease affecting marine fish. In this work, it was found that the mortality of gnotobiotic sea bass larvae challenged with V. anguillarum was dependent on the number of dead fish in the vials at the moment of challenge. Based on this finding, the effect of dead hosts (homogenised sea bass larvae or brine shrimp) on the virulence of V. anguillarum towards sea bass larvae was further investigated. Addition of homogenised hosts led to significantly increased larval mortality of challenged larvae, and this was observed for 3 different V. anguillarum strains, i.e. 43, NB 10 and HI 610. In contrast, the addition of similar levels of tryptone had no effect on mortality. In line with this, the motility of all 3 V. anguillarum strains was significantly increased by the addition of homogenised hosts but not by tryptone. These results suggest that dead hosts increase infectivity of V. anguillarum, not merely by offering nutrients to the bacteria, but also by increasing virulence-associated activities such as motility.