Evaluation of lipophosphoramidates-based amphiphilic compounds on the formation of biofilms of marine bacteria.

The bacteriostatic and/or bactericidal properties of few phosphoramide-based amphiphilic compounds on human pathogenic bacteria were previously reported. In this study, the potential of two cationic (BSV36 and KLN47) and two zwitterionic (3 and 4) amphiphiles as inhibitors of marine bacterial growth and biofilm formation were investigated. Results showed that the four compounds have little impact on the growth of a panel of 18 selected marine bacteria at a concentration of 200 µM, and up to 700 µM for some bacterial strains. Interestingly, cationic lipid BSV36 and zwitterionic lipids 3 and 4 effectively disrupt biofilm formation of Paracoccus sp. 4M6 and Vibrio sp. D02 at 200 µM and to a lesser extent of seven other bacterial strains tested. Moreover, ecotoxicological assays on four species of microalgae highlighted that compounds 3 and 4 have little impact on microalgae growth with EC50 values of 51 µM for the more sensitive species and up to 200 µM for most of the others. Amphiphilic compounds, especially zwitterionic amphiphiles 3 and 4 seem to be promising candidates against biofilm formation by marine bacteria.

Vibrio aestuarianus subsp. cardii subsp. nov., pathogenic to the edible cockles Cerastoderma edule in France, and establishment of Vibrio aestuarianus subsp. aestuarianus subsp. nov. and Vibrio aestuarianus subsp. francensis subsp. nov.

Cockle mortality events have been reported in northern France since 2012. In the present study, we describe and investigate the implication of a potential bacterial causative agent in cockle mortality. Bacteria isolated from five different cockle mortality events were characterized and studied. Using phenotypic analysis combined with DNA-DNA hybridization (DDH) and whole genome sequencing, the isolates were shown to belong to Vibrio aestuarianus, a species regularly detected in France during oyster mortality events. Comparison of the strains from cockles with strains from French oysters and the type strain showed that the strains from cockles were genetically different to those from oysters and also different to the V. aestuarianus type strain. Moreover, the cockle and oyster strains were classified into two different, but close, groups both separated from the type strain by: (1) analyses of the ldh gene sequences; (2) DDH assays between 12/122 3T3T (LMG 31436T=DSM 109723T), a representative cockle strain, 02/041T (CIP 109791T=LMG 24517T) representative oyster strain and V. aestuarianus type strain LMG 7909T; (3) average nucleotide identity values calculated on the genomes; and (4) phenotypic traits. Finally, results of MALDI-TOF analyses also revealed specific peaks discriminating the three representative strains. The toxicity of representative strains of these cockle isolates was demonstrated by experimental infection of hatchery-produced cockles. The data therefore allow us to propose two novel subspecies of Vibrio aestuarianus: Vibrio aestuarianus subsp. cardii subsp. nov. for the cockle strains and Vibrio aestuarianus subsp. francensis subsp. nov. for the Pacific oyster strains, in addition to an emended description of the species Vibrio aestuarianus.

First evidence for a Vibrio strain pathogenic to Mytilus edulis altering hemocyte immune capacities.

Bacterial isolates were obtained from mortality events affecting Mytilus edulis and reported by professionals in 2010-2013 or from mussel microflora. Experimental infections allowed the selection of two isolates affiliated to Vibrio splendidus/Vibrio hemicentroti type strains: a virulent 10/068 1T1 (76.6% and 90% mortalities in 24 h and 96 h) and an innocuous 12/056 M24T1 (0% and 23.3% in 24 h and 96 h). These two strains were GFP-tagged and validated for their growth characteristics and virulence as genuine models for exposure. Then, host cellular immune responses to the microbial invaders were assessed. In the presence of the virulent strain, hemocyte motility was instantaneously enhanced but markedly slowed down after 2 h exposure. By contrast, hemocyte velocity increased in the presence of the innocuous 12/056 M24T1. At the same time interval, 10/068 1T1 invaded hemocytes and was more rapidly internalized than the innocuous strain. Extracellular products (ECPs) prepared from 10/068 1T1 cultures significantly inhibited phagocytic activity while 12/056 M24T1 ECPs had no effect. Furthermore, the pathogenic strain and its ECPs inhibited oxidative burst unlike 12/056 M24T1 strain/ECPs which enhanced ROS production. Taken together, our results suggest that the mussel pathogen 10/068 1T1 may escape immune response by altering hemocytes functions.