Salmonella enterica serovar Typhimurium strains were isolated during a monitoring survey of the carpet shell clam (Ruditapes decussatus) collected from 23 stations along the Tunisian coast. The effect of starvation on Salmonella Typhimurium survival was investigated in vitro using microcosms composed of filtered (0.2 µm) marine water under dark and ambient temperature conditions (25C for 30 days). Eventual changes in physiological, biochemical properties, serotyping, biotyping and antimicrobial sensitivity for the environmentally adapted Salmonella Typhimurium were monitored. In response to stress conditions, the Salmonella Typhimurium strains progressively lost culturability in the absence of notable changes in the total cell count (during all the period of 30 days). Strains appeared to enter into a viable but noncultureable state as determined by epifluorescence method. We have also observed that the T90 value (time required for the reduction of 90% of cells) for the five different Salmonella strains ranged between 25 and 30 h; this indicated the survivability of Salmonella under stress conditions. This state was also characterized by biochemical and antimicrobial changes. PRACTICAL APPLICATION This research highlighted the acquisition of the viable but nonculturable state of bacteria. This form of adaptation of certain pathogens does not really reflect their presence or absence in the marine environment and thus in foods of marine origin. Hence the use of conventional methods of bacterial identification must be completed more advanced research tools before declaring a healthy product or environment.
The use of lactic acid bacteria (LAB) in the prevention or reduction of fish diseases is receiving increasing attention. In the present study, 47 LAB strains were isolated from farmed seabass ( Dicentrarchus labrax ) and were phenotypically and phylogenetically analysed by 16S rDNA and randomly amplified polymorphic DNA - polymerase chain reaction (RAPD-PCR). Their antimicrobial effect was tested in vitro against a wide variety of pathogenic and spoilage bacteria. Most of the strains isolated were enterococci belonging to the following species: Enterococcus faecium (59%), Enterococcus faecalis (21%), Enterococcus sanguinicola (4 strains), Enterococcus mundtii (1 strain), Enterococcus pseudoavium (1 strain), and Lactococcus lactis (1 strain). An Aerococcus viridans strain was also isolated. The survey of their antimicrobial susceptibility showed that all isolates were sensitive to vancomycin and exhibited resistance to between 4 and 10 other antibiotics relevant for therapy in human and animal medicine. Different patterns of resistance were noted for skin and intestines isolates. More than 69% (32 strains) of the isolates inhibited the growth of the majority of pathogenic and spoilage bacteria tested, including Listeria monocytogenes, Staphylococcus aureus, Aeromonas hydrophila, Aeromonas salmonicida, Vibrio anguillarum, and Carnobacterium sp. To our knowledge, this is the first report of bioactive enterococcal species isolated from seabass that could potentially inhibit the undesirable bacteria found in food systems.
In order to test the ability to produce antibacterial substances within marine bacteria, prior to select potential probiotics for use in shellfish farming, we targeted a large collection of bacterial isolates (132 strains), brought from the clam Ruditapes decussatus and 37 reference strains. First, we proceeded to their biochemical identification and the screening of antibiotic resistance profiles. Else, we investigated their inhibitory activity in vitro against several fish and shellfish pathogens, using two methods: the double-layer agar and the direct simultaneous antagonism methods. The results showed high frequencies of inhibitory producing bacteria (IPB) within the isolates. These bacteria (25%) were aerobic mesophylic bacteria belonging to various bacterial groups: 33.7% oxidase-positive Gram-negative bacteria, 7.4% Enterobacteriaceae and 28% lactic acid bacteria. Besides this group, nine strains produced strong inhibition effect. These bacteria belonged to : Aeromonas hydrophila, Aeromonas sobria, Pseudomonas cepacia, Vibrio sp, Serratia liquefaciens and Lactobacillus rhamnosus. They were active against pathogenic bacteria belonging to the genera: Aeromonas, Pseudomonas and Vibrio. These potential probiotics were submitted to further investigations prior to their introduction in larval shellfish farming.
This study investigated survival and virulence of Escherichia coli strains exposed to natural conditions in brackish water. Two E. coli strains (O126:B16 and O55:B5) were incubated in water microcosms in the Bizerte lagoon in Northern Tunisia and exposed for 12 days to natural sunlight in June (231 to 386 W/m2, 26 ± 1 °C, 30 g/L) and in April (227 to 330 W/m2, 17 ± 1 °C, 27 g/L) or maintained in darkness for 21 days (17 ± 1 °C, 27 g/L). The results revealed that sunlight was the most significant inactivating factor (decrease of 3 Ulog within 48 hours for the two strains) compared to salinity and temperature (in darkness). Survival time of the strains was prolonged as they were maintained in darkness. Local strain (E. coli O55:B5) showed better survival capacity (T90 = 52 hours) than E. coli O126:B16 (T90 = 11 h). For both, modifications were noted only for some metabolic activities of carbohydrates hydrolysis. Cytotoxicity of the two strains, tested on Vero cell, was maintained during the period of survival.
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