ABSTRACT:The antibacterial activity present in the s k~n mucus of turbot Scophthalmus maximus, seabream Sparus aurdtd and seabass Dicentrarchus labrdx against Pasteul-ella piscicida and Flex~bac-ter maritimus was evaluated. Using assays on agar plates, none of the mucus samples from the above fish showed any antibacterial activity against E maritimus isolates. Turbot mucus inhibited the growth of the P piscicida but mucus from seabream and seabass did not. Assays in liquid systems to determine the survival of the above pathogens In the presence of skin mucus corroborated the results obtained by the agar plate method. The bactericidal properties of the mucus were lost after heat treatment at pH 3.5 and all skin mucus samples displayed act~vity against Staphylococcus aureus ATCC 25923, a straln resistant to lysozyme. These findings indicated that thermolabile substances other than lysozyme were responsible for the antibacterial activity in mucus of marine fish. Enzymatic and heat treatments of the mucus also showed that factors other than complement were involved and that the active component(s) was likely a glycoprotein. Regardless of the source of isolation and degree of virulence, all P piscicida and E maritimus strains adhered strongly to the skin mucus of the 3 fish species tested. Taking all of the foregoing results into consideration, it appears that whereas a possible portal of entry for E maritlmus Into the fish body is the skin, in P piscicida another pathway must be involved.
The pathobiological activities in aiuo and in uifro of live cells and extracellular products (ECP) of eleven Pasteurellu piscicida strains of different origin were examined. Infectivity trials showed that P. piscicida did not possess strict host specificity since the majority of the isolates were virulent for gilthead seabream, rainbow trout and turbot, with LDS0 values ranging between lo3 and lo6 live cells. However, none of the strains tested were pathogenic for mice (LD5,, > lo* cells). In addition, the ECP were strongly toxic for fish (LDS0 ranging from 1-0 to 4.6 pg protein per g fish), which clearly demonstrates their important role in the pathogenesis of pasteurellosis.All the ECP samples were cytotoxic for fish and homoiothermic cell lines, possessed notable phospholipase activity and displayed haemolytic activity for sheep, salmon and turbot erythrocytes (but not for trout erythrocytes).However, the production of proteolytic enzymes differed among the P. piscicidu strains. Although no strain displayed elastase activity, five isolates (the Japanese and Italian strains) hydrolysed casein and gelatin. All these biological activities in uiuo and in uifru were lost after heat treatment (100 "C for 10 min). The general enzymic patterns of both live cells and ECP evaluated by the API-ZYM system also revealed some variation among the P. piscicida isolates. Generally, whole cells showed a wider range of enzymic activities than ECP. The results presented here are important for the selection of strains in the development of effective polyvalent pasteurellosis vaccines containing both whole cells and ECP.
Skretting, Ctra. de la Estació n, Cojobar, Burgos 09620, Spain A novel Gram-stain-negative rod-shaped gliding bacterial strain, designated 35/09 T , was isolated from diseased European sea bass (Dicentrarchus labrax L.) in Spain. Colonies were pale-yellowpigmented with uneven edges and did not adhere to the agar. The DNA G+C content of the isolate was 31.3 mol%. 16S rRNA gene sequence analysis indicated affiliation to the genus Tenacibaculum (family Flavobacteriaceae, phylum 'Bacteroidetes'). Sequence similarities between the isolate and type strains of other members of the genus were 93.1-97.3 %. The major fatty acids (.5 % of the total fatty acids) were iso-C 15 : 0 (24.8 %), iso-C 15 : 0 3-OH (18.0 %), anteiso-C 15 : 0 (8.1 %), C 15 : 1 v6c (6.9 %) and iso-C 15 : 1 (6.2 %). Genotypic and phenotypic data indicate that strain 35/09 T should be classified as a representative of a novel species in the genus Tenacibaculum, for which the name Tenacibaculum dicentrarchi sp. nov. is proposed; the type strain is 35/09 T (5CECT 7612 T 5NCIMB 14598 T ).
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