Piscirickettsiaceae representative Piscirickettsia salmonis Isolation of the

Piscirickettsiaceae representative piscirickettsia

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Piscirickettsiaceae representative Piscirickettsia salmonis Isolation of the pathogen from the kidney of infected fish was possible in the cytoplasm of salmon cell lines (including CHSE-214) with incubation at 12-2rC (optimally at 15-18°C), whereupon a cytopathic effect was demonstrated in 5-6 days (Fryer et al., 1990). The cell sheet was completely lysed in 14 days. Growth did not occur on bacteriological media, including BHIA, blood agar, mycoplasma medium, charcoal yeast extract agar, Loeffler medium (Appendix 5.1) or Mueller-Hinton agar. Pseudomonadaceae representatives Pseudomonas anguilliseptica Isolation of Ps. anguilliseptica is readily achieved from blood, kidney, liver and spleen samples by use of nutrient agar supplemented with 10% (v/v) horse blood or nutrient agar containing 0.5% (w/v) sodium chloride, and adjusted to a pH of 7.4. Incubation
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Isolation/Detection 171 should be at 20-25°C for at least 7 days, when small (<lmm in diameter), round, raised, entire, shiny, pale-grey colonies develop (Wakabayashi and Egusa, 1972). Pseudomonas chlororaphis This may be accomplished by inoculating homogenates, prepared from the entire fish, onto the surface of nutrient agar plates, with incubation at 25°C for 5 days. It is surprising that this simple method enabled the recovery of pure cultures, because contaminants may be expected from the intestine and body surface (Hatai et al, 1975). Pseudomonas fluorescens Ps. fluorescens was recovered from most organs as pure culture growth on standard bacteriological media, such as Pseudomonas F agar (Appendix 5.1), blood agar, TSA and nutrient agar, following incubation at 22-28°C for 24-28 h (Csaba et al, 1981b; Ahne et al, 1982). Pseudomonas pseudoalcaligenes Ps. pseudoalcaligenes was recovered as mixed cultures with Serratia plymuthica from the ascitic fluid and surface lesions following incubation on TSA at 22° C for 3-4 days (Austin and Stobie, 1992b). Vibrionaceae representatives Generally, vibrios may be isolated on marine 2216E agar (supplied by Difco) with incubation at 25°C for 2-7 days (e.g. Ishimaru et ai, 1996). Vibrio alginolyticus This may be readily achieved from blood by inoculation onto TSA prepared with seawater, TCBS or seawater agar with incubation at 15-25°C for 2-7 days. The precise conditions employed by Colorni et al. (1981) were not stated. However, this group succeeded in isolating V. alginolyticus, V. anguillarum and V. parahaemolyticus from blood, and long, thin, flexible rods from cases of gill rot. In addition, we have isolated pure culture growth of V. alginolyticus from moribund eels. Vibrio anguillarum The pathogen may be readily recovered from infected tissue by use of TSA (Traxler and Li, 1972), nutrient agar (Muroga et al, 1976a, b) and BHIA (Tajima et al, 1981) supplemented with sodium chloride at 0.5-3.5% (w/v), seawater agar and TCBS (Bolinches et al., 1988), with incubation at 15-25°C for periods of up to 7 days. The presumptive identification of V. anguillarum has been achieved using a specially designed medium, designated VAM, which combined bile salts with a high sodium chloride concentration, ampicillin, sorbitol and a high pH (Appendix 5.1;
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