Obviously caution is needed in interpreting serological diagnoses Whenever poss

Obviously caution is needed in interpreting

This preview shows page 231 - 233 out of 594 pages.

Obviously, caution is needed in interpreting serological diagnoses. Whenever poss- ible, culturing should be used for confirmation. Certainly, the selective medium (SKDM; Austin et al, 1983a) has proved useful in isolating Renibacterium from mixed cultures. In addition, during a comparative exercise with KDM2, SKDM consistently enabled a greater recovery of cells from infected fish. In some cases, scant growth of only one or two colonies were recovered from kidney tissue on SKDM, although the parallel KDM2 plates were devoid of any growth (Austin et al, 1983a). Suspect cultures of renibacteria were subsequently confirmed by the characteristic profile on API-zym and other phenotypic traits, namely catalase pro- duction and inabihty to produce oxidase. Whereas debate has centred over the most effective means of detecting BKD, it may be concluded that effective diagnosis should encompass a multiplicity of methods. These include isolation and characterisation, and serology on infected tissue. It is proposed that cHnical cases of disease should be examined by FAT and culturing methods. Asymptomatic cases should be the subject of full bacterio- logical examination.
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210 Bacterial Fish Pathogens WHICH METHOD IS BEST?—FURUNCULOSIS In a comparative study of serodiagnostic techniques, Sakai et al. (1986) reported that iFAT and the peroxidase-antiperoxidase enzyme immunoassay (PAP) were more sensitive (capable of detecting 10^ CFU/ml) than the latex agglutination and co- agglutination techniques. These required 10^ CFU/ml for positive results to be recorded. Nevertheless, with latex agglutination and co-agglutination techniques more (15/15 = 100%) positive samples were detected than by iFAT (10/15 = 67%) or PAP (1/15 = 73%). MOLECULAR TECHNIQUES There is evidence that molecular techniques have been used with increasing regularity for bacterial pathogens. A timely overview of PCR with emphasis on vaHdation of the techniques and problems relating to diagnosis has been published (Hiney and Smith, 1998). A noteworthy advance resulted from the use of PCR technology to identify Mycobacterium spp. in sea bass (Knibb et al, 1993) and Myc. chelonei in a cichlid oscar (Astronotus ocellatus) (McCormick et al., 1995). A PCR followed by reverse cross blot hybridisation identified Mycobacterium sensitively (to lOOfg of DNA, which equated to 20 mycobacterial cells) to the species level (Puttinaowarat et al., 2002). A PCR using a llOObp fragment has distinguished Lactococcus garvieae from Lactococcus lactis (Zlotkin et al., 1998). In terms of sensitivity, the PCR detected Lactococcus garvieae in 1 |il of fish plasma. Another publication reported a PCR based on a 709 bp fragment that was specific for Lactococcus garvieae, enabUng a positive result to be obtained in 4h (Aoki et al, 2000). A multiplex PCR has reportedly been developed, and successfully recognised—from culture and fish tissues—the fish-pathogenic lactococci-streptococci, i.e. Lactococus garvieae, Str.
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