Or bacteriological analyzes. A total of 41 diseased fish were collected, including 10 3 of farm 1, farm 2 6, and 25 of P450 Inhibitors the farm 2.2. Isolation and biochemical characterization For bacterial isolation, samples of brain, kidney, liver, and ascites fluid from diseased fish sampled for fa Aseptic is distributed on a sheep blood agar, and at 25 8C for 72 h Pure colonies were a Gramf Staining, catalase and oxidase tests for the presence of H Thermolysis and growth at different temperatures and on MacConkey agar. The St Strains were at 80 8C in brain heart broth held at 15% glycerol until use. Before the biochemical and molecular tests, the isolates were cultured on Rogosa & Sharpe agar at 25 8C man for 24 h. Biochemical characterization was performed using the Rapid ID32 Strep commercial kit. 2.
3. Molecular analysis of the amplification and phylogenetic tree construction and sequential Age of 16S rRNA gene were for nine Feeder Llig selected Hlten Isolates from the total number of isolates performed. Total DNA was prepared using the commercial kit DNeasy. 16S rRNA was amplified by PCR with primers verst for the universal C70 and B37 Are RKT, as described by Fox et al. The PCR products were purified using a Wizard PCR Preps kit and sequenced using preheating Rts and Rev Rts primers. The sequential reactions Ages were recorded with a terminator cycle sequencing kit BigDyeTM Age and run on an ABI 3730xl Genetic Analyzer. The sequences were then compared to sequences in the NCBI database using BLAST algorithm. The limit for the identification of a bacterial species was 98% identity t of the nucleotides of the 16S rRNA gene.
The phylogenetic relatedness of the isolates was determined by comparison of 16S rRNA sequence analysis of the gene. The sequences of the isolates were aligned with ClustalW with BioEdit sequence of the following bacterial species: Weissella paramesenteroides NRIC 1542, Weissella thailandensis FS61 1, 2973 NCFB hellenica Weissella, Weissella confusa JCM1093, Weissella cibaria LMG 17 699, NRIC 1536 viridensces Weissella, Weissella minor, NRIC 1625, 1627 NRIC halotolerans Weissella, Weissella kandleri NCFB 2753, S 5623 koreensis Weissella, Weissella soli LMG 20 113, LMG 25373T beninensis Weissella, Weissella ghanensis LMG 24 286, 257T fabaria Weissella, Pediococcus, and Weissella sp damnosus strain fish. JZ 1L.
The genetic distance matrix was performed using the Kimura two parameter models, and an evolution Rer tree was with the neighbor joining method MEGA4. Bootstrap values of 1000 are replicated as percentages. Weissella genus-specific PCR was performed for all isolates using the primers and Weir Welf folllowing Jang et al. with some modifications. Briefly, the amplification with a denaturing cycle at 95 8C for 5 min by 30 cycles of 95 8C series for 30 s, 55 8C for 45 s and carried out for 72 8C for 1 min in advance, min with a final extension 7 at 72 8C. 2.4. Antimicrobial susceptibility testing of pathogen resistance to five hours Ufigsten has used antibiotics in aquaculture has been identified globally for all isolates using the disk diffusion tests. The tests were carried out in accordance with M42-A guidelines recommended by all Changes to the bacteria streptococcus. Hard drives with the antimicrobi