Identification of genes in a KG- phenotype of Lactococcus garvieae, a fishpathogenic bacterium, whose proteins react with anti-KG- rabbit serum

Five different clones (SA1B05, SA1B10, SA2F01, SA8A11 and SA9H10) were isol ated from the gene library of the Lactococcus garvieae SA8201 (KG-) strain by immunological screening using rabbit serum against L. garvieae (KG-) phe notype cells. A Western blot analysis indicated that the molecular sizes of immunologically detected proteins of SA1B05, SA1B10, SA2F01, SA8A11 and SA 9H10, which were fused with LacZ protein, were 25, 30, 23, 26 and 13 kDa, r espectively. The amino acid sequences of the immunologically detected prote ins of SA1B05, SA1B10, SA2F01 and SA8A11 were homologous to a processing pr otease of Bacillus subtilis (36.6%), dihydropteroate synthase of Escherichi a coli (34.6%), trigger factor of B. subtilis (45.8%) and N-acetylglucosami ne-6-phosphate deacetylase of Vibrio furnissii (37.1%), respectively. There was no significant homologous sequence of SA9H10 in DDBJ/EMBL/GenBank and SwissProt. We cloned and sequenced a longer DNA fragment (SASH10L) of SA9H1 0 from the gene library. The predicted amino acid sequence of this clone wa s weak homology to M protein of Streptococcus pyogenes (22.7%). Five genes were specifically expressed in the KG- phenotype strains. However, SA8A11 a nd SA9H10 was expressed in the mutated strain SA8201-TTC, whose serological phenotype was changed from KG- to KG+ by 2,3,5-triphenyltetrazolium chlori de. (C) 1999 Academic Press.