Seco-Mediavilla Patricia, Verger Jean-Michel, Grayon Maggy, Cloeckaert Axel, Marín Clara M, Zygmunt Michel S, Fernández-Lago Luis, Vizcaíno Nieves
Departamento de Microbiología y Genética, Universidad de Salamanca, 37007 Salamanca, Spain.
Clin Diagn Lab Immunol. 2003 Jul;10(4):647-51. doi: 10.1128/cdli.10.4.647-651.2003.
Sequencing of bp26, the gene encoding the Brucella sp. immunogenic BP26 periplasmic protein, was performed in the reference strains of Brucella abortus, B. suis, and B. ovis. The three bp26 sequences were almost identical to that published for B. melitensis 16M bp26, and only minor nucleotide substitutions, without modifying the amino acid sequence, were observed between species. The bp26 genes of the seven B. abortus biovar reference strains and B. abortus S19 and RB51 vaccine strains were also sequenced. Again, only minor differences were found. Surprisingly, the bp26 nucleotide sequence for B. abortus S19 was almost identical to that found for B. melitensis 16M and differed from the sequence described previously by others (O. L. Rossetti, A. I. Arese, M. L. Boschiroli, and S. L. Cravero, J. Clin. Microbiol. 34:165-169, 1996) for the same B. abortus strain. The epitope mapping of BP26, performed by using a panel of monoclonal antibodies and recombinant DNA techniques, allowed the identification of an immunodominant region of the protein interesting for the diagnosis of B. melitensis and B. ovis infection in sheep. A recombinant fusion protein containing this region of BP26 reacted indeed, in Western blotting, as the entire recombinant BP26 against sera from B. melitensis- or B. ovis-infected sheep while it avoided false-positive reactions observed with sera from Brucella-free sheep when using the entire recombinant BP26. Thus, use of this recombinant fusion protein instead the entire recombinant BP26 could improve the specific serological diagnosis of B. melitensis or B. ovis infection in sheep.
对编码布鲁氏菌属免疫原性周质蛋白BP26的bp26基因进行了测序，所用菌株为流产布鲁氏菌、猪布鲁氏菌和绵羊布鲁氏菌的参考菌株。这三个bp26序列与已发表的羊种布鲁氏菌16M bp26序列几乎相同，不同种之间仅观察到少量核苷酸替换，且未改变氨基酸序列。还对7株流产布鲁氏菌生物变种参考菌株以及流产布鲁氏菌S19和RB51疫苗株的bp26基因进行了测序。同样，仅发现了微小差异。令人惊讶的是，流产布鲁氏菌S19的bp26核苷酸序列与羊种布鲁氏菌16M的序列几乎相同，与其他人先前描述的同一流产布鲁氏菌菌株（O. L. Rossetti、A. I. Arese、M. L. Boschiroli和S. L. Cravero，《临床微生物学杂志》34:165 - 169，1996年）的序列不同。通过使用一组单克隆抗体和重组DNA技术对BP26进行表位定位，确定了该蛋白的一个免疫显性区域，该区域对诊断绵羊的羊种布鲁氏菌和绵羊布鲁氏菌感染具有重要意义。在蛋白质印迹法中，含有BP26这一区域的重组融合蛋白确实与来自羊种布鲁氏菌或绵羊布鲁氏菌感染绵羊的血清发生反应，就如同整个重组BP26一样，而当使用整个重组BP26时，它避免了与未感染布鲁氏菌的绵羊血清出现假阳性反应。因此，使用这种重组融合蛋白而非整个重组BP26可以改善绵羊羊种布鲁氏菌或绵羊布鲁氏菌感染的特异性血清学诊断。
