副猪嗜血杆菌的MLST分析及数据库建立

Analysis and creation of database about MLST typing of haemophilus parasuis

对副猪嗜血杆菌86个分离株的7个看家基因，包括ATP合成酶J9链（atpD）、翻译起始因子IF-2（in-fB）、核糖体蛋白口亚基（rpoB）、苹果酸脱氢酶（mdh）、6-磷酸葡萄糖脱氢酶（6pgd）、3-磷酸甘油醛脱氢酶（g3pd）和延胡索酸还原酶B（frdB）进行了PCR扩增；对扩增产物进行了测序；对序列编辑、队列比对、建立MI．ST定义、等位基因序列文件等，与牛津大学动物系KeithJolley合作建立了副猪嗜血杆菌86个分离株的MI。ST数据库。经MLST分析发现所研究的分离株有高度的遗传异源性，86个菌株被分成74个序列类型。基因平均距离为0．674，h-0．804。每个基因位点存在64～76不等数目的等位基因。基因序列多态性位点最大（fedB）达325和（6pgd）219，最小（in，-B）为77；用UPGMA分析确定了5个菌群。

Seven housekeeping genes （fl chain of ATP synthase, （atpD）, translation initiation factor IF-2, （infB）, malate dehydrogenase gene, （mdh）, ribosomal protein subunit, rpoB, 6-phosphogluconate dehydrogenase, （6pgd）, glyceraldehyde-3-phosphate dehydrogenase, （g3pd） and fumarate reductase B, （frdB） were amplified and sequenced from 86 H. parasuis isolaties. Seven selected genes were sequenced from the 86 H. parasuis isolates. The Bioedit, Mega 4.0 and START Ilsoftwares were used to edit, assemble and align, define MLST schemes and allele squences profile. We and cooperated with Keith Jolley in department of zoology, university of Oxford eonstuctrued H. parasuis isolates PubMLST database and H. parasuis PubMLST locus/sequence difinitions database. The results showed that the high heterogeneity of H parasuis,ie. 86 isolates studied, were assigned to 74 STs, with a high level of diversity by MLST analysis. Allele average genetic distance was 0.674, IA =0.804. The numbers of alleles per locus were ranged from 64 to 76. The locus with the highest diversity were frdB （325） and 6pgd （219） ,and locus with the lowest polymorphy was infB （77）. Five different clusters were fixed by use of UPGMA.