Photorhabdus insect-related (Pir) toxin-like genes in a plasmid of Vibrio parahaemolyticus, the causative agent of acute hepatopancreatic necrosis disease (AHPND) of shrimp.
Photorhabdus insect-related (Pir) toxin-like genes in a plasmid of Vibrio parahaemolyticus, the causative agent of acute hepatopancreatic necrosis disease (AHPND) of shrimp.
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DOI：
10.3354/dao02830
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发表时间：
2015-02-10
影响因子：
1.4
通讯作者：
Lightner DV
中科院分区：
文献类型：
--
作者：
Han JE;Tang KF;Tran LH;Lightner DV
从引起急性肝胰腺坏死病（AHPND）的副溶血性弧菌（Vibrio parahaemolyticus）13-028/A3菌株中鉴定出69 kb质粒pVPA 3 -1。这种疾病是造成养殖对虾大量死亡的原因，被称为早期死亡综合征（EMS）。该质粒的GC含量为45.9%，每个细菌细胞的拷贝数为37，这是通过比较定量PCR分析确定的。它由92个开放阅读框组成，编码动员蛋白、复制酶、转座酶、毒力相关蛋白和类似于光杆状病毒昆虫相关（Pir）毒素的蛋白。在副溶血性弧菌中，这些Pir毒素样蛋白由位于3.5 kb片段内的2个基因（pirA样和pirB样）编码，该片段侧翼为转座酶编码序列的反向重复序列（1 kb）。这两个基因的GC含量仅为38.2%，显著低于质粒的其余部分，这表明这些基因是最近获得的。基于蛋白质组学分析，pirA样（336 bp）和pirB样（1317 bp）基因分别编码13和50 kDa的蛋白质。在副溶血性弧菌13-028/A3的实验室培养物中，两种蛋白质均分泌到培养基中。我们开发了一种双重PCR诊断方法，检测限为105 CFU ml−1，并靶向该副溶血性弧菌菌株中的pirA和pirB样基因。该PCR方案可以可靠地检测引起AHPND的副溶血性弧菌菌株，并且不与非致病性菌株或从虾池中分离的其他种类的弧菌交叉反应。
The 69 kb plasmid pVPA3-1 was identified in Vibrio parahaemolyticus strain 13-028/A3 that can cause acute hepatopancreatic necrosis disease (AHPND). This disease is responsible for mass mortalities in farmed penaeid shrimp and is referred to as early mortality syndrome (EMS). The plasmid has a GC content of 45.9% with a copy number of 37 per bacterial cell as determined by comparative quantitative PCR analyses. It consists of 92 open reading frames that encode mobilization proteins, replication enzymes, transposases, virulence-associated proteins, and proteins similar to Photorhabdus insect-related (Pir) toxins. In V. parahaemolyticus, these Pir toxin-like proteins are encoded by 2 genes ( pirA- and pirB-like) located within a 3.5 kb fragment flanked with inverted repeats of a transposase-coding sequence (1 kb). The GC content of these 2 genes is only 38.2%, substantially lower than that of the rest of the plasmid, which suggests that these genes were recently acquired. Based on a proteomic analysis, the pirA-like (336 bp) and pirB-like (1317 bp) genes encode for 13 and 50 kDa proteins, respectively. In laboratory cultures of V. parahaemolyticus 13-028/A3, both proteins were secreted into the culture medium. We developed a duplex PCR diagnostic method, with a detection limit of 105 CFU ml−1 and targeting pirA- and pirB-like genes in this strain of V. parahaemolyticus. This PCR protocol can reliably detect AHPND-causing strains of V. parahaemolyticus and does not cross react with non-pathogenic strains or with other species of Vibrio isolated from shrimp ponds.