Genome organization of the Kresse strain of porcine parvovirus: Identification of the allotropic determinant and comparison with those of NADL-2 and field isolates
Genome organization of the Kresse strain of porcine parvovirus: Identification of the allotropic determinant and comparison with those of NADL-2 and field isolates
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DOI：
10.1128/jvi.70.4.2508-2515.1996
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发表时间：
1996-04-01
影响因子：
5.4
通讯作者：
Tijssen, P
中科院分区：
文献类型：
--
作者：
Bergeron, J;Hebert, B;Tijssen, P
将猪腮腺病毒（PPV）克雷斯（Kresse）株克隆到pUC19中，并进行了独立的感染性克隆测序。具有不同致病性的PPV Kresse和NADL-2株具有相同的基因组组织和高度的序列同源性。在实验室菌株序列差异显著的区域，用PCR扩增了15株田间分离株的部分基因组（1.5或1.6 kb）。在Kresse菌株和毒田分离株的VP1/VP2编码区一致存在5个氨基酸差异。在田间分离株的基因组中也发现了一些不一致的点突变。此外，在那些具有疫苗氨基酸谱的病毒中，除了一个分离株（iff -3）外，其余的都含有一个127 bp的衣壳蛋白基因下游的非编码直接重复序列。唯一的例外也是唯一从木乃伊胎儿中获得的疫苗型PPV。为了鉴定导致Kresse菌株明显趋向性（以及可能的病理）的遗传因素，建立了将毒力与疫苗菌株区分开来的体外细胞系统。随后，利用Kresse和NADL-2株的嵌合感染克隆鉴定了位于VP1/VP2区的同素异形体决定因子。将含有三个氨基酸差异的Kresse基因组的BglII片段转移到NADL-2背景或相反的结构中，导致目标基因组的表型恢复到BglII片段的亲本菌株的表型。基于犬细小病毒衣壳结构对氨基酸差异定位的预测表明，它们都位于病毒粒子的外表面或靠近外表面。特别是，一个突变（S-436—>P）的位置通过类比映射到衣壳最容易接近的三倍尖峰区域。
The Kresse strain of porcine parovirus (PPV) was cloned into pUC19, and independent infectious clones were sequenced. The PPV Kresse and NADL-2 strains, which have different pathogenicities, shared an identical genomic organization and a high degree of sequence identity. Partial genomes (1.5 or 1.6 kb) of 15 field isolates were also amplified by PCR in regions with significant sequence differences between the laboratory strains. Five amino acid differences were consistently present within the VP1/VP2 coding region of the Kresse strain and virulent field isolates. A number of inconsistent point mutations were also found throughout the genomes of field isolates. In addition, among those with the vaccine amino acid profile, all but one isolate (IAF-3) contained a 127-bp noncoding direct repeat downstream of the capsid protein gene. The one exception was also the only vaccine-type PPV obtained from a mummified fetus. In order to identify genetic elements responsible for the distinct tropism (and possibly the pathology) of the Kresse strain, in vitro cell systems which differentiated the virulent from the vaccinal strains were established. Subsequently, chimeric infectious clones of the Kresse and NADL-2 strains were used to identify the allotropic determinant located in the VP1/VP2 region. The transfer of the BglII fragment of the Kresse genome, containing three amino acid differences, into the NADL-2 background, or the opposite construct, caused the phenotype of the target genome to revert to that of the parent strain of the BglII fragment. Prediction of the localization of amino acid differences on the basis of canine parvovirus capsid structure indicates that each is located on or near the outer surface of the virion. In particular, the position of one mutation (S-436-->P) maps by analogy to the threefold spike, the most accessible region of the capsid.