DNA polymorphism in mitochondrial NADH dehydrogenase subunit-1 within and among Toxocara and Toxascaris species

Китайский. 对弓首属犬弓首蛔虫、猫弓首蛔虫、马来西亚弓首蛔虫、牛弓首蛔虫和狮弓属狮弓蛔虫线粒体DNA（mtDNA）中烟酰胺脱氢酶亚基I（nad1）基因部分序列（pnad1）进行比较研究，分析它们之间的序列差异和种群遗传关系。先用同位素（γ33P）标记上下游引物，通过PCR方法扩增出单个虫体线粒体pnad1片段，然后采用单链构象多态性（SSCP）分析技术对pnad1片段进行多态性研究，最后挑选出代表性样品进行测序，并利用基因分析软件DNAStar (版本5.01)对序列进行分析比较。犬弓首蛔虫种内pnad1基因序列差异为0.3%～1.9%，与猫弓首蛔虫、马来亚弓首蛔虫、牛弓首蛔虫和狮弓蛔虫的种间序列差异分别为11.0%～14.1%、10.7%～12.4%、12.6%～13.7%和17.5%～18.2%。猫弓首蛔虫种内pnad1基因序列差异为2.8%，与马来西亚弓首蛔虫、牛弓首蛔虫和狮弓蛔虫的种间序列差异分别是12.0%～13.0%、15.1%和18.6%～21.2%。马来西亚弓首蛔虫种内pnad1基因序列差异为0～0.3%，与牛弓首蛔虫和狮弓蛔虫的种间序列差异分别是12.4%～12.8%和18.6%～19.0%。犬弓首蛔虫、猫弓首蛔虫和马来西亚弓首蛔虫不同地方虫株的pnad1序列都有一定差异（0～2.8%）。但种间pnad1的序列差异（10.7%～21.2%）明显高于种内的序列差异，说明nad1基因可以作为弓首蛔虫和狮弓蛔虫分子分类及种群遗传关系研究的遗传标记。

Английский. The objectives of the present study were to examine sequence variation in a portion of the mitochondrial NADH dehydrogenase subunit 1 gene (pnad1) within and among Toxocara canis, T. cati, T. malaysiensis, T. vitulorum and Toxascaris leonina, and to study their systematic and population genetics. Total genomic DNA was extracted from individuals of Toxocara and Toxascaris species from different geographical locations. The pnad1 of each sample was amplified by a polymerase chain reaction (PCR) using primers end-labelled with γ33P-ATP. Sequence variation in the pnad1 was examined and compared among these nematodes using single-strand conformation polymorphism (SSCP) analysis, followed by DNA sequencing. While the average intraspecific variations in the pnad1 sequences were 0.3%-1.9% for T. canis, 2.8% for T. cati, 0-0.3% for T. malaysiensis from different geographical origins, the interspecific differences were 11.0%-14.1% between T. canis and T. cati, 10.7%-12.4% between T. canis and T. malaysiensis, 12.6%-13.7% between T. canis and T. vitulorum, 17.5%-18.2% between T. canis and T. leonina, 12.0%-13.0% between T. cati and T. malaysiensis, 15.1% between T. cati and T. vitulorum, 18.6%-21.2% between T. cati and T. leonina, 12.4%-12.8% between T. malaysiensis and T. vitulorum, and 18.6%-19.0% between T. malaysiensis and T. leonina. The findings of this study showed that the intras-pecific variations in pnad1 within these ascaridoid species were subtle, and significantly lower than interspecific differences, indicating that the pnad1 provides genetic markers for the identification of these ascaridoid nematodes, and should have an important implication for studying systematic, population genetic structures and the molecular ecology of Toxocara and Toxascaris species.