永州地区犬球虫的感染情况调查、虫种鉴定及种系发育关系研究

Infection investigation,species identification,and study on the phylogenetic relationship of canine coccidia in Yongzhou area

为了了解永州地区犬球虫的感染情况,确定优势虫种,并探究不同球虫虫种之间的种系发育关系,试验利用饱和重铬酸钾溶液漂浮法对采自永州地区的362份犬粪便进行虫卵检测,通过宏基因组测序技术鉴定虫种,PCR扩增球虫的18S r RNA、ITS-1基因,经测序、比对确定永州地区流行的优势虫种,并对其种系发育关系进行分析。结果表明:有124份犬粪便为阳性,总体感染率为34.25%,其中宠物犬的感染率为24.31%,流浪犬的感染率为44.20%;在永州地区有犬等孢球虫与俄亥俄等孢球虫流行,其中犬等孢球虫33株(占比26.61%),俄亥俄等孢球虫91株(占比73.39%);犬等孢球虫18S r RNA、ITS-1基因序列的长度分别为738 bp与1000 bp,俄亥俄等孢球虫18S r RNA、ITS-1基因序列的长度分别为1051 bp和398 bp。两种犬球虫18S r RNA、ITS-1基因序列的相似性分别为98.53%~98.76%和99.58%~99.72%。此外,在基于18S r RNA、ITS-1基因序列所组建的2个种系发育树中,犬等孢球虫与俄亥俄等孢球虫均处于每个种系发育树的同一分支上。说明在永州地区犬球虫的感染率较高,俄亥俄等孢球虫为该地区的优势虫种,且犬等孢球虫与俄亥俄等孢球虫之间存在较近的亲缘关系。

In order to understand the infection of canine coccidia in Yongzhou area,to identify the dominant species,and to investigate the phylogenetic relationship between different coccidia species,in the test,the floating method of potassium dichromate solution was used to detect eggs in 362 dog feces collected from Yongzhou area;the species were identified by metagenome sequencing technology;the ribosomal 18S r RNA and ITS-1 genes of coccidia were amplified by PCR;and the dominant species prevalent in Yongzhou area were identified by sequencing and comparison,and their phylogenetic relationship were analyzed.The results showed that 124 canine feces were positive,with an overall infection rate of 38.27%;the infection rate was 27.16%in pet dogs and 49.38%in stray dogs.In Yongzhou area,there were 2 types of canine coccidia,Cystoisospora canis and Cystoisospora ohioensis,of which 33 strains of Cystoisospora canis(26.61%)and 91 strains of Cystoisospora ohioensis(73.39%)were prevalent.The lengths of the 18S r RNA and ITS-1 gene sequences of Cystoisospora canis were 738 bp and 1000 bp,respectively;the lengths of the 18S r RNA and ITS-1 gene sequences of Cystoisospora ohioensis were 1051 bp and 398 bp,respectively.The homology of the 18S r RNA and ITS-1 gene sequences of the two species of canine coccidia was 98.53%to 98.76%and 99.58%to 99.72%,respectively.In addition,in the two phylogenetic trees based on the 18S r RNA and ITS-1 gene sequences,Cystoisospora canis and Cystoisospora ohioensis were in the same branch of each phylogenetic tree.The results suggested that the infection rate of canine coccidia was high in Yongzhou area,and Cystoisospora canis was the dominant species in this area,and there was a close relationship between Cystoisospora canis and Cystoisospora ohioensis.