Identification of Haemaphysalis longicornis, H.flava and H.campanulata based on molecular markers of 16S rDNA and CO Ⅰ gene
Objective To establish a molecular biological method for identifying Haemaphysalis longicornis,H.flava and H.campanulata with markers of 16S rDNA gene and cytochrome oxidase subunit Ⅰ gene (CO Ⅰ),and analyze the phylogenetic relationship of these three species of ticks.Methods Ticks were collected from the body of animals in Shanghai,and identified by their morphological characteristics under the dissecting microscope.Then the genomic DNA was extracted from these samples and the 16S rDNA and CO Ⅰ gene were amplified by PCR from the genomes of the three species of ticks and they were sequenced for homology analysis.Mega 6.0 software was used to construct the phylogenetic trees of the gene sequences for phylogenetic analysis.Results In this three species of Haemaphysalis ticks,the identities of the 16S rDNA gene and CO Ⅰ gene of H.longicornis to the 16S rDNA gene and CO Ⅰ gene of other isolates in GenBank were 96.0％～97.2％ and 96.9％～98.8％,respectively.The identities of the 16S rDNA gene and CO Ⅰ gene of H.flava to the 16S rDNA gene and CO Ⅰ gene of other isolates in GenBank were 95.9％～98.3％ and 87.1％～91.9％,respectively.The identities of the 16S rDNA gene of H.campanulata to that from Japan (AB819170) in GenBank was 97.5％.The identities between the three species of Haemaphysalis ticks were 88.0％,87.0％,86.9％ for 16S rDNA and 80.4％,80.8％,82.8％ for CO Ⅰ.The phylogenetic trees were constructed with the nucleotide sequence of 16S rDNA and CO Ⅰ gene among three species of ticks and other Haemaphysalis genus ticks,respectively.The results showed that H.longicornis and H.flava were clustered together respectively and they all had formed independent branches.Conclusions It can more accurately identify ticks species by combination of molecular biological identification methods and traditional morphological classification.
NETL
NSTL
万方数据
