Mitochondrial Gene Heterogeneity and Population Genetics of Haemaphysalis longicornis (Acari: Ixodidae) in China.

INTRODUCTION: Haemaphysalis longicornis is an important ectoparasite of domestic and wild animals that can transmit many pathogens including viruses, fungi, bacteria and protozoa. MATERIALS AND METHODS: In this study, we examined genetic variation and population genetics in three mitochondrial (mt) genes [cox1 (cytochrome c subunit 1), rrnL (large subunit ribosomal RNA) and nad5 (NADH dehydrogenase 5)] among four H. longicornis populations from China. RESULTS: The sizes of the partial sequences of cox1, rrnL and nad5 were 776 bp, 409 bp, 510 bp, respectively. Among the obtained sequences, we identified 22 haplotypes for cox1, 2 haplotypes for rrnL and 17 haplotypes for nad5. Low gene flow and significant genetic differentiation (66.2%) were detected among H. longicornis populations. There was no rapid expansion event in the demographic history of four H. longicornis populations in China. In addition, phylogenetic analyses confirmed that all the Haemaphysalis isolates were H. longicornis which were segregated into two major clades. CONCLUSION: The mt DNA genes provide a potential novel genetic marker for molecular epidemiology of H. longicornis and assist in the control of tick and tick-borne diseases in humans and animals.

First Report of Ixodes nipponensis Infection in Goats in China.

Ticks are obligate blood-sucking ectoparasites that infect a wide range of animals and humans, causing a variety of both human and animal diseases around the world. Ixodes nipponensis is the most commonly reported tick in Korea and Japan, but it is very rare in China. In this study, six I. nipponensis samples were collected from three black goats in Hunan province, China. Ticks identified morphologically as I. nipponensis were then examined by PCR with two different molecular markers: mitochondrial cox1 and the second internal transcribed spacer of ribosomal DNA genes. Sequence comparison and phylogenetic analysis of the cox1 sequences confirmed that all of the examined hard Ixodes ticks represented I. nipponensis. This finding indicates a potential risk of zoonotic I. nipponensis infection in humans and animals in China. To our knowledge, this is the first report documenting the occurrence of I. nipponensis infection in goats in China.

Molecular characterization of hard tick Haemaphysalis longicornis from China by sequences of the internal transcribed spacers of ribosomal DNA.

In the present study, the entire first and second internal transcribed spacer (ITS-1 and ITS-2) regions of nuclear ribosomal DNA (rDNA) of Haemaphysalis longicornis from China were amplified by polymerase chain reaction. The 45 representative amplicons were sequenced, and sequence variation in the ITS was examined. The ITS sequences of H. longicornis were 3644 bp in size, including the part of 18S rDNA, 28S rDNA sequences and the complete ITS-1, 5.8S rDNA and ITS-2 sequences. Sequence analysis revealed that the ITS-1, 5.8S rDNA and ITS-2 of this hard tick were 1582, 152, and 1610 bp in size, respectively. The intra-specific sequence variations of ITS-1 and ITS-2 within H. longicornis were 0-2 and 0-2.2%; however, the inter-specific sequence differences among members of the genus Haemaphysalis were significantly higher, being 35.1-55.2 and 37-52% for ITS-1 and ITS-2, respectively. The molecular approach employed in this study provides the foundation for further studies of the genetic variation of H. longicornis from different hosts and geographical origins in China.

Genetic variation in mitochondrial genes of the tick Haemaphysalis flava collected from wild hedgehogs in China.

The tick Haemaphysalis flava (Acari: Ixodidae) is an important ectoparasite, which causes direct damage to their hosts and also acts as a vector of various infectious disease agents in China. Despite its significance, the epidemiology, genetics and biology of H. flava has not been studied in detail. In the present study, the genetic variation in three mitochondrial (mt) DNA regions, namely cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 1 and 4 (nad1 and nad4), was examined in H. flava ticks collected from wild hedgehogs in China. A portion of cox1 (pcox1), nad1 (pnad1) and nad4 (pnad4) genes were PCR amplified from individual H. flava ticks and the amplicons were sequenced. The length of the sequences of pcox1, pnad1 and pnad4 were 849, 285 and 626 bp, respectively. The intra-specific sequence variation within H. flava was 0-0.4% for pcox1, 0-0.4% for pnad1 and 0-0.3% for pnad4. However, the inter-specific variation was significantly higher, 12.5-14.3%, 13.6-24.8% and 14.8-19% for pcox1, pnad1 and pnad4, respectively. Phylogenetic analysis based on Maximum likelihood (ML) method using the combined target mt gene sequences confirmed that all isolates of Haemaphysalis were H. flava. The molecular approach employed in this study provides a tool for further elucidating the molecular diversity of H. flava in China and elsewhere in Asia.