A novel defensin‐like peptide from salivary glands of the hard tick,Haemaphysalis longicornis

A novel defensin‐like antimicrobial peptide named longicornsin was isolated from the salivary glands of the hard tick, Haemaphysalis longicornis, using a 10‐kDa cut‐off Centriprep filter and reversed‐phase high‐performance liquid chromatography (RP‐HPLC). Its amino acid sequence was determined as DFGCGQGMIFMCQRRCMRLYPGSTGFCRGFRCMCDTHIPLRPPFMVG by Edman degradation. The cDNA encoding longicornsin was cloned by cDNA library screening. The predicted protein from the cDNA sequence was composed of 78 amino acids including a mature longicornsin. It showed similarity with defensin‐like peptides from other ticks by BLAST search. Different from most other tick defensin‐like peptides, longicornsin had a C‐terminal extension. Purified longicornsin exerted potent antimicrobial activities against bacteria and fungi. Interestingly, it even showed strong antimicrobial ability against drug‐resistant microorganisms and Helicobacter pylori. The results of this study indicated that longicornsin is a potential candidate for novel antimicrobial drug design.     

Screening and Molecular Cloning of a Protective Antigen from the Midgut of Haemaphysalis longicornis

Vaccination is considered a promising alternative for controlling tick infestations. Haemaphysalis longicornis midgut proteins separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membrane were screened for protective value against bites. The western blot demonstrated the immunogenicity of 92 kDa protein (P92). The analysis of the P92 amino acid sequence by LC-MS/MS indicated that it was a H. longicornis paramyosin (Hl-Pmy). The full lenghth cDNA of Hl-Pmy was obtained by rapid amplification of cDNA ends (RACE) which consisted of 2,783 bp with a 161 bp 3'' untranslated region. Sequence alignment of tick paramyosin (Pmy) showed that Hl-Pmy shared a high level of conservation among ticks. Comparison with the protective epitope sequence of other invertebrate Pmy, it was calculated that the protective epitope of Hl-Pmy was a peptide (LEEAEGSSETVVEMNKKRDTE) named LEE, which was close to the N-terminal of Hl-Pmy protein. The secondary structure analysis suggested that LEE had non-helical segments within an α-helical structure. These results provide the basis for developing a vaccine against biting H. longicornis ticks.     

Ticks collected from selected mammalian hosts surveyed in the Republic of Korea during 2008-2009

A tick survey was conducted to determine the relative abundance and distribution of ticks associated with selected mammals in the Republic of Korea (ROK) during 2008-2009. A total of 918 ticks were collected from 76 mammals (6 families, 9 species) captured at 6 provinces and 3 Metropolitan Cities in ROK. Haemaphysalis longicornis (54.4%) was the most frequently collected tick, followed by Haemaphysalis flava (28.5%), Ixodes nipponensis (7.6%), Ixodes pomerantzevi (4.8%), Ixodes persulcatus (4.6%), and Haemaphysalis japonica (0.1%). Adults (57.0%) and nymphs (28.7%) of Ixodes and Haemaphysalis spp. were collected most frequently from medium or large mammals in this survey, while few larvae (14.3%) were collected. Hydropotes inermis was the most frequently captured mammal (52.6%), with a 16.4 tick index and 5 of 6 species of ticks collected during this survey. H. longicornis (69.7%) was the predominant tick collected from H. inermis, followed by H. flava (22.2%), I. persulcatus (6.1%), I. nipponensis (1.8%), and H. japonica (0.2%).     

Proteomic Screening of Antigenic Proteins from the Hard Tick,Haemaphysalis longicornis (Acari: Ixodidae)

Proteomic tools allow large-scale, high-throughput analyses for the detection, identification, and functional investigation of proteome. For detection of antigens from Haemaphysalis longicornis, 1-dimensional electrophoresis (1-DE) quantitative immunoblotting technique combined with 2-dimensional electrophoresis (2-DE) immunoblotting was used for whole body proteins from unfed and partially fed female ticks. Reactivity bands and 2-DE immunoblotting were performed following 2-DE electrophoresis to identify protein spots. The proteome of the partially fed female had a larger number of lower molecular weight proteins than that of the unfed female tick. The total number of detected spots was 818 for unfed and 670 for partially fed female ticks. The 2-DE immunoblotting identified 10 antigenic spots from unfed females and 8 antigenic spots from partially fed females. Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF) of relevant spots identified calreticulin, putative secreted WC salivary protein, and a conserved hypothetical protein from the National Center for Biotechnology Information and Swiss Prot protein sequence databases. These findings indicate that most of the whole body components of these ticks are non-immunogenic. The data reported here will provide guidance in the identification of antigenic proteins to prevent infestation and diseases transmitted by H. longicornis.     

Comparative microbiome analysis of Haemaphysalis longicornis ticks at the Korea Combat Training Center in 2022

The Korea Combat Training Center (KCTC), located in Gangwon Province, is a restricted military training facility where research on the environmental conditions and health risks to military personnel has been limited. In this study, using iSeq 100, we investigated the bacterial abundance and microbiome of Haemaphysalis longicornis specimens collected at the KCTC from June to August 2022, to assess current and potential public health risks to military personnel. Our results show that adult ticks had significantly greater species richness compared with larvae and nymphs, with no notable differences in diversity across developmental stages. Principal coordinate analysis of the microbial communities did not show differences attributable to any single factor, such as collection location or date. Coxiella-like endosymbionts (AB001519) were identified in all 13 samples, and Jatrophihabitans, Sphingomonas, and Spirosoma were consistently found across all samples. In addition, iSeq 100 also identified Rickettsia rickettsii and Borrelia spp., which were not detected with conventional polymerase chain reaction (PCR).     

Detection of tick‐borne Anaplasma bovis,Anaplasma phagocytophilum and Anaplasma centrale in Spain

The genus Anaplasma (Rickettsiales: Anaplasmataceae) includes species of medical and veterinary importance. The presence of Anaplasma spp. in ticks from birds, as well as in Haemaphysalis punctata (Ixodida: Ixodidae) specimens collected from cattle and vegetation in northern Spain was investigated. A total of 336 ticks from birds [174 Ixodes frontalis (Ixodida: Ixodidae), 108 H. punctata, 34 Hyalomma marginatum (Ixodida: Ixodidae), 17 Ixodes ricinus (Ixodida: Ixodidae) and three Ixodes spp.], and 181 H. punctata specimens collected from cattle (n = 71) and vegetation (n = 110) were analysed. Anaplasma bovis was detected in five H. punctata, including two from birds (1.9%) and three from vegetation (2.7%). Four I. frontalis (2.3%) (one co‐infected with ‘Candidatus Midichloria mitochondrii’) and one I. ricinus (5.9%) removed from birds, as well as four H. punctata (5.6%) collected from cattle showed Anaplasma phagocytophilum infection. In addition, Anaplasma centrale was found in two H. punctata, one from a cow (1.4%) and the other from vegetation (0.9%). This study represents the first evidence of the presence of A. bovis in European ticks, and reports the first detection of A. bovis and A. centrale in H. punctata, and the first finding of A. phagocytophilum and ‘Ca. Midichloria mitochondrii’ in I. frontalis.     

Seasonal dynamics of the tick Haemaphysalis tibetensis in the Tibetan Plateau,China

The tick Haemaphysalis tibetensis (Acari: Ixodidae) Hoogstraal is an important arthropod vector widespread in the Qinghai‐Tibet Plateau, and knowledge of its seasonal dynamics is still poor. The current study investigated the seasonal dynamics of the parasitic and non‐parasitic H. tibetensis over a 2‐year period from March 2014 to February 2016 in the Tibetan Plateau, China. During this timeframe, non‐parasitic ticks were collected weekly by flag‐dragging in grassland and shrubland areas, and parasitic ticks were removed weekly from selected sheep. Plateau pikas were captured using traps and examined for immature ticks between May to September 2014. Results suggest that non‐parasitic H. tibetensis were mainly distributed in the grassland, and the parasitic adults and nymphs were found mostly on sheep. Larvae were usually found on Plateau pikas and the prevalence of infestation and mean parasitic intensity were 72.1 and 1.81%, respectively. Adults were observed from March to July with the major peak occurring in mid‐April. Nymphs were found from March to August and reached a peak in late June. Larvae were collected from April to September, and their numbers peaked in late May. In the parasitic and non‐parasitic period, the overall sex ratio of males to females was 1.62 and 1.30, respectively. Results show that H. tibetensis can complete one generation per year, with a population overlap between stages over the spring‐summer months. These findings provide additional information on the biology and ecology of H. tibetensis as well as insights on its control in the environment and on sheep.     

Immunization effect of recombinant P27/30 protein expressed in Escherichia coli against the hard tick Haemaphysalis longicornis (Acari: Ixodidae) in rabbits

We investigated the induction of resistance to Haemaphysalis longicornis infestation in rabbits that had been immunized with recombinant H. longicornis P27/30 protein. The success of immunological control methods is dependent upon the use of potential key antigens as tick vaccine candidates. Previously, we cloned a gene encoding 27 kDa and 30 kDa proteins (P27/30) of H. longicornis, and identified P27/30 as a troponin I-like protein. In this study, rabbits that were immunized with recombinant P27/30 expressed in Escherichia coli showed the statistically significant longer feeding duration for larval and adult ticks (P<0.05), low engorgement rates in larval ticks (64.4%), and an apparent reduction in egg weights, which suggest that H. longicornis P27/30 protein is a potential candidate antigen for a tick vaccine. These results demonstrated that the recombinant P27/30 protein might be a useful vaccine candidate antigen for biological control of H. longicornis.     

实验室条件下长角血蜱的生物学特性研究

实验室条件[(27±1)℃;75%RH;6L：18D]下,长角血蜱完成其生活史需135.8d：卵的孵化期为38.5d：幼虫吸血前期、吸血期和蜕皮前期分别为5.3d,3.8d和13.9d;若虫吸血前期、吸血期和蜕皮前期分别为7.2d,5.4d和16.9d;成虫吸血前期、吸血期、产卵前期和产卵期分别为7.6d,9.4d,7.8d和20.0d。雌虫饱食体重与产卵量之间存在非常显著的正相关(r＝0.9496,P<0.001)。雌虫的生殖效率指数REI＝11.06,生殖适合度指 数RFI＝7.19。生活周期在不同季节无明显变化。雄虫吸血期受雌虫吸引而成功地交配,这一过程是在吸引性信息素的作用下完成的。