Growth of Haemaphysalis (Kaiseriana) longicornis on cattle.

Increases were recorded of lengths of larvae, nymphs and females of the tick, Haemaphysalis (Kaiseriana) longicornis during feeding on cattle, so that categories could be established for estimating the ages of ticks in natural populations. Most larvae and nymphs engorged in 4 days and females in 5 or 6 days. Four age categories were defined for larvae and 3 for nymphs. The growth of females was variable but 4 stages of physiological development were defined based on changes in their lengths. The lengths were determined of each instar 24 h before detachment to enable estimation of the numbers maturing each day on cattle in the field.     

Hemalin, a thrombin inhibitor isolated from a midgut cDNA library from the hard tick Haemaphysalis longicornis

A full-length sequence of a thrombin inhibitor (designated as hemalin) from the midgut of parthenogenetic Haemaphysalis longicornis has been identified. Sequence analysis shows that this gene belongs to the Kunitz-type family, containing two Kunitz domains with high homology to boophilin, the thrombin inhibitor from Rhipicephalus (Boophilus) microplus. The recombinant protein expressed in insect cells delayed bovine plasma clotting time and inhibited both thrombin-induced fibrinogen clotting and platelet aggregation. A 20-kDa protein was detected from the midgut lysate with antiserum against recombinant hemalin. The gene is expressed at all stages of the tick except for the egg stage, and hemalin mRNA mainly in the midgut of the female adult tick. Real-time PCR analysis shows that this gene has a distinctly high expression level in the rapid bloodsucking period of the larvae, nymphs, and adults. Disruption of the hemalin gene by RNA interference led to a 2-day extension of the tick blood feeding period, and 27.7% of the RNA-treated ticks did not successfully complete the blood feeding. These findings indicate that the newly identified thrombin inhibitor from the midgut of H. longicornis might play an important role in tick blood feeding.     

Identification of two forms of cyclophilin from the hard tick Haemaphysalis longicornis

We have shown here the identification and characterization of two cyclophilin, cyclophilin A (CyPA) and B (CyPB), from the ixodid tick, Haemaphysalis longicornis. Both CyPA and CyPB contain the conserved peptidyl-prolyl isomerase (PPIase) domain, with CyPA consisting of 188 amino acids and CyPB of 216 amino acids. CyPA and CyPB share 50–67% amino acid sequence identity with the cyclophilins of other organisms, and are found in multiple organs throughout the developmental stages of ixodid ticks. In addition, recombinant CyPA and CyPB exhibited PPIase activity that could be inhibited by the cyclic peptides cyclosporin A (CsA). Silencing of CyPA through RNA interference has led to a significant reduction in the body weight of engorged ticks and their failure to lay eggs, in contrast to CyPB whose silencing did not result in any detectable phenotypic changes. Our results indicate that CyPA represents the major cyclophilin protein in H. longicornis involved in blood ingestion, viability, and oocyte development. This is the first report of cyclophilin proteins from ixodid and argasid ticks.     

Biosteres longicaudatus: Developmental dependence on host (Anastrepha suspensa) physiology

Larvae of Anastrepha suspensa that were in the first day of the third instar were parasitized by females of the solitary endoparasitoid, Biosteres longicaudatus. At the end of the 6-hr oviposition period, larvae were ligated posterior to the ring gland so that some larvae had parasitoids anterior to the ligature while in others, the parasitoids were in the abdomen, posterior to the ligature. Ninety-two percent of the parasitoids anterior to the ligature hatched to the first through third instars. Parasitoids posterior to the ligature had a 75% egg hatch to the first instar only. No larval molts to the second or subsequent instars occurred in these parasitoids. Upon parabiosis to 3-day-old, unparasitized host pupae, the ligated larvae pupated and 97% of the first-instar parasitoids in these parabiosed larval abdomens molted to the second instar. Newly laid parasitoid eggs transplanted to 3-day-old pupal hosts had less than one-third of the egg hatch of those transplanted to first-day third-instar hosts. The data implicate the physiological state of the host (vis-a-vis pupation and associated events) as being an important factor in the development of the endoparasitoid.     

Multiple vitellogenins from the Haemaphysalis longicornis tick are crucial for ovarian development

Ovarian development and egg maturation are crucial processes for the success of reproduction in ticks. Three full-length cDNAs encoding the precursor of major yolk protein, vitellogenin, were obtained from cDNA libraries of the Haemaphysalis longicornis tick and designated as HlVg-1, HlVg-2 and HlVg-3. The HlVg mRNAs were found in fed females with major expression sites in the midgut, fat body and ovary. Native PAGE and Western blot demonstrated that HlVgs in the hemolymph, fat body and ovary of fed females consisted of four major polypeptides. RNAi results showed that HlVg dsRNA-injected ticks obtained lower body weight, egg weight and showed higher mortality of engorged females after blood sucking than control groups. Our results indicate that all HlVgs are essential for egg development and oviposition.     

长角血蜱幼虫中的蜕皮甾类(英文)

本文用放射免疫分析和高效液相色谱法对长角血蜱幼虫中的蜕皮甾类进行了测定。　蜕皮甾类存在于吸血和饱血幼虫,并且随生理时期而变化。吸血期和饱血后前３天,激素含量低（小于 １４． ３５ｐｇ ＥＥ／只）;饱血后５天迅速上升（２９．８７ｐｇ ＥＥ／只）;饱血后７天达到高峰（５６． ０４ｐｇ ＥＥ／只）;高峰后又下降到低水平（１９．３８ ｐｇＥＥ／只）。高峰期的蜕皮甾类主要为两种成分,２０Ｅ和Ｅ（２０Ｅ∶Ｅ＝３．０７∶１）。２０Ｅ可能是幼虫发育中起主要作用的激素。     

The development of oocytes in the ovary of a parthenogenetic tick, Haemaphysalis longicornis

Haemaphysalis longicornis is an important vector of various pathogens in domestic animals and humans. The tick is a unique species with bisexual and parthenogenetic races. Although mating induces oocyte development, it is possible in the parthenogenetic race to complete oogenesis without copulation. Here we examined the developmental process of oocytes from unfed to the oviposition period in parthenogenetic H. longicornis. We classified the developmental stages of oocytes into five stages: stage I, germinal vesicle occupies more than half of the cytoplasm; stage II, germinal vesicle occupies less than half of the cytoplasm; stage III, germinal vesicle migrates from the center in the oocyte to the vicinity of the pedicel cells; stage IV, the cytoplasm is filled with yolk granules of various sizes; stage V, the cytoplasm is occupied by large yolk granules. Oocytes at the unfed period were undeveloped and classified as stage I. Stage I and II oocytes were observed at the rapid feeding period, indicating that oocyte development began after the initiation of blood feeding. All developmental stages of oocytes were observed at the pre-oviposition period. At 10?days after the beginning of the oviposition period, the ratios of stage I and II oocytes were higher than those of the previous period, suggesting that the ovarian development and activity may be continuing. Based on these findings, we propose classification criteria for the oocyte development in the parthenogenetic H. longicornis. The criteria will be useful for understanding the mechanisms of tick reproduction and transovarial transmission of pathogens.     

The effect of density on the survival of BoophilusMicroplus on previously unexposed cattle

Previously unexposed ‘naive’ cattle were infested with between 500 and 20,000 larvae of the tick Boophilus microplus, to observe whether density dependent mortality of the tick occurs on cattle which have not acquired resistance. Similar proportions of larvae matured at different densities, which contrasts with previous findings of increased mortality at higher densities on cattle with acquired resistance to B. microplus. Major differences were observed between the proportions of larvae maturing in different experiments.     

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。     

A history of the introduction,establishment, dispersal and management of Haemaphysalis longicornis Neumann, 1901 (Ixodida: Ixodidae) in New Zealand

ABSTRACT

The earliest concerns New Zealand farmers had about ticks are chronicled here, together with the eventual invasion and establishment of Haemaphysalis longicornis, which, still today, remains the only livestock tick parasite in this country. Early attempts, legislative and practical, to restrict the spread of the tick, and later attempts to manage a permanent problem are presented and discussed. Brief biographies of many of the main persons mentioned in this history are presented in an Appendix, as is a reconsideration of how the tick may have arrived.     

Evaluation of the acaricidal activity of 63 commercialized pesticides against Haemaphysalis longicornis (Acari: Ixodidae)

The acaricidal activity and residual effects of 63 commercialized pesticides against the Cheongju strain of Haemaphysalis longicornis in Korea were examined. Twenty‐two pesticides (4 carbamates, 5 organophosphates, 10 pyrethroids, 1 amitraz, 1 diamide, and 1 unclassified pesticide) caused greater than 80% mortality in H. longicornis adults under laboratory conditions using the spray method. These 22 pesticides were used to treat grassland under field conditions for investigation of the residual effects, and 100% mortality in H. longicornis adults was observed with all the carbamates (carbosulfan, benfuracarb, fenobucarb, and carbaryl) and γ‐cyhalothrin after 3 days of pesticide treatment. γ‐Cyhalothrin exhibited 56.7% mortality after 10 days of treatment, which was the longest residual effect of treated grass on H. longicornis adults. With regard to the residual effects on H. longicornis under field conditions, most of the 22 pesticides exhibited higher mortality in nymphs than in adults. In particular, benfuracarb exhibited 96.7% acaricidal activity until 10 days after treatment. These results indicate that carbamates are highly likely to be available under field conditions and, based on this preliminary data, could be used for the control of H. longicornis adults and nymphs.     

Tritrichomonas foetus: Stable anaerobic resistance to metronidazole in vitro

Stable anaerobic resistance of Tritrichomonas foetus to metronidazole was induced in vitro by cultivation of trichomonads in the Diamond's TYM medium with metronidazole in concentrations sublethal to the parasites. Nine metronidazole-resistant strains were derived from four drug-susceptible clones of the T. foetus strain KV-1. Subculturing the parasites at both increasing and constant pressure of the drug resulted in development of resistance if the medium contained at least 3 μg ml^?1 of metronidazole and the organisms were exposed to the drug for 3 to 8 months. The development of resistance was gradual and in all clones investigated proceeded through similar sequence of stages: (1) Survival without growth and subsequent reproduction at low metronidazole concentrations (1 to 5 μg ml^?1. (2) Survival and reproduction at moderate concentrations of the drug (10 to 15 μg ml^?1. (3) Resistance to 100 μg ml^?1 metronidazole, unstable in absence of selective pressure of the drug. (4) Resistance to high concentrations of metronidazole, stable when the organisms were maintained under nonselective conditions. The trichomonads with fully developed resistance were able to grow in anaerobic culture at 100 μg ml^?1 metronidazole and could be maintained indefinitely under these conditions. The minimal lethal concentrations for metronidazole obtained with these strains in an anaerobic in vitro assay were, at 48 h, 500 to 1000 μg ml^?1. This is 100 to 400 times higher than those obtained with the parent clones. The fully developed resistance was stable in organisms maintained in the absence of the drug over 2 years. The substrains with unstable resistance regained the susceptibility to high concentrations of metronidazole after 80 to 100 transfers in drug-free media. These strains, however, retained their resistance to moderate doses of metronidazole and full resistance could be restored by subculture in the presence of 10 μ ml^?1 metronidazole.     

The effect of varying the number of Schistosoma mansoni miracidia on the reproduction and survival of Biomphalaria pfeifferi

The number of young snails emerging from egg masses of Biomphalaria pfeifferi (Kampala strain) is reduced by exposure to an increasing number of miracidia of Schistosoma mansoni (West Nile strain). Snails exposed to four miracidia are rendered sterile. Survival of infected snails is inversely proportional to the number of miracidia to which an individual snail is exposed.     

长角血蜱卵黄蛋白单克隆抗体的制备及鉴定

为研究蜱类的卵黄发生及其机理,用长角血蜱Haemaphysalis longicornis卵黄蛋白（vitellin, Vn）免疫BALB/c小鼠,取免疫鼠脾细胞与骨髓瘤细胞SP2/0进行融合,经3次克隆化筛选,获得6株能稳定分泌抗Vn的单克隆抗体（McAb）,即1B5,2A7,2B8,2F2,3A1和3G1。1B5,2B8和2F2亚型为IgGA;2A7亚型为IgG1;3A1和3G1亚型为IgG2a。6株McAb均具有高度特异性,效价在1∶10.5以上。选取效价和特异性最好的1株抗体1B5进行SDS-PAGE分析和亲和力测定,测得1B5重链和轻链的分子量分别为58 kD和21 kD,亲和常数为2.8993×10^-6。Western免疫印迹分析发现6株单抗均与Vn的8个亚基发生免疫反应。本研究成功制备了6株抗长角血蜱Vn的单克隆抗体,为深入阐明长角血蜱卵黄发生的过程与调控机理提供了重要的工具。     

Acquired resistance in snails. Induction of resistance to Schistosoma mansoni in Biomphalaria glabrata

Acquired resistance to Schistosoma mansoni PR-I strain has been induced in Biomphalaria glabrata 442132 strain by infecting the snails with irradiated homologous miracidia. Present and previous results support the hypothesis that acquired resistance to trematodes in snails is an enhancement of the host's natural resistance to the parasite.     

长角血蜱不同发育期盾窝超微结构的比较研究

为阐明蜱类盾窝及其发育特点,用扫描电镜观察了长角血蜱Haemaphysalis longicornis不同发育期盾窝的结构,并分析了血餐对盾窝发育的影响.结果表明：幼蜱仅具1对盾窝原基,且每个盾窝原基有1个盾窝孔;若蜱盾窝有了一定的发育,面积（长径×短径）增大且盾窝孔数增多（2~6个）;成蜱盾窝面积最大,且盾窝孔数达21~35个.盾窝的发育主要在幼蜱蜕皮阶段及若蜱的吸血和蜕皮阶段,雌蜱盾窝孔径显著大于雄蜱（P<0.01）,成蜱、若蜱和幼蜱的盾窝孔孔径在吸血过程中（交配雌蜱除外）各虫期均无显著变化 （P>0.05）.综合分析成蜱与未成熟蜱盾窝孔径,发现它们之间无显著差异 （P>0.05）,这在一定程度上说明蜱类的盾窝孔径在未成熟期可能已经有了雌雄分化.     

性信息素2，6-二氯酚在长角血蜱交配行为中的作用

长角血蜱Haemaphysalis longicornis的交配行为包括7个时期,行为的完成依赖于性信息素的调节。生物测定表明：雄蜱的行为反应受雌蜱分泌的性信息素影响。堵塞雌蜱盾窝其行为受到抑制,点滴2,6-DCP或雌蜱盾窝腺提取物则被恢复。用气相色谱法测定了雌蜱盾窝腺中2,6-DCP的含量;吸血后1～２天含量最高(11.12 ng/只);吸血后3～５天即交配前下降交维持在一较恒定的水平;吸血后6～７天即交配后明显降低;饱血后检测不到2,6-DCP。2,6-DCP是长角血蜱性信息素的一种成分。     

长角血蜱卵黄蛋白的纯化及其性质

用凝胶过滤与离子交换层析、蛋白质电泳和糖脂蛋白染色等方法提取纯化长角血蜱Haemaphysalis longicornis卵黄蛋白,并对其性质进行了研究。PAGE和SDS-PAGE分析表明,长角血蜱的卵黄蛋白只有一种,由8个亚基组成,亚基的相对分子质量分别为112 kD, 103 kD, 80 kD, 78 kD, 71 kD, 68 kD, 62 kD和52 kD,卵黄蛋白经苏丹黑B和希夫试剂染色呈阳性,表明是一种含血红素的糖脂蛋白。     

长角血蜱雌蜱感染嗜菌异小杆线虫后血淋巴的变化

用嗜菌异小杆线虫Heterorhabditis bacteriophora E-6-7(Hb E-6-7)感染长角血蜱Haemaphysalis longicornis雌蜱,测定感染后雌蜱血淋巴总蛋白含量、酯酶活性及酯酶同工酶的变化,以探讨昆虫病原线虫对蜱的致病机理。结果显示,雌蜱血淋巴总蛋白含量在Hb E-6-7感染后12 h显著增加,达到最大值92.21 μg/μL;在感染后24 h明显下降（49.06 μg/μL）;至感染后48 h降至34.25 μg/μL。雌蜱血淋巴酯酶活性在线虫感染后0～12 h内无显著变化;在感染24 h后迅速增加（OD_24h=0.1840,OD_36h=0.1940,OD_48h=0.2165）,与对照组差异显著。PAGE结果表明,线虫感染后导致雌蜱血淋巴酯酶同工酶电泳图谱发生变化,主要为电泳图谱两端a带和b带的消失以及一条c带的增加。上述结果表明,长角血蜱雌蜱被Hb E-6-7感染后其血淋巴总蛋白含量和酯酶发生变化,这种变化可能与蜱的防御和对昆虫病原线虫的适应有关。