Import and efflux of flubendazole in Haemonchus contortus strains susceptible and resistant to anthelmintics

Drug entry into the body of a helminth is a key factor in the efficacy of anthelmintics. The present project was designed to study the ex vivo uptake and efflux of the benzimidazole anthelmintic flubendazole (FLU) in four strains of H. contortus: the ISE strain (fully susceptible to anthelmintics), the ISE-S strain (resistant to ivermectin), the BR strain (resistant to benzimidazoles) and the WR strain (multi-resistant). The transport of FLU between dead and living nematodes was also compared as well as the effect of verapamil, an inhibitor of the main efflux ABCB1 transporter (P-glycoprotein), on FLU accumulation in nematodes. The obtained results showed that FLU is able to effectively enter H. contortus adults due to high FLU lipophilicity. Passive diffusion is probably the only mechanism in both FLU import and efflux from nematodes. No differences in FLU transport were found among four H. contortus strains with different sensitivity to anthelmintics. No active FLU efflux from H. contortus and no effect of verapamil were observed, indicating that H. contortus cannot protect itself against FLU by the active removal of this anthelmintic from its body.     

Decreased ivermectin and moxidectin sensitivity in Haemonchus contortus selected with moxidectin over 14 generations.

Ivermectin resistance in the nematode Haemonchus contortus has been reported in many parts of the world and many ivermectin resistant isolates have been found to have reduced sensitivity to moxidectin. However, it is unclear whether parasites that are selected with moxidectin would demonstrate reduced sensitivity to ivermectin. In this study, the effects of moxidectin and ivermectin on an unselected strain and a strain of H. contortus derived from the unselected strain but selected over 14 generations with moxidectin, were compared in jirds. The recovery of adult worms and fourth stage (L4) larvae following treatment were compared between strains and anthelmintics. Moxidectin-selected H. contortus showed reduced sensitivity to ivermectin as well as to moxidectin. Doses of 0.1 mg/kg of moxidectin and 0.4 mg/kg of ivermectin were necessary to obtain an efficacy of 95% or above against the moxidectin-selected strain of H. contortus compared with 0.025 mg/kg for moxidectin and 0.1 mg/kg for ivermectin required for a similar efficacy in the unselected strain.     

Rumen function and digestion of Merino sheep divergently selected for genetic difference in resistance to Haemonchus contortus

The aim of this experiment was to investigate the effect of Haemonchus contortus infection on rumen function and digestion of Merino sheep from lines divergently selected for genetic resistance to H. contortus. Rumen function and whole-tract digestibility were determined in 29 Merino weaner wethers from the CSIRO Haemonchus selection flock, increased resistance to Haemonchus (IRH), decreased resistance to Haemonchus (DRH) and random bred control (C) selection lines. Wethers were fed a restricted low quality roughage diet (5.9 MJ ME/kg DM, 29 g MP/kg DM) to maintain weight and were either uninfected (NIL) or infected with H. contortus (INF). Measurements were taken at week 7 of infection, while animals were housed in metabolic crates. Animals were then euthanased to determine abomasal worm counts. IRH had significantly lower worm egg count than DRH and C lines however, adult worm and larval counts, though lower in IRH animals, did not differ significantly. DM and OM digestibility and in sacco degradability of IRH and DRH selection lines were greater than C animals. Rumen function of animals from the IRH line was altered in response to H. contortus infection with an increase in fluid outflow and turnover rate and a decrease in propionic acid concentration. These changes may be a component of a greater host resistance and begin to explain the failure of IRH animals to translate lower worm egg count into greater bodyweight gain.     

The establishment and development of Haemonchus contortus in goats

Twelve goats were inoculated with 40,000 third-stage Haemonchus contortus larvae and two were killed on each of Days 4, 7, 11, 14, 18 and 21 after inoculation (DAI). The number of worms that established, and the site of development were recorded. More worms established in the fundic, than in the middle or pyloric thirds of the abomasum. Early development occurred within the mucosa; emergence into the lumen started between 7 and 11 days after infection. By 4 DAI, all worms had completed the third moult to the L4 stage. At 11 DAI the majority of the worms were adults. A mean of 13.2% of the female worms had eggs in their uteri at 18 DAI; by 21 DAI more than half of the female worms had eggs in their uteri. The development of H. contortus was essentially similar to that described in sheep.     

Hypergastrinaemia of sheep infected with Haemonchus contortus

Radioimmunoassay of plasma gastrin has shown that a hypergastrinaemia occurs in yearling sheep, after infection with 10(6) infective third stage larvae of Haemonchus contortus. Plasma gastrin concentrations began to rise two to four days after infection, that is, at about the same time as, or slightly before, the abomasal pH was observed to increase.     

捻转血矛线虫ZJ株滞育虫体的获得及其形态特征

为进一步研究寄生性线虫的滞育机制,参照Adams(1986)的方法,通过瘤胃灌注,将捻转血矛线虫感染性三期幼虫攻入绵羊体内,对绵羊进行处理后,获得捻转血矛线虫滞育虫体。滞育虫体发育程度一致,虫体长(1 067±97)μm(n=18),肠道细胞中含有大量棒状结晶块;性腺开始发育,出现雌驱器原基;颈部出现颈乳突。通过温度检测分析,温度可能不是滞育现象的主要刺激因素。棒状结晶块可以被DAPI特异性着色,其成分可能含有核酸。本试验将为进一步研究捻转血矛线虫的滞育机制及其防治奠定基础。     

Factors affecting the prevalence and severity of footrot in a Merino flock selected for resistance to Haemonchus contortus

SUMMARY An outbreak of virulent footrot occurred in three lines of sheep selected for different levels of resistance to Haemonchus contortus , providing an opportunity to examine the factors affecting footrot severity. The selection lines did not differ significantly in average footrot score, but various other factors were found to have an effect. In breeding ewes, the average score was 10.2 (maximum possible score 20) and the prevalence of footrot was 96%. There was a significant age effect, with average score increasing from 9.1 ± 0.8 in 2-year-old ewes, to 13.4 ± 0.9 in 7-year-old ewes. Ewes that lost their lambs tended to have higher scores than those that either failed to lamb or successfully reared their litter. Significant differences in footrot score were caused by the lambing paddock in which the ewes had been, during the period seven weeks to one week before infection. Among lambs, aged about 20 weeks, the average footrot score was lower (5.0), as was the prevalence (75%). The score increased by an average of 0.09 ± 0.03 per day of age and the prevalence increased by an average of 0.6%± 0.3% per day of age. Lambs born and reared as twins had significantly lower scores than singles, and progeny of maiden dams had lower scores than progeny of older ewes. There was no significant difference because of the sex of the lamb. The paternal half-sib estimate of heritability of footrot score was 0.20 ± 0.14 and of the genetic correlation between footrot score and worm egg count was essentially zero (0.02 ± 0.20), but these estimates were probably biased by the confounding of sires with lambing paddocks. Nevertheless, the results strongly suggest that there are no adverse genetic associations between resistance to footrot and resistance to internal parasites.     

Rapid selection for ivermectin resistance in Haemonchus contortus

Lambs infected with two isolates, one British and one American, of Haemonchus contortus were treated with increasing doses of ivermectin. Eggs from the highest dose that had not eliminated the infection were cultured and larvae used to infect another lamb. After three generations the H. contortus was resistant to 0.2 mg/kg ivermectin. The results stress the ease with which ivermectin resistance can be selected if high selection pressure is applied.     

捻转血矛线虫Hc-TTR-51蛋白的定位及功能

捻转血矛线虫(Haemonchus contortus)是最重要的家畜寄生虫之一。已报道秀丽隐杆线虫(Caenorhabditis elegans)转甲状腺素蛋白(transthyretin,TTR)参与虫体细胞间的相互作用。目前捻转血矛线虫转甲状腺素蛋白(Hc-TTR-51)的定位及功能尚未见报道。研究首先克隆捻转血矛线虫Hc-ttr-51基因,然后分析其蛋白在捻转血矛线虫、HEK 293T细胞内的定位情况,最后RNA干扰Hc-ttr-51基因分析对秀丽隐杆线虫生长发育的影响,从而对其功能进行预测分析。结果表明,捻转血矛线虫Hc-ttr-51基因在虫体各发育阶段均有表达,与其他发育期虫体相比,L1、L3、雄成虫该基因的转录水平显著上升。在L3期虫体体内该蛋白广泛表达,L4期和成虫体内该蛋白特异性表达于性腺。在HEK 293T细胞中,该蛋白在细胞质中呈明显的点状分布。RNA干扰秀丽隐杆线虫Hc-ttr-51基因表达后,虫体产卵量极显著下降,体长体宽显著缩短,咽泵率显著下降。以上研究为捻转血矛线虫生长发育特性、疫苗研究等提供依据。     

Serial passage of Haemonchus contortus in resistant and susceptible sheep

A recently isolated field strain of Haemonchus contortus was passaged through resistant (repeatedly reinfected) and susceptible (immunosuppressed) sheep for six and nine generations, respectively. Infectivity of the resulting serially passaged strains was not significantly different when tested in groups of 1-year-old susceptible sheep.     

欧洲盘羊捻转血矛线虫感染的诊断及病理学观察

对1只急性死亡的幼年欧洲盘羊进行剖检观察,皱胃内可见大量粉白色线虫,采用形态学及分子生物学相结合的方法对虫体进行鉴定,诊断为捻转血矛线虫;对虫体寄生部位的病理学进行观察,在皱胃黏膜间及黏膜固有层内均可见虫体及虫卵。     

Voluntary feed intake and diet selection of Merino sheep divergently selected for genetic difference in resistance to Haemonchus contortus

This research was designed to determine if divergent selection for resistance to Haemonchus contortus had produced correlated changes in voluntary feed intake and diet selection. Voluntary feed intake, diet selection and production were determined in 54 Merino weaner rams from the CSIRO Haemonchus selection flock, increased resistance to Haemonchus (IRH), decreased resistance to Haemonchus (DRH) and random bred control (C) selection lines. Weaner rams were fed ad libitum either a high (9.2 MJ ME/kg DM, 90 g MP/kg DM) or moderate (6.3 MJ ME/kg DM, 30 g MP/kg DM) quality diet and given the choice between the two diets, when uninfected (NIL) or infected with H. contortus (INF). Symmetrical response to divergent selection for worm egg count (WEC) was not matched by a symmetrical change in feed intake and there was no difference in diet selection between selection lines. Feed intake, growth and wool production of DRH animals remained the same as that of IRH, yet DRH animals had five times greater WEC than IRH. This study begins to explain the mechanisms that allow resistant animals to effectively prevent establishment and/or development of H. contortus, by maintaining a greater immune response to infection through higher circulating eosinophils, plasma globulin and IgG(1) antibody titres. Susceptible animals have displayed resilience by improving feed conversion efficiency and increasing protein synthesis.