Genetics of vulvar morph types in Haemonchus contortus: Haemonchus contortus from the Northern Tablelands of New South Wales

Le Jambre L. F. and Royal W. M. 1977. Genetics of vulvar morph types in Haemonchus contortus: Haemonchus contortus from the Northern Tablelands of New South Wales. International Journal for Parasitology7: 481–487. The inheritance of the vulvar morph types, smooth, knobbed, linguiform A-right, linguiform A-left and linguiform C-type was studied in Haemonchus contortus from the Northern Tablelands of New South Wales. The dominance hierarchy of these characters was found to be smooth > knobbed > linguiform. The linguiform phenotype was the most common in the wild type population. The modifications on the linguiform phenotype were found to be highly heritable with A-right responding most to selection. Realised heritabilities for these characters was A-right 0.37, A-left 0.15, and C-type 0.21. It appeared that the dominance of a phenotype increased following selection.     

Optimum temperature for egg development of phenotypes in Haemonchus contortus cayugensis as determined by Arrhenius diagrams and Sacher's entropy function

Lejambre L. P. and Whitlock J. H. 1973. Optimum temperature for egg development of phenotypes in Haemonchus contortus cayugensis as determined by Arrhenius diagrams and Sacher's entropy function. International Journal for Parasitology3: 299–310. Oxygen uptake, maturation time and percentage hatch were determined on eggs from “wild-type” Haemonchus contortus cayugensis populations as well as those from the morph types smooth, linguiform B and linguiform A. Sacher's organizational entropy, which is essentially a sum of oxygen utilized by a system corrected for the number of viable units in the system, was used to determine the optimum temperature for development of the eggs. Although the eggs from “wild-type” H. contortus cayugensis hatched well across a broad temperature range; individual phenotypes showed a much narrower range. The optimum temperature for the development of eggs from smooth worms was approximately 30°C while linguiform A and B hatched best at 25°C. Linguiform A eggs had a broader temperature tolerance than linguiform B so that at temperatures of 17°C they hatched with a greater efficiency than did either smooth or linguiform B. It is argued that H. contortus follows a “strategy” of having many phenotypes, each one of which appears to be adapted to a different temperature. This would allow a population to maintain a broad range of optimal temperatures without the expense of maintaining the cybernetic machinery which a single individual would have to have if it were to tolerate the same range of temperature.     

Haemonchus contortus: Immunodiffusion patterns of antigens from phenotypically different females

Immunodiffusion patterns of the different morphological types of female Haemonchus contortus (Rudolphi, 1803) indicate that the three phenotypes—smooth, linguiform, and knobbed—differ from each other serologically. Linguiform antigens gave five precipitin lines, knobbed gave four lines, and smooth gave three precipitin lines with rabbit antisera. Also, lines unite in a manner indicating marked antigenic differences between the three types. Since serological studies are considered to be sensitive tools in taxonomy, the present work indicates that taxonomic importance should be attached to the vulvar configurations in female H. contortus.     

Haemonchus contortus: New data on its genetic constitution

Studies on the genetic constitution of single smooth-bodied females, appearing from time to time in the homozygous (aa) linguiform strain “C,” revealed that these worms have the constitution of the linguiform (aa) morph but the character “flap” in them has failed to develop because of the undetermined effect of some additional factors. Experiments were also performed to check the dominance to recessiveness ratios of the characters “flap,” “knob,” and “smooth.” With the Haemonchus types present in Bulgaria these ratios have persisted over seven or eight years. It is stated that they are the same in populations whose morphologic types in female specimens show different percentage distributions.     

Hybridization of Australian Haemonchus placei (place, 1893), Haemonchus contortus cayugensis (Das & Whitlock, 1960) and Haemonchus contortus (Rudolphi, 1803) from Louisiana

There were distinct ecotypie differences in the ability to develop to third stage larvae at a constant 11 or 13°C for two weeks. H. conforms cayugensis could develop at both 11 and 13°C; H. contortus from Louisiana could develop at 13°C but not 11°C and H. placei could not complete development at either temperature. Eggs produced from the first cross between ecotypes had the same cold tolerance as their maternal parent. F₁ eggs were intermediate between parental ecotypes, while F₂ eggs, when fertile, developed at the same temperature as the most cold-tolerant parent in the first cross. The H. placei knobbed vulvar morph type was dominant to H. contortus cayugensis smooth morph type and recessive to the H. contortus (Louisiana) smooth morph type. Knobbed morph type of H. contortus (Louisiana) was dominant to smooth of H. contortus cayugensis. Dominance of morph type was enhanced by a matroclinous effect in the between species matings but not in the within species matings. No fertile eggs were produced by the F1 of the mating between female H. placei × male H. contortus (Louisiana) or by the F₂ of the reciprocal mating. Reciprocal matings between H. placei × H. contortus cayugensis produced an F₁ and F₂ that had reduced fertility but were not completely sterile. No decrease in fertility was observed in the F₂ and F₁ from the H. contortus cayugensis × H. contortus(Louisiana)matings. Cytological studies revealed several kinds of meiotic disturbances in the between species F₁ and F₂. The most frequent were aneuploidy, failure of chromosome pairing, and pairing between non-homologous chromosomes. Males were more seriously affected than females by meiotic disturbances. The H. placei X chromosome appeared to be selected against by the hybrid genetic background. Within species hybrids showed no meiotic disturbances and the percentage of male offspring from these crosses was the same as for female offspring.     

Description of Haemonchus placei (Place, 1893) (Nematoda, Trichostrongylidae, Haemonchinae), identification and intra-specific morphologic variability

Haemonchus placei in cattle has never been completely described, possibly due to great morphological similarity with small ruminants Haemonchus contortus. It is newly described on one isolate from Argentina. It has clear distinct morphological features from sheep and goats Haemonchus contortus and presents only two female morphotypes (linguiform and knobbed) instead of three recorded in H. contortus. A key is proposed to identify females. Female as well as male Haemonchus placei from New World (Argentina, Mexico, USA) are morphologically different from those of Old World (Africa: Burkina-Faso, Mauritania and Ivory Coast) or Australia, possibly due to local evolution since their introduction several centuries ago from Africa or India. We propose to differentiate three sub-species, H. placei placei in Australia, H. placei africanus in western Africa and H. placei argentinensis in the New World.     

Norcantharidin analogues with nematocidal activity in Haemonchus contortus

With the major problems with resistance in parasitic nematodes of livestock to anthelmintic drugs, there is an urgent need to develop new nematocides. In the present study, we employed a targeted approach for the design of a series of norcantharidin analogues (n = 54) for activity testing against the barber’s pole worm (Haemonchus contortus) of small ruminants in a larval development assay (LDA) and also for toxicity testing on nine distinct human cell lines. Although none of the 54 analogues synthesized were toxic to any of these cell lines, three of them (N-octyl-7-oxabicyclo(2.2.1)heptane-2,3-dicarboximide (B2), N-decyl-7-oxabicyclo(2.2.1)heptane-2,3-dicarboximide (B3) and 4-[(4-methyl)-3-ethyl-2-methyl-5-phenylfuran-10-oxa-4-azatricyclo[5.2.1]decane-3,5-dione (B21) reproducibly displayed 99-100% lethality to H. contortus in LDA, with LD_50s of 25-40 μM. The high ‘hit rate’ (5.6%) indicates that the approach taken here has advantages over conventional drug screening methods. A major advantage of norcantharidin analogues over some other currently available anthelmintics is that they can be produced in one to two steps in large amounts at low cost and high purity, and do not require any additional steps for the isolation of the active isomer. This positions them well for commercial development.     

Inheritance of avermectin resistance in Haemonchus contortus

A larval development assay was used to compare the responses of the Chiswick Avermectin Resistant (CAVRS) isolate of Haemonchus contortus, an avermectin-susceptible isolate (VRSG) and their crosses to avermectins. The F(1) and F(2) generations of reciprocal crosses between CAVRS and VRSG were denoted as CAVRS malesxVRSG females=CXV, and VRSG malesxCAVRS females=VXC. The levels of avermectin resistance in the developing larvae of the F(1) of both CXV and VXC were indistinguishable from that in the avermectin-resistant parent, indicating that the resistance trait is completely dominant. Avermectin dose-response curves for the CXV F(1) did not show a 50% mortality rate at low concentrations, indicating that avermectin resistance is not sex-linked. This conclusion was confirmed when adult male worms of the F(1) of the CXV mating were found to have survived treatment of the host with 200microgkg(-1) ivermectin. This dose rate (200microgkg(-1) ivermectin) caused a 50% reduction in the number of adult males in the F(1) from both CXV and VXC crosses, but only a non-significant reduction in the number of adult females in the F(1). Dose-response curves obtained for the F(2) generations in the larval development assay indicated the presence of 25% of avermectin-susceptible individuals, suggesting that a single major gene largely controls the avermectin-resistance trait. This genetic analysis of avermectin resistance in an Australian H. contortus isolate indicates that the expression of the gene for avermectin resistance is an autosomal, complete dominant in the larvae; however, in adults its expression is sex-influenced, with males having a lower resistance to avermectin than females.     

Feed resource selection of Criollo goats artificially infected with Haemonchus contortus: nutritional wisdom and prophylactic self-medication

Previous cafeteria studies suggested that a moderate natural gastrointestinal nematode (GIN) infection did not modify the resource selection of adult Criollo goats towards tannin-rich plants compared with worm-free goats. A higher infection with Haemonchus contortus could trigger a change in the resource selection behaviour towards tannin-rich foliage. Alternatively, goats might select plant species solely to meet their nutritional requirements. A cafeteria study investigated the effect of a high artificial infection with H. contortus on the feed resource selection of goats. Adult Criollo goats (37.5±4.8 kg BW) with browsing experience were distributed in two groups: the infected group (IG) with six animals artificially infected with H. contortus (6000 L₃/animal); and the non-infected group (NIG) with six animals maintained worm-free. The experiment included two 5-day periods with additional 5-day adaptation period. In the first period, animals were offered foliage of five plant species with a decreasing gradient of condensed tannins (CT) (Mimosa bahamensis, Gymnopodium floribundum, Havardia albicans, Acacia pennatula, Lysiloma latisiliqum), and three plant species with negligible CT content (Leucaena leucocephala, Piscidia piscipula and Brosimum alicastrum). In the second period the foliage of B. alicastrum was withdrawn. A grain-based concentrate feed was offered daily at 1% BW in DM basis. Dry matter and nutrient intake was determined. Foliage selection of each experimental group was determined using the Chesson selection index. The H. contortus egg count per gram of faeces (EPG) was determined for infected goats twice daily. Chesson index showed a similar pattern of foliage selection on periods 1 and 2. Mean EPG of goats in IG was 2028±259 EPG during period 1 and 1 293±198 EPG during period 2 (P>0.05). During period 1, the selection pattern was highest for B. alicastrum (tannin-free), followed by a tannin-rich plant (M. bahamensis). These two plants remained as highly selected during period 2. The Chesson index showed that both experimental groups (IG and NIG) selected the same plant species in both periods. Thus, a high H. contortus infection did not affect selection of goats fed with CT-rich plants. Apparently, goats balanced their nutrient intake with the plants selected, showing evidence of nutritional wisdom. This balance may have helped to prevent excess protein in the diet and also to maintain a low GIN infection, both considered as examples of prophylactic self-medication.     

The mechanism of hatching of eggs of Haemonchus contortus

Fluid collected from hatching eggs of Haemonchus contortus contained a lipase which hydrolysed 2-naphthyl laurate (about 0·7 μmol naphthol freed /h/10⁶ eggs). The fluid also hydrolysed l-leucinamide (about 2·3 μmol leucine freed/h/10⁶ eggs). The fluid when added to normal or heated eggs caused ‘hatching’. ‘Hatching’ also occurred in exsheathing fluid from infective juveniles and in a preparation of pancreatic lipase containing leucine aminopeptidase. A purified mammalian leucine aminopeptidase in combination with several different lipases did not attack egg shells.The ‘spontaneous’ hatching of eggs of H. contortus was strongly inhibited by 1,10-phenanthroline, 10^?3M, and this inhibition was reversed by Zn²⁺. However, the inhibition of ‘hatching’ of eggs in externally applied hatching fluid, or the hydrolysis of leucinamide in hatching fluid was generally less marked.     

Effects of temperature on the expression of elytral colour polymorphism in the ladybird beetle,Menochilus sexmaculatus (Coleoptera: Coccinellidae)

The ladybird beetle, Menochilus sexmaculatus (Fabricius), has a remarkable elytral colour polymorphism composed of black and red. In the present study, we investigated the effect of temperature on growth from the first instar larva to the pupal stage, as well as maternal morph types on the phenotypic expression of the elytral colour morph in a polymorphic population from Osaka, Japan. Female individuals of three different elytral colour morphs were collected from a wild population, and hatchlings from each female were divided into three groups, which were reared at three constant temperatures: 20 °C, 25 °C, and 30 °C. The phenotypic frequency of F₁ adults indicated that the elytral morph type was determined by genetic factors, but not by growth temperatures. Namely, type A (almost black morph) was the most abundant in F₁ from type A mothers (Male: 52.6%; Female: 32.3%); and types B (four small-dotted morph) and F (four medium-dotted morph) were the most abundant from type B (Male: 56.7%; Female: 53.3%) and type G (four larger-dotted morph) mothers (Male: 33.3%; Female: 31.3%), respectively. Therefore, the expression of elytral colour polymorphism in the Osaka, Japan population is likely to have a genetic basis contingent on parental morphs, rather than a phenotypic plasticity associated with growth temperatures.     

Somatic proteome of Haemonchus contortus

Currently, there is a dearth of proteomic data to underpin fundamental investigations of parasites and parasitism at the molecular level. Here, using a high throughput LC–MS/MS-based approach, we undertook the first reported comprehensive, large-scale proteomic investigation of the barber's pole worm (Haemonchus contortus) – one of the most important parasitic nematodes of livestock animals worldwide. In total, 2487 unique H. contortus proteins representing different developmental stages/sexes (i.e. eggs, L3s and L4s, female (Af) and male (Am) adults) were identified and quantified with high confidence. Bioinformatic analyses of this proteome revealed substantial alterations in protein profiles during the life cycle, particularly in the transition from the free-living to the parasitic phase, and key groups of proteins involved specifically in feeding, digestion, metabolism, development, parasite-host interactions (including immunomodulation), structural remodelling of the body wall and adaptive processes during parasitism. This proteomic data set will facilitate future molecular, biochemical and physiological investigations of H. contortus and related nematodes, and the discovery of novel intervention targets against haemonchosis.     

The inheritance of thiabendazole resistance in Haemonchus contortus.

Haemonchus contortus worm populations isolated from naturally infected sheep at the Pastoral Research Laboratory, Armidale, N.S.W., were found to contain approximately 20% of worms resistant to a 50 mg/kg dose of thiabendazole. Following 3 generations of selection with 50 mg/kg thiabendazole the number of worms removed by the anthelmintic was too small to detect differences between treated and control groups. After more than 15 generations of selection, matings between males from the selected strain and non-resistant females produced resistant males and females in equal numbers. Thus, thiabendazole resistance does not appear to be sex-linked. A dose--response assay on the F2 adults indicated that worms from female resistant x male non-resistant crosses were more resistant than F2 adults of the reciprocal cross. An in vitro technique that identified thiabendazole-resistant eggs by their ability to hatch in a solution containing thiabendazole and 0.1% NaCl solution was also used to study the inheritance of resistance. F1 eggs had similar LC50's to the resistant parents. F2 and back-cross eggs from an original mating of thiabendazole-resistant females x non-resistant males had a higher LC50 than F2 and back-cross eggs from the reciprocal mating, indicating a degree of matroclinous inheritance of resistance. However, the resistant parents had tolerances to thiabendazole exceeding those of F2. F3 eggs had a resistance distribution that ranged from that of the resistant to the non-resistant parent. No significant deviation from linearity was observed in any of the dose--response lines. These results indicate that thiabendazole resistance in H. contortus worms is inherited as an autosomal and semi-dominant trait.     

Ecoepidemiology of Haemonchus contortus bahiensis, ecotype present in sheep of arid zones of Venezuela

The frequency distribution of female Haemonchus contortus bahiensis Grisi, 1974 in sheep from Venezuelan arid zones is 15.32% for the type with vulvar flap, 51.6% for the vulvar-knob and 33.07% for smooth type. A Shannon-Weaver diversity index corresponding to 1.44 bits was calculated for these forms with similarities in the general size, egg-size and in the number of the longitudinal cuticular ridges. An aggregated kind of distribution in the host population according to the K parameter of a negative binomial distribution was recorded for male and female worms. A complicated interaction was observed between the abundance, aggregation and prevalence of this ecotype and the importance of these findings is discussed with regard to host-parasite relationships.     

Developmental arrest of Haemonchus contortus in sheep treated with a corticosteroid

Adams D. B. 1986. Developmental arrest of Haemonchus contortus in sheep treated with a corticosteroid. International Journal for Parasitology16: 659–664. Developmental arrest of the nematode, Haemonchus contortus, at the fourth larval stage within sheep appears to be controlled by a complex of signals and events in which both host-associated and parasite-associated factors are essential participants. Treatment of worm-free sheep with the corticosteroid, dexamethasone, during infection demonstrated the existence of an arrest-prone state in infective larvae and a host-associated influence actuating the phenomenon of arrest. Not all subpopulations of the parasite responded to the influence of dexamethasone and a selection experiment in which the parasite was passaged through immune and non-immune sheep showed that the arrest-prone state was largely determined by genetic factors. The host-associated factor initiating or actuating arrest and identified by dexamethasone-treatment could be ascribed more satisfactorily to a manifestation of immune responsiveness than to a direct effect of the corticosteroid.     

A unique thioredoxin of the parasitic nematode Haemonchus contortus with glutaredoxin activity

The dependency of parasites on the cellular redox systems has led to their investigation as novel drug targets. Defence against oxidative damage is through the thioredoxin and glutathione systems. The classic thioredoxin is identified by the active site Cys-Gly-Pro-Cys (CGPC). Here we describe the identification of a unique thioredoxin in the parasitic nematode, Haemonchus contortus. This thioredoxin-related protein, termed HcTrx5, has an arginine in its active site (Cys-Arg-Ser-Cys; CRSC) that is not found in any other organism. Recombinant HcTrx5 was able to reduce the disulfide bond in insulin, and be regenerated by mammalian thioredoxin reductase with a K_m 2.19 ± 1.5 μM, similar to the classic thioredoxins. However, it was also able to reduce insulin when glutathione and glutathione reductase replaced the thioredoxin reductase. When coupled with H. contortus peroxiredoxin, HcTrx5 was active using either the thioredoxin reductase or the glutathione and glutathione reductase. HcTrx5 is expressed through the life cycle, with highest expression in the adult stage. The unique activity of this thioredoxin makes it a potential drug target for the control of this parasite.     

The inheritance of intrasexual dimorphism in female diving beetles (Coleoptera: Dytiscidae)

Many species of Dytiscus diving beetles exhibit intrasexual dimorphism, e.g., the elytra is smooth in some females and grooved in others. However, the expression of the grooves and whether they are a product of heredity or the environment remain unknown. One Japanese species, Dytiscus sharpi sharpi Wehncke, 1875 , also shows female dimorphism, with grooved and smooth morphs, while D. sharpi validus Régimbart, 1899, only has a single morph (the grooved type). A hybrid of the two species should therefore provide a means of sorting out how the grooves are inherited. We found two independent wetlands of D. sharpi sharpi in Chiba Prefecture, Japan. One was a place where a high proportion of grooved females lived, and the others had high proportions of smooth females. After five to eight generations of beetles from two populations with different proportions of grooved females were reared under aquarium conditions constituting a common garden design, i.e., water temperature, water depth, and presence of a plant for oviposition, the differences remained. We mated smooth virgin females of D. sharpi sharpi with males of D. sharpi validus to obtain hybrid offspring. The elytral traits of the hybrid females produced only grooved forms. These results suggested that the female dimorphism is determined by genetics, and that the grooved morph was dominant over the smooth one, independent of environmental factors. In addition, the hybrid insects did not differ from the two subspecies insects in larval survivorship, pupation success, or sex ratio. They also showed neither morphological abnormality nor reduced survival.     

Haemonchus contortus: enzymes. 3. Glutamate dehydrogenase

Adult male and female Haemonchus contortus were homogenized and subjected to differential centrifugation. The crude, high-speed, supernatant fraction contained more than 95% of the glutamate dehydrogenase activity. The enzyme was purified through use of DEAE-cellulose columns and sucrose density gradient centrifugation. The enzyme from both crude and purified preparations was detected as a single band of activity following starch or polyacrylamide-gel electrophoresis. The Haemonchus enzyme was compared with ovine and bovine liver glutamate dehydrogenases. The three enzymes were similar in molecular size, Michaelis constants, and pH optimums but differed in electrophoretic mobility in polyacrylamide-gels, activity with NADP as coenzyme, and effect of AMP and ADP on activity. Sheep anti-Haemonchus glutamate dehydrogenase serum inhibited Haemonchus glutamate dehydrogenase, but did not inhibit the ovine or bovine enzymes.