The aim of the study was to compare the relative gene expression of Haemonchus contortus P-glycoprotein genes (Hco-pgp) between fourth (L4), infective (L3), and transitory infective (xL3) larval stages as laboratory models to study ivermectin (IVM) resistance. The H. contortus resistant to IVM (IVMr) and susceptible to IVM (IVMs) strains were used to develop xL3 in vitro culture and to infect Meriones unguiculatus (gerbils) to collect L4 stages. Morphometric differences were evaluated from 25 individuals of H. contortus from each strain. Relative gene expression from xL3 and L4 was determined between comparison of IVMr stages and from IVMr vs IVMs stages. Seven Hco-pgp genes (1, 2, 3, 4, 9, 10, and 16) were analysed by RT-qPCR using L3 stage as control group, per strain, and GAPDH and β-tubulin as constitutive genes. Morphological changes were confirmed between xL3 and L4 developing oral shape, oesophagus, and intestinal tube. In addition, the body length and width showed statistical differences (p < 0.05). The Hco-pgp1, 2, 3, and 4 genes (p < 0.05) were upregulated from 7.1- to 463.82-fold changes between IVMr stages, and Hco-pgp9 (13.12-fold) and Hco-pgp10 (13.56-fold) genes showed differences between L4 and xL3, respectively. The comparative study between IVMr vs IVMs strains associated to xL3 and L4 displayed significant upregulation for most of the Hco-pgp genes among 4.89–188.71 fold-change. In conclusion, these results suggest the use of H. contortus xL3 and L4 as suitable laboratory models to study IVMr associated with Hco-pgp genes to contribute to the understanding of anthelmintic resistance.

As genomic research continues to improve our understanding of the genetics of anthelmintic drug resistance, the revolution in DNA sequencing technologies will provide increasing opportunities for large-scale surveillance for the emergence of drug resistance. In most countries, parasite control in cattle and bison has mainly depended on pour-on macrocyclic lactone formulations resulting in widespread ivermectin resistance. Consequently, there is an increased interest in using benzimidazole drugs which have been used comparatively little in cattle and bison in recent years. This situation, together with our understanding of benzimidazole resistance genetics, provides a practical opportunity to use deep-amplicon sequencing to assess the risk of drug resistance emergence. In this paper, we use deep-amplicon sequencing to scan for those mutations in the isotype-1 β-tubulin gene previously associated with benzimidazole resistance in many trichostrongylid nematode species. We found that several of these mutations occur at low frequency in many cattle and bison parasite populations in North America, suggesting increased use of benzimidazole drugs in cattle has the potential to result in widespread emergence of resistance in multiple parasite species. This work illustrates a post-genomic approach to large-scale surveillance of early emergence of anthelmintic resistance in the field.

The efficacy of eight essential oils (EOs) (Solidago canadensis, Eucalyptus globulus, Pelargonium asperum, Ocimum basilicum, Thymus vulgaris, Mentha piperita, Cymbopogon citratus and Cymbopogon martinii) against gastrointestinal nematodes (GINs) was evaluated using eggs collected from naturally infected cattle and cultured infective larvae (L3). The larvae species cultured from the faecal samples and subjected to two in vitro tests were Haemonchus spp. (55.5%), Trichostrongylus spp. (28.0%), Cooperia spp. (15.0%) and Oesophagostomum spp. (1.5%). The genus of EO Cymbopogon (C. citratus and C. martinii) showed the highest anthelmintic activity at the dose of 8.75 mg/ml, for the egg hatch, the larval migration and mortality assays. All of the EOs tested reduced egg hatching to rates <19.0%, compared to the controls (water and water + Tween 20) that had rates >92.0%. Cymbopogon citratus and C. martinii treatments resulted in 11.6 and 8.1% egg hatch, had the lowest migration of larvae through sieves, 60.5 and 54.9%, and the highest mortality rates, 63.3 and 56.3%, respectively. Dose–response tests showed that EO from C. citratus had the lowest larval LC50 and migration inhibition concentration (IC50) values of 3.89 and 7.19 mg/ml, respectively, compared to two other EOs (C. martinii and O. basilicum). The results suggest that EOs from the genus Cymbopogon can be interesting candidates for nematode control in cattle, although it may prove challenging to deliver concentrations to the gastrointestinal tract sufficient to effectively manage GINs.

Premunition is the state in a disease where an existing infection protects the host from reinfection with the same species. The cause of premunition is not clearly understood. In this study, we hypothesized that kin-selection might be a contributing factor in premunition. To test this theory, sheep were infected either once with a linguiform or smooth vulval morphotype of Haemonchos contortus, twice with the same morphotype or twice with different morphotypes. All infections resulted in a similar number of adult parasites. However, there were differences in the morphotypes recovered providing potential evidence of kin selection. Negative interference competition might also contribute to the reduction of the incoming population. Allelopathic or physical interactions between the parasites may be the mechanism behind the observed phenomena.

The levels of nematode egg production in goats and the availability of infective larvae (L3) on pasture were investigated on a dairy unit in New South Wales, Australia. The output of eggs by adult goats was always above 300 epg. The profile of the graph of larval availability in herbage paralleled those for temperature and rainfall, suggesting that larval peaks occurred when the temperature and availability of moisture were optimal. The dominant genus was Trichostrongylus, followed by Haemonchus, then Ostertagia. A larger proportion of Haemonchus larvae in the cultures of faeces were collected during the summer months.

Five experimental grass paddocks were sequentially contaminated with fresh bovine faeces containing known numbers of eggs of predominantly Haemonchus and Cooperia spp. during the 1984/85 dry season (November to March). Faecal, herbage and soil samples were examined at regular intervals between November and June in order to determine the rate of development and mortality of infective larvae (L3) in faeces, the pattern of herbage infestation with L3 and the role of faeces and soil as reservoirs of L3 during the dry season and early rains. L3 first appeared in faeces approximately 4 days post contamination (PC) and peak counts were obtained 28, 14, 7 and 14 days PC in the paddocks contaminated in December, January, February and March, respectively. The counts initially declined linearly at the rate of approximately 7535 L3/week and 10 947 L3/week in P2 and P4 respectively, due primarily to mortality but later there was an accelerated fall in the counts as the surviving L3 moved out of the faecal pads onto herbage. The overall trend of faecal larval populations in each paddock was therefore distinctly curvilinear. Although large numbers of L3 were present inside dry faecal pads in most paddocks throughout the dry season, none migrated on to herbage at that time of the year. Translation of L3 to herbage was very rapid and occurred simultaneously in all the paddocks 24 hours following the first heavy rainfall in late March. Consequently peak herbage infestations in all paddocks were coincident and occurred a few days after commencement of larval migration. The closer to the end of the dry season the contaminations were carried out, the larger were the subsequent early rains rise and the peak herbage infestation and the longer this infestation survived on herbage. No L3 were recovered from soil throughout the study, which suggests that faecal pads were the sole reservoir of L3 during the dry season and hence the source of the early rains herbage infestation.

A cDNA-AFLP (cDNA-Amplified Fragment Length Polymorphism)-based strategy has been used to identify levamisole (LEV) resistance markers in the nematode Haemonchus contortus. Transcript profiles of adult nematodes from two LEV-resistant and two susceptible isolates were compared. Among the 17 280 transcript-derived fragments (TDFs) amplified, 26 presented a polymorphic pattern between resistant and susceptible nematodes: 11 TDFs were present in both resistant isolates and absent from both susceptible isolates whereas 15 TDFs were present in both susceptible isolates and absent from both resistant isolates. 8 TDFs specifically present in resistant isolates were cloned and sequenced. Some of these TDFs could represent novel genes, as their sequences presented no homologies in databases. Interestingly, specific expression of one candidate (HA17) in resistant nematodes from different isolates was confirmed by RT-PCR experiments. The finding that HA17 expression correlates with LEV resistance in three H. contortus isolates vs five susceptible isolates strongly suggest that we identified a new potential marker of LEV resistance. This differential approach at the transcriptome level could be of great interest for the identification of the molecular mechanism involved in this phenotype.

This paper summarises the progress towards vaccine development against the major blood-feeding nematodes of man and livestock, the hookworms and Haemonchus contortus, respectively. The impact of the diseases and the drivers for vaccine development are summarized as well as the anticipated impact of the host immune response on vaccine design. The performance requirements are discussed and progress towards these objectives using defined larval and adult antigens, many of these being shared between species. Specific examples include the Ancylostoma secreted proteins and homologues in Haemonchus as well as proteases used for digestion of the blood meal. This discussion shows that many of the major vaccine candidates are shared between these blood-feeding species, not only those from the blood-feeding stages but also those expressed by infective L3s in the early stages of infection. Challenges for the future include: exploiting the expanding genome information for antigen discovery, use of different recombinant protein expression systems, formulation with new adjuvants, and novel methods of field testing vaccine efficacy.

The evidence for both between- and within- breed genetic variation for resistance to gastrointestinal (GI) nematode parasites is reviewed. It is concluded that much of the published research on breed characterisation for resistance suffers from poor experimental design. Prior to the initiation of a 6-year study that has just been completed in coastal Kenya there were no estimates of within-breed genetic variation (i.e. heritabilities) of resistance in sheep or goats in the tropics. This study has confirmed that Red Maasai sheep and Small East African (SEA) goats are more resistant to GI parasites (predominantly Haemonchus contortus) than Dorper sheep and Galla goats. Heritability estimates for logarithm transformed faecal egg counts (an indicator of resistance) in 8-month-old lambs was 0.18±0.08 for all lambs, but higher in the susceptible Dorper-sired lambs (0.35±0.16) than in the resistant Red Maasai-sired lambs (0.06±0.07). This difference in heritability suggests that many centuries of natural selection have fixed most of the genes for resistance in the Red Maasai sheep. The resistant Red Maasai sheep and SEA goats were two to three times more productive than the susceptible Dorper sheep and Galla goats in the sub-humid coastal Kenya environment.