Goats are more susceptible to Haemonchus contortus infection than sheep under similar experimental settings.

Due to differences in their feeding behavior, sheep and goats are often assumed to respond differently to helminth infections. The present study compared Haemonchus contortus infection profile between sheep and goats under the same experimental setting. Experimental infection was conducted using a randomized block design in four groups of intact sheep (InfSH and ConSH) and goats (InfG, and ConG). Groups InfSH and InfG (N = 7 each) received 10,000L3 of H. contortus whereas the control groups ConSH and ConG (N = 7 each) remained uninfected. Faecal egg counts and PCV were measured from Day 0 to day 56 post infection (PI). On day 56 PI, animals were humanely slaughtered and abomasal contents were recovered to measure worm burden, worm length and in utero egg count. The findings show that: (1) Haemonchus infected animals showed an increase in FEC starting from day 21 PI, (2) progressive reduction in PCV was registered from day 7 PI and continued to the end of the experiment while this remained at pre-infection levels in control groups, (3) FEC was much higher (P < 0.001) and PCV was significantly lower (P < 0.05) in infected goats than in infected sheep, (4) at necropsy, total worm burden with worm establishment rates of 63% and 28.87% were registered respectively for infected goats and sheep with significant difference (P < 0.05), (5) Female worms were significantly longer (P < 0.05) in InfG (22.8(± 1.2) compared to InfSH (20.5 ± 0.67 mm) while (6) Mean worm fecundity was 974.8 ± 239.4 and 1162.5 ± 89.4 respectively for groups InfSH and InfG with no significant difference (P = 0.07), and (7) Parasite traits such as worm burden, FEC and female worm length were well correlated in sheep whereas few such patterns were observed in goats. In conclusion, under the same experimental infection, Arsi-Bale goats are more susceptible to H. contortus infection than Arsi-Bale sheep and hence deserve special attention when they are forced to live on grazing rather than browsing.

Prevalence status and detection of benzimidazole resistance using AS-PCR in Haemonchus contortus of goats from Marathwada region, Maharashtra, India.

This study examined Haemonchus contortus prevalence and benzimidazole resistance in eight districts of Marathwada, Maharashtra, India. A comprehensive investigation of 264 abomasa of goats collected from abattoirs and goats necropsied at the College of Veterinary Sciences and Animal Husbandry, Parbhani, revealed 21.21 % a prevalence of H. contortus. The incidence of H. contortus did not vary much across seasons and it was highest in summer (23.42 %), followed by monsoon (22.89 %), and lowest in winter (15.71 %). Statistically non-significant (p < 0.05) prevalence was observed in male and female animals. A detailed examination of 168 adult H. contortus worms from eight districts revealed the presence of all conceivable genotypes including homozygous resistant (rr), susceptible (SS), and heterozygous (Sr) BZ susceptible genotypes. The rr was the most frequent at 50 %, followed by SS at 27 % and Sr at 22 %. The presence of the SNP was observed in in all eight randomly selected and sequenced samples.

Characterisation of seryl tRNA synthetase (srs-2) in Haemonchus contortus and Teladorsagia circumcincta.

The aim of the study was to purify and characterise recombinant proteins with the potential as an anti-parasite vaccine. Full-length cDNAs encoding seryl-tRNA synthetase (srs-2) were cloned from Haemonchus contortus (HcSRS-2) and Teladorsagia circumcincta (TcSRS-2). TcSRS-2 and HcSRS-2 cDNA (1458bp) encoded proteins of 486 amino acids, each of which was present as a single band of about 55 kDa on SDS-PAGE. Multiple alignments of the protein sequences showed homology of 94% between TcSRS-2 and HcSRS-2, 76-93% with SRS-2s of eight nematodes and 68% with Mus musculus SRS-2. The predicted three-dimensional structures revealed an overall structural homology of TcSRS-2 and HcSRS-2, highly conserved binding and catalytic sites, and minor differences in the tautomerase binding site residues in other nematode SRS-2 homologues. A phylogenetic tree was constructed using helminth and mammalian SRS-2 sequences. Soluble C-terminal SRS-2 proteins were expressed in Escherichia coli strain AY2.4 and purified. Recombinant HcSRS-2 assay shows that the recombinant enzyme was active and stable. The Km and Vmax for ATP were 3.9 ± 1.0 µM and 2.7 ± 0.1 µmol min-1 mg-1 protein, respectively. Antibodies in serum and saliva from field-immune, but not nematode-naïve, sheep recognised recombinant HcSRS-2 and TcSRS-2 in enzyme-linked immunosorbent assays. Recognition of the recombinant proteins by antibodies generated by exposure of sheep to the native enzyme indicates similar antigenicity of the two proteins.

The first molecular detection of benzimidazole resistance in Haemonchus contortus from sheep in Hejing and Minfeng counties of Southern Xinjiang.

To understand the benzimidazole (BZ) resistance of Haemonchus contortus in Southern Xinjiang, three single nucleotide polymorphisms (SNPs) designated as F167Y, E198A, and F200Y, in the isotype-1 ß-tubulin gene which are associated with BZ resistance, were investigated for H. contortus populations from sheep in Hejing and Minfeng counties of Southern Xinjiang. In brief, a total of 190 H. contortus adults were collected from 52 out of 70 slaughtered sheep in city abattoirs across two regions in Southern Xinjiang. The species identity of each adult worm was confirmed by PCR amplification of ITS-2 using H. contortus-specific primers targeting the ITS-2. The samples were then investigated for BZ-related SNPs at locus 167, 198, and 200, by PCR-sequencing of the isotype-1 ß-tubulin gene. The results showed that only E198A and F200Y mutations were detected in the investigated H. contortus populations. The E198A mutation (homozygous and heterozygote resistant: found in 40% and 30% of sequenced samples from Minfeng and Hejing counties, respectively) was predominant compared with the F200Y mutation (homozygous and heterozygote resistant: found in 14% and 13.3% of sequenced samples from Minfeng and Hejing counties, respectively). The results indicate a high prevalence of BZ resistance in H. contortus populations from certain areas of Southern Xinjiang. Our findings provide valuable information for the prevention and control of H. contortus in areas with similar conditions.

Expression of Toll-like receptors in Haemonchus Contortus resistant sheep: An innate immune parameter for host defense against gastrointestinal nematode infection.

Innate immune parameters, a first line of defense against invading pathogens like bacteria, parasites, fungi, etc, play a significant role in the prevention and elimination of aetiological agents primarily by recognition of invading pathogen-specific molecules by different pattern recognition receptors. Toll-like receptors (TLRs), a type-I transmembrane glycoprotein, cause innate immune responses mainly by produing inflammatory cytokines, chemokines and interferons. The objective of present study was to determine the role of TLRs in parasite resistance in Malpura sheep. In the current study, transcript variation of TLRs and its downstream signalling molecules namely MyD88, TRIF, IRF-3, TRAF, TGF-ß, NFκB, and CD14 were ascertained by real-time PCR in Haemonchus contortus resistant (R) and susceptible (S) Malpura sheep. Results have shown significantly (P<0.05) up-regulated expression of TLR-2, TLR-4, TLR-5, TLR-8 and TLR-10 in July however down-regulated patterns were observed in August and September in R-line sheep compared to S-line sheep. This indicates that at more or less equal parasite load, the TLR genes in R sheep produce more transcripts, but after parasite loads have increased hugely in the S line, they easily surpass the levels seen in the S line. Result suggests that transcriptional activity of the TLR genes was related to parasite load and there were differences between the lines at different infection intensities. Three-point transcript expression observation of the signalling molecules namely TRIF, IRF-3, TRAF, a similar pattern was observed in R sheep compared with S sheep.

Immunohistochemistry analyses of the abomasal mucosa show differences in cellular-mediated immune responses to Haemonchus contortus infection in resistant and susceptible young lambs.

Haemonchus contortus is known for its high pathogenicity in sheep, and the uncontrolled use of anthelmintics resulted in the emergence of multiple drug-resistant populations. Breeding sheep for gastrointestinal nematode resistance is a sustainable alternative to reduce dependence of anthelmintic drugs, and differences in the degree of resistance between breeds have been reported. Here we compare two sheep breeds (Santa Ines and Ile de France), concerning the differences in innate and adaptive immune response involved in the resistance against H. contortus infection. Immunohistochemical analyses of the abomasum were conducted in naïve Santa Ines (n = 14) and Ile de France (n = 12) lambs randomized into four groups: infected Santa Ines (n = 8), non-infected control Santa Ines (n = 6), infected Ile de France (n = 8), and non-infected control Ile de France (n = 4). The infected lambs were initially infected with H. contortus infective larvae at 14 days of age, and multiple infections were conducted every second day until they reached 66 days of age. There was a significant effect (P < 0.001) of the infection with increase in numbers of CD3+ T; CD79α+ B; GATA3+ Th2/ILC2; POU2F3+ tuft cells; FOXP3+ T reg; and IgE + cells in the fundus of the abomasal mucosa in both Santa Ines and Ile de France lambs. Nevertheless, the infected Santa Ines lambs presented the highest averages for CD79α+ B; GATA3+ Th2/ILC; IgE + cells; and POU2F3+ tuft cells and there was a significant association of the breed and infection status with regards to POU2F3+ tuft cells, with the highest mean in the infected Santa Ines group. The infected Santa Ines group had three lambs with high degree of resistance and five lambs that showed a moderate infection. Our results suggest a mechanism of synergistic coordination between different immune-cell types in promoting resistance of suckling lambs under H. contortus infection.

Molecular signatures of Haemonchus contortus infection in sheep: A comparative serum proteomic study on susceptible and resistant sheep breeds.

Due to the negative impact of Haemonchus contortus in the tropics and subtropics, the detection of serum protein profiles that occur in infected sheep is of high relevance for targeted selective treatment strategies (TST). Herein, we integrated proteomics with phenotypic traits to elucidate physiological mechanisms associated to H. contortus infection in susceptible (Dorper - D) and resistant (Santa Inês - S) sheep breeds. Naïve female lambs were infected with H. contortus third-stage larvae on day zero (D0), and samples were collected weekly, for 28 days. Feces were used for individual fecal egg counts (FEC) blood for packed cell volume (PCV) and serum for specific antibody quantification through ELISA. Sera was collected on D0 (-) and D21 (+), and analyzed using a LC-MS/MS based proteomics approach. FEC, PCV, and anti-H. contortus antibody levels confirmed the absence of infection on D0. On D28 there was a significant difference between the two breeds for logFEC means (D = 3774 and S = 3141, p=0.033) and PCV means (D = 16.3 % and S = 24.3 %, p=0.038). From a total of 754 proteins identified, 68 differentially abundant proteins (DAPs) were noted. Phosphopyruvate hydratase (ENO3) was a DAP in all comparisons, while S+ vs D+ and S- vs D- shared the highest number of DAPs (8). Each of the four experimental groups clustered separately in a principal component analysis (PCA) of protein profile. Among the DAPs, proteins associated with the innate and adaptive immune system were detected when comparing S- vs D- and S+ vs D+. In D-, some proteins were linked to stress response to handling, sampling and heat. Focusing on the consequences of infection in each breed, in the D+ vs D- comparison, upregulated proteins were associated with inflammation control and immune response, where downregulated proteins pointed to a negative impact of infection on tissue anabolism, compromising muscle growth and fat deposition. In the S+ vs S- comparison, upregulated proteins were related to immune response, while the downregulated proteins were possibly linked to muscular development and growth, impaired by infection. Collectively, it can be concluded that ENO3 regulation emerges as a potential factor underlying the differential immune response observed between Santa Inês and Dorper sheep infected with H. contortus. In turn, detected acute phase proteins (APPs) reinforce their relation with infection, inflammation and stress conditions, whereas THEMIS-like may contribute to the immune system in Dorper. GSDMD, Guanylate-binding protein and ACAN warrant further investigation as possible biomarkers for TST strategy development.

Haemonchus contortus alters distribution and utilization of protein and amino acids in different tissues of host sheep.

The objective was to determine host animal protein/amino acid redistribution and use among the abomasum, duodenum and muscle of sheep infected with Haemonchus contortus. Sixteen male Ujumqin sheep (32.4 ± 3.9 kg) were dewormed and randomly assigned to two groups, infected or not infected with H. contortus (GIN and CON). The GIN group had lower (P < 0.05) dry matter intake, average daily gain, and live body weight than CON, with extensive focal infiltration of lymphocytes in the lamina propria and bottom of the abomasal epithelium. In the abomasum and duodenum, there were 100 and 220 genes, respectively, that were up-regulated, whereas 56 and 149 were down-regulated. In the abomasum, the most enriched KEGG pathways were related to immunity and inflammation reaction, including: viral protein interaction with cytokine and cytokine receptor (P = 0.017), influenza A (P = 0.030), IL-17 signaling pathway (P = 0.030). In the duodenum, KEGG pathways were more enriched in nutrient metabolism, including pancreatic secretion (P < 0.001), protein digestion and absorption (P < 0.001), graft-versus-host disease (P = 0.004). Furthermore, most genes related with the above KEGG pathways were increased in the abomasum but decreased in the duodenum. Amino acid profiles in abomasum and duodenum of CON and GIN groups were clustered in a partial least-squares discriminant analysis model, with significant changes in 36 and 19 metabolites in abomasal and duodenal chyme, respectively. Further confirmed by transcriptome-targeted metabolome association analysis, GIN mainly enhanced metabolism of arginine and sulphur amino acids in abomasum and those metabolic pathways were associated. Meanwhile, GIN mainly decreased pyruvate related amino acid metabolism in duodenum. Moreover, concentrations of Arg (P = 0.036), His (P = 0.027), and Cys (P = 0.046) in longissimus thoracis et lumborum were decreased in GIN, whereas concentrations of Gly (P = 0.012) and Ala (P = 0.046) were increased. In conclusion, H. contortus enhanced metabolism of arginine and sulphur amino acids in the abomasum; decreased pyruvate metabolism in the duodenum; and drove more protein/amino acids for abomasal tissues to resist physical and immune damage, reducing protein and amino acids in duodenum and muscle for support host growth. Specific nutrients (such like arginine, histidine, and cysteine) may play important role in control gastrointestinal nematode infection for ruminant.

In vivo anthelmintic activity of Combretum mucronatum schumach & Thonn leaf extract against Haemonchus contortus in goats.

Parasitic infections with gastrointestinal nematodes are a serious problem for the health and welfare of domestic animals and negatively affect the economics of animal production. Haemonchus contortus is a haematophagous nematode of small ruminants responsible for significant mortality and morbidity. In addition, the widespread resistance to synthetic anthelmintic drugs emphasizes the urgent need of alternative treatment options against haemonchosis. This work aims to investigate the anthelmintic activity of an hydroethanolic Combretum mucronatum leaf extract (CMLE) against Haemonchus contortus in goats. Goats were artificially infected with 3500 third-stage larvae of H. contortus, and 21 days later, treated with CMLE (1000, 500, 250 mg/kg) for 4 consecutive days. Different parameters such as faecal egg count reduction, weight and haematocrit were monitored during the experimental period. The number of eggs per gram of faeces (EPG) was concentration-depended lower and significantly reduced compared to the untreated control (p < 0.0001). The effect of the highest CMLE dose (4 ×1000 mg/kg body weight) was similar to the effect of albendazole (1 ×5 mg/kg of body weight). The ED50 and ED90 values calculated were 189.17 and 392.33 mg/kg body weight respectively. ED50 and ED90 values were time-dependent. Moreover, CMLE improved haematocrit and weight of goats in dose-dependent and time-dependent manner. These results showed that CMLE could be used for haemonchosis treatment in small ruminants.

Efficacy of a Haemonchus contortus vaccine under field conditions in young alpacas.

Anthelmintic resistance to Haemonchus contortus creates increasing management challenges with small ruminants and camelids. The commercial vaccine, Barbervax®, contains H11 and H-gal-GP antigens, derived from gut mucosal membrane enzymes of H. contortus involved in digesting blood. Antibody neutralization of these antigens causes failure of H. contortus to digest blood, resulting in parasite death. H11 and H-gal-GP are considered "hidden" antigens, meaning the host immune system does not encounter these proteins under natural infection. Therefore, repeat immunization is required to maintain protective humoral responses. One previous study evaluated the safety of Barbervax® in camelids but the efficacy could not be assessed due to lack of successful infection in the controls. The objective of the current study was to evaluate clinical parameters of anemia, fecal egg counts (FECs), and humoral immune responses of healthy alpacas after immunizing with Barbervax® compared to non-vaccinated controls, all under natural environmental exposure on parasite-laden pastures. A crossover-like study was performed where twenty alpacas (298 ± 66 days of age) were assigned to be initially vaccinated with Barbervax® (n=10) or receive no treatment (n=10). Three doses of Barbervax® were administered at three-week intervals. Feces and blood were collected on Day -10, 0, 21, 43, 64, 85, 106, and 135 to evaluate FECs, packed cell volume (PCV), and antibody titers. Each group was kept on separate adjacent pastures. Tracer sheep (n=2 per study group) were introduced on Day 43 for a three-week period to ensure parasite acquisition. For the crossover-like component on Day 85, the initial non-vaccinated group was administered Barbervax® with dosing repeated on Day 106 and 135. Results indicated all initially vaccinated alpacas produced antibody titers to vaccine antigen that corresponded to lower mean FECs compared to the initially non-vaccinated group. A reduced mean FEC in the vaccinate group was observed 21 days after peak antibody titers. Similarly, when pooled vaccinate antibody titers were noted to wane on Day 106, an increase in FEC was observed at the following time point (Day 135). Conclusions from our study support the use of Barbervax® to reduce H. contortus burdens in alpacas. Furthermore, a less than 30-day lag time between antibody titer and resultant effect in FECs was observed. Additional studies assessing the ability of Barbervax® to reduce H. contortus burdens during subsequent grazing seasons would provide even greater information regarding the use of Barbervax® within alpaca herds to modulate H. contortus infections, refugia, and anthelmintic use.

High rates of benzimidazole-resistance-associated alleles in Haemonchus contortus and detection of resistance against macrocyclic lactones in strongylids from German alpaca herds.

The population of South American camelids (SAC) has been steadily growing in Europe, where they are confronted with the regional endoparasite population of ruminants. As there are no anthelmintic drugs registered for use against nematode infections in SACs, anthelmintics (AH) available for ruminants or horses are usually applied. Reports indicating potential failures in administered AH are increasing. However, the generally low egg counts in SACs complicate the application of resistance tests in the field. The present study reports a follow-up study on SAC farms where anthelmintic resistance (AR) was suspected. The aims were (i) to repeat faecal egg count reduction tests (FECRTs) on potentially affected farms identified in a previous study with larger sample sizes, (ii) to verify suspected AR of Haemonchus contortus against benzimidazoles (BZ) by performing a single-nucleotide polymorphism (SNP) analysis using digital polymerase chain reaction (dPCR), and (iii) to apply the mini-FLOTAC technique for more reliable results at low egg counts in line with current recommendations. Seven farms (9-46 animals each) were examined by coproscopy, larval differentiation and SNP analysis. A FECRT was performed on six of these farms with moxidectin (three farms), monepantel (two farms) and ivermectin (one farm). The FEC was calculated according to the current World Association for the Advancement of Veterinary Parasitology (WAAVP) guidelines with the clinical protocol (a newly introduced variant of FECRT which can be used for smaller sample sizes and lower egg counts on the cost of sensitivity) and an expected efficacy of 99%. A high level (> 90%) of BZ-resistance-associated SNPs on codon 200 of H. contortus was observed on all farms. With the FECRT, resistance was demonstrated for ivermectin (74% FECR), while it remained inconclusive for one farm for moxidectin treatment. Sustained efficacy was demonstrated for the remaining treatments. This study showed an advanced level of BZ resistance in H. contortus of SACs and the development of AR against macrocyclic lactones on some farms. Thus, constant monitoring of AH treatment and sustainable worm control methods both need to be applied.

Regulatory effect of Balanites aegyptiaca ethanol extract on oxidant/antioxidant status, inflammatory cytokines, and cell apoptosis gene expression in goat abomasum experimentally infected with Haemonchus Contortus.

This experiment aimed to assess the regulatory effects of treatment with Balanites aegyptiaca fruit ethanol extract (BA-EE) on oxidant/antioxidant status, anti-inflammatory cytokines, and cell apoptosis gene expression in the abomasum of Haemonchus contortus-infected goats. Twenty goat kids were assigned randomly to four equal groups: (G1) infected-untreated, (G2) uninfected-BA-EE-treated, (G3) infected-albendazole-treated, (G4) infected-BA-EE-treated. Each goat in (G1), (G3), and (G4) was orally infected with 10,000 infective third-stage larvae. In the fifth week postinfection, single doses of albendazole (5 mg/kg.BW) and BA-EE (9 g/kg.BW) were given orally. In the ninth week postinfection, the animals were slaughtered to obtain abomasum specimens. The following oxidant/antioxidant markers were determined: malondialdehyde (MDA), glutathione (GSH), glutathione-S-transferase (GST), superoxide dismutase (SOD), catalase (CAT). The mRNA gene expression of cytokines (IL-3, IL-6, IL-10, TNF-α) and cell apoptosis markers (Bax, Bcl-2) were estimated. (G1) showed significantly reduced GSH content and GST and SOD activities but a markedly increased MDA level. (G3) and (G4) revealed a markedly lower MDA level with pronouncedly elevated GSH, SOD, and GST levels. The antioxidant properties of BA-EE were superior to those of albendazole. The mRNA gene expressions of IL-3, IL-6, IL-10, TNF-α, and Bax-2 were upregulated in (G1) but downregulated in (G3) and (G4). Bcl-2 and Bcl-2/Bax ratio expression followed a reverse course in the infected and both treated groups. We conclude that BA-EE treatment has a protective role in the abomasum of H. contortus-infected goats. This could be attributed to its antioxidant properties and ability to reduce pro-inflammatory cytokines and cell apoptosis.

Molecular study of the transcription factor SKN-1 and its putative relationship with genes that encode GST and antioxidant enzymes in Haemonchus contortus.

Haemonchus contortus is a parasitic nematode of ruminants. Once inside its host, it is exposed to reactive oxidative species and responds by synthesising antioxidant enzymes as a defence. In Caenorhabditis elegans, antioxidant genes are regulated by the transcription factor skinhead-1 (Cel-SKN-1). However, there is little information about the function of SKN-1 in H. contortus (Hco-SKN-1). Therefore, we performed a molecular investigation to characterise Hco-SKN-1 and its putative relationship with genes encode antioxidant enzymes, namely glutathione S-transferases (Hco-GSTs, n = 3), superoxide dismutase (Hco-SOD) and catalase (Hco-CAT), which are involved in haematophagy and defence against the host. We used in silico sequence analysis of Hco-SKN-1 and Hco-GSTs to design and perform relative expression assays involving H. contortus eggs, infective larvae (L3) and adults. Furthermore, we exposed H. contortus transitional infective larvae (xL3) to erythrocytes or hydrogen peroxide (H2O2) and evaluated the relative expression of antioxidant genes at 24 or 48 h. Gene Ontology (GO) analysis revealed 31 functions associated with Hco-SKN-1 and Hco-GSTs, including stress resistance, larval development and the active immune response. Hco-GST-5957 and Hco-SOD showed the highest expression in adults, indicating a relationship with specific functions at this mature stage. xL3 exposed to erythrocytes or H2O2 showed significant upregulation of Hco-SKN-1, but it occurred after upregulation of the antioxidant genes, indicating that these genes are not regulated by Hco-SKN-1 during the blood-feeding stage. Additional investigation is necessary to understand the putative regulation of antioxidant genes by Hco-SKN-1 during the blood-feeding stage.

Comparison of P-glycoprotein gene expression of two Haemonchus contortus isolates from Yucatan, Mexico, with resistant or susceptible phenotype to ivermectin in relation to a susceptible reference strain.

The variability in the expression of different P-glycoprotein (P-gp) genes in parasitic nematodes of ruminants such as Haemonchus contortus (Hco-pgp) may be caused by different factors including nematode biology, geographical region and anthelmintic pressure. This study analysed the relative expression level of 10 P-gp genes in two H. contortus (Hco-pgp) field isolates from Yucatan, Mexico: 1) PARAISO (IVM-resistant) and 2) FMVZ-UADY (IVM-susceptible). These isolates were compared with a susceptible reference isolate from Puebla, Mexico, namely "CENID-SAI". In all cases H. contortus adult males were used. The Hco-pgp genes (1, 2, 3, 4, 9, 10, 11, 12, 14 and 16) were analysed for each isolate using the RT-qPCR technique. The Hco-pgp expressions were pairwise compared using the 2-ΔΔCt method and a t-test. The PARAISO isolate showed upregulation compared to the CENID-SAI isolate for Hco-pgp 1, 3, 9, 10 and 16 (P < 0.05), and the PARAISO isolate showed upregulation vs. FMVZ-UADY isolate for Hco-pgp 2 and 9 (P < 0.05), displaying 6.58- and 5.93-fold differences (P < 0.05), respectively. In contrast, similar Hco-pgp gene expression levels were recorded for FMVZ-UADY and CENID-SAI isolates except for Hco-pgp1 (P <0.1), which presented a significant upregulation (6.08-fold). The relative expression of Hco-pgp allowed confirming the IVM-resistant status of the PARAISO isolate and the IVM-susceptible status of the FMVZ-UADY isolate when compared to the CENID-SAI reference isolate. Therefore, understanding the association between the Hco-pgp genes expression of H. contortus and its IVM resistance status could help identifying the genes that could be used as molecular markers in the diagnosis of IVM resistance. However, it is important to consider the geographic origin of the nematode isolate and the deworming history at the farm of origin.

Variants of NLRP3 Protein in Haemonchus contortus Infected Sheep: Impact on Immune Cell Responsiveness to LPS In Vitro.

Pathogen recognition is an essential component to achieve the desired outcome of host protection. Nod-like receptor pyrin containing domain 3 (NLRP3) is a cytoplasmic pattern recognition receptor (PRR) with a wide array of agonists, such as PAMPs, DAMPs, ATP, bacterial product and viral products. Stimulation of the NLRP3 inflammasome results in proteolytic activation of IL-1ß and IL-18, cell pyroptosis and classically, the induction of proinflammatory responses. St. Croix (STC) sheep have resistance traits exhibiting the appropriate T-helper type 2 immune response ensuing protection during helminth parasitic infection whereas parasite-susceptible Suffolk (SUF) sheep have an impaired response resulting in parasite establishment and adverse symptoms. The objective of these experiments was to determine if NLRP3 protein in H. contortus-infected SUF sheep was defective using the classical activation pathway of NLRP3 inflammasome. Peripheral blood mononuclear cells (PBMCs) derived from H. contortus-infected STC and SUF sheep were isolated from whole blood and treated (MCC950 treatment for 2 h followed by LPS treatment for 3 h, 1400 W treatment for 2 h followed by LPS treatment for 3 h, LPS treatment for 3 h or culture media for 3 h). qPCR analysis of LPS-stimulated PBMC revealed an upregulation in inflammatory associated genes IL-1ß, TLR4, TNFα and NFκB (p < 0.0001) in STC PBMC and downregulation in IFNγ, IL-6 and iNOS for SUF PBMC. Pharmacological inhibition of iNOS in SUF PBMC resulted in an upregulation in the expression of IFNγ. These preliminary data begin to discover a relationship between NLRP3 activation and TLR4 signalling in PBMC of STC and SUF sheep.

Beyond parasitism: Exploring the microbial profile of Haemonchus contortus and its predilection site (abomasum) in Kashmir Merino sheep.

Gastrointestinal helminth infection, particularly by Haemonchus contortus, poses significant challenges to sheep farming worldwide. While anthelmintic drugs have been traditional control measures, the emergence of resistance calls for alternative strategies. Understanding the interaction between parasites, host, and their microbiome is crucial for management of helminth infection. This study intricately explores the interactions between microbial communities in Kashmir Merino sheep infected with H. contortus, to understand the complex interplay between host, parasite, and their microbiome. Sheep abomasal contents and H. contortus were collected from infected and control groups, processed for DNA extraction, and subjected to metagenomic sequencing of the 16 S rRNA gene. Downstream analysis unveils distinct microbial patterns, where Proteobacteria were dominant in H. contortus, while Bacteroidota and Firmicutes prevailed in the sheep abomasum. The revelation of unique genera and shifts in diversity indices underscored helminth-induced disruptions in the host. Beta diversity analysis further showed significant variations in bacterial profiles, providing insights into the intricate host, parasite, and microbiome dynamics. Additionally, this study elucidated the presence of pathogenic bacteria within H. contortus, accentuating their potential role in exacerbating sheep health issues. This finding underscores the complexity of the host-parasite-microbiome interaction showing helminth-induced microbiome alterations of the host.

Divergent humoral responses between males and females against 24 kDa excretory-secretory protein of Haemonchus contortus and influence of ovine ß-globin polymorphism.

Lambs harboring the Hb-AA ß-globin haplotype present improved cell-mediated responses and increased resistance against Haemonchus contortus infection. The aim of the present study was to compare the effect of sex and ß-globin haplotypes on specific humoral responses and phenotypes of resistance during H. contortus infection in Morada Nova sheep. As expected, females displayed stronger resistance during the first and second experimental challenges. Differential systemic humoral immune responses were observed comparing sex groups, in which higher levels of specific antibodies targeting 24 kDa excretory-secretory (ES24) protein of H. contortus of IgG and IgM antibodies were respectively observed as predominant isotypes in males and females. The IgM levels were significantly correlated with phenotypes of resistance, evaluated by packed cell volume and fecal egg counts. To our knowledge this is the first study reporting divergent humoral responses profiles to H. contortus infection between male and female sheep. The impact of ß-globin haplotypes was less pronounced in females compared to males. Notably, only males showed significant weight differences across haplotypes, with Hb-AA lambs being the heaviest. Additionally, Hb-AA males had significantly higher PCV (indicating better red blood cell health) and lower FEC (indicating lower parasite burden). These findings suggest a more pronounced effect of ß-globin polymorphisms on H. contortus infection in males, potentially due to their generally weaker resistance compared to females. This study highlights the importance of sex and ß-globin haplotypes in shaping immune responses to H. contortus infection. Specifically, IgM antibodies targeting the ES24 protein appear to play a crucial role in host-parasite interactions and may hold promise for therapeutic development.

Rodents Used for the Propagation of Hookworms and Haemonchus contortus.

Helminthiasis due to hookworm infestations in humans and Haemonchus contortus in sheep cause untold levels of disease and economic losses worldwide. Drug resistance is an ever-growing problem with pathogenic helminths. Thus, there is a critical need for new treatment strategies for hookworms and H. contortus that depends on animal models. Because hook- worms and H. contortus are obligate parasites, they can only be maintained using live animals. This review describes use of the Syrian golden hamster to propagate hookworms and Mongolian gerbil to propagate H. contortus.

Analysis of lncRNA-related studies of ivermectin-sensitive and -resistant strains of Haemonchus contortus.

In this study, 858 novel long non-coding RNAs (lncRNAs) were predicted as sensitive and resistant strains of Haemonchus contortus to ivermectin. These lncRNAs underwent bioinformatic analysis. In total, 205 lncRNAs significantly differed using log2 (difference multiplicity) > 1 or log2 (difference multiplicity) < - 1 and FDR < 0.05 as the threshold for significant difference analysis. We selected five lncRNAs based on significant differences in expression, cis-regulation, and their association with the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathways. These expressions of lncRNAs, namely MSTRG.12610.1, MSTRG.8169.1, MSTRG.6355.1, MSTRG.980.1, and MSTRG.9045.1, were significantly downregulated. These findings were consistent with the results of transcriptomic sequencing. We further investigated the relative expression of target gene mRNAs and the regulation of mRNA and miRNA, starting with lncRNA cis-regulation of mRNA, and constructed a lncRNA-mRNA-miRNA network regulation. After a series of statistical analyses, we finally screened out UGT8, Unc-116, Fer-related kinase-1, GGPP synthase 1, and sart3, which may be involved in developing drug resistance under the regulation of their corresponding lncRNAs. The findings of this study provide a novel direction for future studies on drug resistance targets.

Potential distribution modelling for Haemonchus contortus (Nematoda: Trichostrongylidae) in South America.

The species Haemonchus contortus occurs in many regions worldwide, mainly parasitising small ruminants and economically impacting animal production. Climate change is considered a driving force for the risk of diseases caused by helminths and can also affect relationships between parasites and their hosts, with the potential to cause losses in both animal production and biodiversity in general. The aim of this study was to model the potential distribution of H. contortus in South America. We used MaxEnt to perform the analyses and describe the contribution of important bioclimatic variables involved in the species distribution. Our results show that H. contortus colonised most of the areas with habitats that suit the species' environmental requirements and that this parasite presents habitat suitability in a future scenario. Understanding the effects of climate change on the occurrence and distribution of parasite species is essential for monitoring these pathogens, in addition to predicting the areas that tend to present future parasite outbreaks and identify opportunities to mitigate the impacts of the emergence of diseases caused by these organisms.