Anthelmintic resistance of gastrointestinal cattle nematodes

Anthelmintic resistance of parasites in small ruminants, cattle and horses is increasing worldwide as a consequence of the over usage of the currently available products. In Belgium, Cooperia oncophora is the most common cattle nematode in which resistance, especially against macrocyclic lactones, occurs. Once resistance has been diagnosed, a change to another drug with a different mode of action is advised. However, effective anthelmintics will be hardly available in the near future. Therefore, it is important that farmers and veterinarians find a balance between achieving good parasite control and the sustainability of their control strategies. In this way, anthelmintic resistance may be delayed, and the effectiveness of anthelmintic drugs may be prolonged. This requires sensitive detection tools. With a sensitive detection technique, anthelmintic resistance can be diagnosed in a very early stage. Hence, the spread of resistance alleles in the parasite population may be prevented. In this review, different diagnostic assays for the detection of anthelmintic resistance are discussed, an overview is given of the current status of anthelmintic resistance in Belgian cattle, and measures are suggested to avoid or delay the development of anthelmintic resistance

Transcriptome analyses reveal protein and domain families that delineate stage-related development in the economically important parasitic nematodes, Ostertagia ostertagi and Cooperia oncophora

BACKGROUND: Cooperia oncophora and Ostertagia ostertagi are among the most important gastrointestinal nematodes of cattle worldwide. The economic losses caused by these parasites are on the order of hundreds of millions of dollars per year. Conventional treatment of these parasites is through anthelmintic drugs; however, as resistance to anthelmintics increases, overall effectiveness has begun decreasing. New methods of control and alternative drug targets are necessary. In-depth analysis of transcriptomic data can help provide these targets. RESULTS: The assembly of 8.7 million and 11 million sequences from C. oncophora and O. ostertagi, respectively, resulted in 29,900 and 34,792 transcripts. Among these, 69% and 73% of the predicted peptides encoded by C. oncophora and O. ostertagi had homologues in other nematodes. Approximately 21% and 24% were constitutively expressed in both species, respectively; however, the numbers of transcripts that were stage specific were much smaller (~1% of the transcripts expressed in a stage). Approximately 21% of the transcripts in C. oncophora and 22% in O. ostertagi were up-regulated in a particular stage. Functional molecular signatures were detected for 46% and 35% of the transcripts in C. oncophora and O. ostertagi, respectively. More in-depth examinations of the most prevalent domains led to knowledge of gene expression changes between the free-living (egg, L1, L2 and L3 sheathed) and parasitic (L3 exsheathed, L4, and adult) stages. Domains previously implicated in growth and development such as chromo domains and the MADF domain tended to dominate in the free-living stages. In contrast, domains potentially involved in feeding such as the zinc finger and CAP domains dominated in the parasitic stages. Pathway analyses showed significant associations between life-cycle stages and peptides involved in energy metabolism in O. ostertagi whereas metabolism of cofactors and vitamins were specifically up-regulated in the parasitic stages of C. oncophora. Substantial differences were observed also between Gene Ontology terms associated with free-living and parasitic stages. CONCLUSIONS: This study characterized transcriptomes from multiple life stages from both C. oncophora and O. ostertagi. These data represent an important resource for studying these parasites. The results of this study show distinct differences in the genes involved in the free-living and parasitic life cycle stages. The data produced will enable better annotation of the upcoming genome sequences and will allow future comparative analyses of the biology, evolution and adaptation to parasitism in nematodes

Detection and mechanisms of macrocyclic lactone resistance in the bovine nematode Cooperia oncophora

Wereldwijd beperken infecties met gastro-intestinale nematoden het welzijn en de productiviteit van de veestapel. De meest voorkomende gastro-intestinale rondwormen bij rundvee in gematigde klimaatzones zijn Ostertagia ostertagi (in de lebmaag) en Cooperia oncophora (in de dunne darm). Controle van deze parasieten steunt voornamelijk op het gebruik van breedspectrum anthelminthica met een persisterende werking, met name de macrocyclische lactones (ML’s) zoals ivermectine, abamectine, doramectine, eprinomectine en moxidectine. Het intensieve en frequente gebruik van deze anthelminthica heeft echter geleid tot de selectie van ML-resistentie in deze economisch belangrijke maag-darm nematoden

Molecular mechanism of macrocyclic lactone resistance in the cattle parasites Ostertagia ostertagi and Cooperia oncophora

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Gene expression analysis of ABC transporters in a resistant Cooperia oncophora isolate following in vivo and in vitro exposure to macrocyclic lactones

Members of the ATP-binding cassette (ABC) transporter family (P-glycoproteins, Half-transporters and Multidrug Resistant Proteins) potentially play a role in the development of anthelmintic resistance. The aim of this study was to investigate the possible involvement of ABC transporters in anthelmintic resistance in the bovine parasite, Cooperia oncophora. Partial sequences of 15 members of the ABC transporter protein family were identified, by mining a transcriptome dataset combined with a degenerate PCR approach. Reverse transcriptase PCR showed that most of the ABC transporters identified were constitutively transcribed throughout the life cycle of C. oncophora. Constitutive differences in gene transcript levels between a susceptible and resistant isolate were only observed for Con-haf-9 and Con-mrp-1 in eggs of the resistant isolate, while no differences were observed in L3 or the adult life stage. Analysis of resistant adult worms, collected from calves 14 days after treatment with either ivermectin or moxidectin, showed a significant 3- to 5-fold increase in the transcript levels of Con-pgp-11 compared to non-exposed worms. Interestingly, a 4-fold transcriptional up-regulation of Con-pgp-11 was also observed in L3 of the resistant isolate, after in vitro exposure to different concentrations of ivermectin, whereas this effect was not observed in exposed L3 of the susceptible isolate. The results suggest that the worms of this particular resistant isolate have acquired the ability to up-regulate Con-pgp-11 upon exposure to macrocyclic lactones. Further work is needed to understand the genetic basis underpinning this process and the functional role of PGP-11

Potential contribution of P-glycoproteins to macrocyclic lactone resistance in the cattle parasitic nematode <em>Cooperia oncophora</em>

International audienceResistance against macrocyclic lactones such as ivermectin is widespread among parasitic gastrointestinal nematodes of small ruminants and is rapidly increasing in cattle parasites. ABC transporters of the subfamily B, the so-called P-glycoproteins (Pgps) have been frequently implicated in ivermectin resistance and are a major cause of multi-drug resistance in protozoa and helminths. The Pgp inhibitor verapamil (VPL) dramatically enhanced susceptibility of the cattle parasitic nematode Cooperia oncophora to ivermectin in vitro as measured in a larval developmental assay and a larval migration inhibition assay using third stage larvae. Moreover, VPL completely restored susceptibility to ivermectin in a resistant isolate resulting in virtually identical dose-response curves of susceptible and resistant isolates in the presence of VPL. Further characterisation of the molecular mechanisms resulting in Pgp-mediated ivermectin resistance is still hampered by the lack of molecular and biochemical information for Pgps of parasitic nematodes. Using PCR with degenerate primers, fragments of four different C. oncophora Pgps could be amplified and the Conpgp-2, previously implicated in macrocyclic lactone resistance in Haemonchus contortus, and Conpgp-3 full-length cDNAs were obtained by RACE PCR. The pgp sequences presented here contribute important data required to systematically screen resistant C. oncophora isolates for up- or down-regulation of Pgps and for the detection of single nucleotide polymorphisms in Pgps to detect selection of specific Pgp alleles by anthelmintics as early as possible