An application of metabolomics to the detection of anthelmintic resistance of gastrointestinal nematodes in goats (METABOL-AR)

main classes of anthelmintics, including most commonly used – benzimidazoles. While the genetic background of resistance to benzimidazoles has been well clarified, little is known about the consequences of anthelmintic resistance for metabolism of gastrointestinal nematodes. The objective of METABOL-AR project is to quantify the metabolome of the larval stages of gastrointestinal nematodes susceptible and resistant to benzimidazoles. Therefore, in the METABOL-AR project we aim to employ currently the most advanced and sophisticated tool for investigating gastrointestinal nematodes metabolome – metabolomic MxP Quant 500 XL assay allowing to quantify over 1000 metabolites from over 30 chemical classes in the larvae of gastrointestinal nematodes. Larvae will be collected from various geographical and climatic regions of Europe so that an influence of various potential confounders (such as gastrointestinal nematodes species composition, temperature, and rainfall) on gastrointestinal nematodes metabolome can be precisely studied. A main goal will be to identify a set of metabolites significantly and independently linked to the presence of resistance to benzimidazoles. In the future quantification of these metabolites could serve as a simple, rapid and accurate method for diagnosing resistance to benzimidazoles in larvae isolated from fecal samples. This would provide an alternative for traditional in vitro and in vivo tests for resistance to benzimidazoles. To achieve this objective METABOL-AR team brings together 7 partners (research institutions) from 6 different countries representing different regions of Europe with different climate from humid subtropical and maritime to temperate continental. Additionally, the involvement of a private company as a subcontractor is assumed. Since the larvae will come from different countries the METABOL-AR will also provide a highly complex view of gastrointestinal nematodes species composition (referred to as nemabiome) and resistance to benzimidazoles prevalence and extent in various geographical and climatic regions of Europe.

The research objectives include:

a) determining the influence of geographical and climatic factors on larval metabolism,

b) identifying subtle differences in metabolomic profiles associated with changes in the species composition of gastrointestinal nematodes,

c) isolating a set of metabolites that enable precise differentiation between larvae susceptible and resistant to benzimidazoles,

d) obtaining data confirming that metabolomics can be a rapid and practical method for diagnosing anthelmintic resistance,

e) disseminating knowledge, implementing new methods, and conducting training on the diagnosis of anthelmintic resistance in countries where such techniques have not been previously applied,

f) collecting data on the prevalence of benzimidazole resistance in goat herds in regions where such information has so far been unavailable (Estonia, Latvia) or limited (Turkey),

g) gathering detailed data on the species composition and interactions of gastrointestinal nematode populations in goats across different climatic regions of Europe.

The results of the project will enable the development of a new diagnostic tool for detecting anthelmintic resistance at a very early stage of its development, using material that can be easily collected from animals in a non-invasive manner. This tool will be entirely based on the larval stages of parasites. The METABOL-AR project may make it possible to accurately diagnose drug resistance without the need for costly and labor-intensive in vivo or in vitro methods, as well as invasive sampling procedures. This approach will significantly reduce the number of animals required (by using pooled fecal samples and larval parasite stages), which may have a substantial positive impact on animal welfare.

Our research will also provide insights into new metabolic pathways associated with anthelmintic resistance and the adaptation of gastrointestinal nematodes to survive under different environmental and climatic conditions, indicating new directions for future research on gene expression related to these pathways in other groups of anthelmintic drugs as well