Dataset Information

Absence of mutations associated with resistance to benzimidazole in the beta-tubulin gene of Ascaris suum.

ABSTRACT: INTRODUCTION:Benzimidazoles are commonly used for the control of veterinary nematodes. Resistance to benzimidazoles has been associated with three single nucleotide polymorphisms in the ?-tubulin gene of common nematodes. However, these mutations are infrequent in the genus Ascaris spp. METHODS:In order to determine mutations associated with benzimidazole resistance in Ascaris suum, worms were collected from slaughtered pigs and a partial region of the ?-tubulin gene was sequenced. RESULTS:All parasites showed the wildtype genotype for codons 167, 198, and 200 of the ?-tubulin gene. CONCLUSIONS:This is the first report of genetic sequences associated with benzimidazole resistance in A. suum.

SUBMITTER: Palma A

PROVIDER: S-EPMC7094045 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Absence of mutations associated with resistance to benzimidazole in the beta-tubulin gene of Ascaris suum.

Palma Adalid A Matamoros Gabriela G Escobar Denis D Sánchez Ana Lourdes AL Fontecha Gustavo G

Revista da Sociedade Brasileira de Medicina Tropical 20200316

<h4>Introduction</h4>Benzimidazoles are commonly used for the control of veterinary nematodes. Resistance to benzimidazoles has been associated with three single nucleotide polymorphisms in the β-tubulin gene of common nematodes. However, these mutations are infrequent in the genus Ascaris spp.<h4>Methods</h4>In order to determine mutations associated with benzimidazole resistance in Ascaris suum, worms were collected from slaughtered pigs and a partial region of the β-tubulin gene was sequenced ...[more]

PMID: 32187331

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Project description:BackgroundThe treatment coverage of control programs providing benzimidazole (BZ) drugs to eliminate the morbidity caused by soil-transmitted helminths (STHs) is unprecedently high. This high drug pressure may result in the development of BZ resistance in STHs and so there is an urgent need for surveillance systems detecting molecular markers associated with BZ resistance. A critical prerequisite to develop such systems is an understanding of the gene family encoding β-tubulin proteins, the principal targets of BZ drugs.Methodology and principal findingsFirst, the β-tubulin gene families of Ascaris lumbricoides and Ascaris suum were characterized through the analysis of published genomes. Second, RNA-seq and RT-PCR analyses on cDNA were applied to determine the transcription profiles of the different gene family members. The results revealed that Ascaris species have at least seven different β-tubulin genes of which two are highly expressed during the entire lifecycle. Third, deep amplicon sequencing was performed on these two genes in more than 200 adult A. lumbricoides (Ethiopia and Tanzania) and A. suum (Belgium) worms, to investigate the intra- and inter-species genetic diversity and the presence of single nucleotide polymorphisms (SNPs) that are associated with BZ resistance in other helminth species; F167Y (TTC>TAC or TTT>TAT), E198A (GAA>GCA or GAG>GCG), E198L (GAA>TTA) and F200Y (TTC>TAC or TTT>TAT). These particular SNPs were absent in the two investigated genes in all three Ascaris populations.SignificanceThis study demonstrated the presence of at least seven β-tubulin genes in Ascaris worms. A new nomenclature was proposed and prioritization of genes for future BZ resistance research was discussed. This is the first comprehensive description of the β-tubulin gene family in Ascaris and provides a framework to investigate the prevalence and potential role of β-tubulin sequence polymorphisms in BZ resistance in a more systematic manner than previously possible.

Project description:Infections by parasitic nematodes inflict a huge burden on the health of humans and livestock throughout the world. Anthelmintic drugs are the first line of defense against these infections. Unfortunately, resistance to these drugs is rampant and continues to spread. To improve treatment strategies, we must understand the genetics and molecular mechanisms that underlie resistance. Studies of the fungus Aspergillus nidulans and the free-living nematode Caenorhabditis elegans discovered that a beta-tubulin gene is mutated in benzimidazole (BZ) resistant strains. In parasitic nematode populations, three beta-tubulin alleles, F167Y, E198A, and F200Y, have long been correlated with resistance. Additionally, improvements in sequencing technologies have identified new alleles - E198V, E198L, E198K, E198I, and E198Stop - also correlated with BZ resistance. However, none of these alleles have been proven to cause resistance. To empirically demonstrate this point, we independently introduced the F167Y, E198A, and F200Y alleles as well as two of the newly identified alleles, E198V and E198L, into the BZ susceptible C. elegans N2 genetic background using the CRISPR-Cas9 system. These genome-edited strains were exposed to both albendazole and fenbendazole to quantitatively measure animal responses to BZs. We used a range of concentrations for each BZ compound to define response curves and found that all five of the alleles conferred resistance to BZ compounds equal to a loss of the entire beta-tubulin gene. These results prove that the parasite beta-tubulin alleles cause resistance. The E198V allele is found at low frequencies along with the E198L allele in natural parasite populations, suggesting that it could affect fitness. We performed competitive fitness assays and demonstrated that the E198V allele reduces animal health, supporting the hypothesis that this allele might be less fit in field populations. Overall, we present a powerful platform to quantitatively assess anthelmintic resistance and effects of specific resistance alleles on organismal fitness in the presence or absence of the drug.

Dataset Information

Absence of mutations associated with resistance to benzimidazole in the beta-tubulin gene of Ascaris suum.

Publications

Absence of mutations associated with resistance to benzimidazole in the beta-tubulin gene of Ascaris suum.

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