Genetic variants in dyf-7 validated by droplet digital PCR are not drivers for ivermectin resistance in Haemonchus contortus

International Journal for Parasitology. Drugs and Drug Resistance
Safaa T ElmahalawyJohan Höglund

Abstract

Resistance to ivermectin (IVM) in the nematode Haemonchus contortus in small ruminants is an increasing problem throughout the world. Access to molecular diagnostics will allow early detection of IVM resistance, which in turn can limit the spread of resistant isolates. One candidate gene which has recently been suggested as a marker for IVM resistance is that for dye-filling protein (dyf-7). In this study, we critically investigated the suitability of A141G and G153T single nucleotide polymorphisms (SNP) of dyf-7 as a marker in larval cultures collected from sheep farms in Sweden, involving several isolates for which resistance status had been characterised by the faecal egg count reduction test (FECRT). Initially, we designed dyf-7 primers from a worldwide collection of adult Haemonchus contortus DNA. With the sequence data, we created a haplotype network. We then optimised and used the same sets of primers and probes in a droplet digital PCR (ddPCR) assay for precise quantification of dyf-7 allele frequencies in pre- and post-anthelmintic treatment faecal larval cultures. The fractional abundance (FA) of the mutant SNP was within the range 7.8 and 31%. However, the FA was generally stable in samples collected from the same fa...Continue Reading

Datasets Mentioned

BETA
KF927016.1
KF927017.1
KF927018.1
and
KF927019.1
MH172331-MH172361

Methods Mentioned

BETA
PCR
electrophoresis

Software Mentioned

Primer3Plus
QuantaSoft
Codon Code Aligner
Popart

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