Genotypic and phenotypic evaluation for benzimidazole resistance or susceptibility in Haemonchus contortus isolates

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• 作者：Waleed M. Arafa ; Patricia J. Holman ; Thomas M. Craig
• 关键词：Anthelmintic resistance ; Benzimidazole ; Beta tubulin gene ; Haemonchus contortus ; Larval development assay ; PCR ; Ruminant
• 刊名：Parasitology Research
• 出版年：2017
• 出版时间：February 2017
• 年：2017
• 卷：116
• 期：2
• 页码：797-807
• 全文大小：
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Medical Microbiology; Microbiology; Immunology;
• 出版者：Springer Berlin Heidelberg
• ISSN：1432-1955
• 卷排序：116

文摘

Haemonchus contortus isolates were evaluated for benzimidazole (BZ) resistance or susceptibility by allele-specific PCR based on β-tubulin isotype 1 gene polymorphisms at the F167Y, E198A, and F200Y sites. Two isolates, one presumed susceptible from wild pronghorn antelope (PH) and one known to be resistant from goats (VM), were also assayed phenotypically for BZ resistance or susceptibility in the larval development assay (Drenchrite®). The BZ EC50 was 0.198 μM (intermediate between susceptible and weak resistant) for PH with critical well 5 (intermediate between susceptible and weak resistant) and 1.456 μM (intermediate weak resistant and resistant) for VM with critical well 8.5 (resistant). Genotypically, DNA extracted from pooled VM L3 larvae in the Drenchrite® wells with the highest BZ concentration was homozygous susceptible (SS) at the F167Y and E198A sites and homozygous resistant (RR) at the F200Y site by PCR, and sequence analysis bore this out. PH L3 larvae DNA from a control well (no BZ) was SS at all three sites by PCR, confirmed by sequence analysis. All single adult worm samples (N = 21) from PH, VM, Egypt goat (EG), and a Texas llama were SS at F167Y and E198A by PCR; however, only 3 PH worms and 1 EG worm were SS at F200Y. Three additional PH worms were RS and upon cloning two clones were identified as resistant by sequencing and two as susceptible. Clones from single adult worms VM, llama, and EG samples that were RR by PCR at F200Y were sequence verified as resistant. In this study, F200Y was the most frequently found genotypic marker for BZ resistance or susceptibility in the different Haemonchus isolates.