Available online 3 May 2024, 108763

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Nearly half of the 35 brown dog tick isolates screened displayed amitraz resistance

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Exon-3 region of putative octopamine receptor gene was amplified and sequenced

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Three novel non-synonymous mutations detected in the exon-3 region of resistant samples

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Molecular docking revealed decreased binding efficiency of mutated receptor with amitraz

Abstract

The brown dog tick or Rhipicephalus sanguineus sensu lato is an ixodid tick, responsible for the dissemination of pathogens that cause canine infectious diseases besides inflicting the direct effects of tick bite. The hot humid climate of Kerala, a south Indian state, is favorable for propagation of tick vectors and acaricides are the main stay of tick control. Though the resistance against synthetic pyrethroids is reported among these species, the status of amitraz resistance in R. sanguineus s. l. in the country is uncertain due to the lack of molecular characterization data and scarce literature reports. Hence the present study was focused on the phenotypic detection and preliminary genotypic characterisation of amitraz resistance in the R. sanguineus s. l. A modified larval packet test (LPT) on a susceptible isolate was performed to determine the discriminating dose (DD). Further LPT-DD on 35 tick isolates was carried out to detect amitraz resistance robustly, along with that full dose response bioassays on the resistant isolates were performed. The results indicated that amitraz resistance is prevalent with 49 per cent of the samples being resistant. Amplification of exon 3 of octopamine receptor gene from both the susceptible and resistant larval isolates was carried out. Amplicons of ten pooled amitraz susceptible and ten pooled amitraz resistant representative samples were sequenced and analysed, unveiling a total of three novel non-synonymous mutations in the partial coding region at positions V32A, N41D and V58I in phenotypically resistant larval DNA samples. In silico analysis by homology modelling and molecular docking of the mutated and unmutated receptors showed that these mutations had reduced the binding affinity to amitraz. However, lack of mutations in the octopamine receptor gene in three of the pooled low order resistant R. sanguineus s. l. larval samples could be suggestive of other mechanisms associated with amitraz resistance in the region. Hence, further association studies should be carried out to confirm the association of these mutations with target insensitivity in R. sanguineus s. l. ticks, along with exploring the status of metabolic resistance and other mechanisms of resistance.

Section snippetsINTRODUCTION

Ticks are obligate blood sucking ectoparasites globally recognized as vectors of various pathogens. Recent studies have highlighted the predominantly zoonotic nature of tick-borne diseases, which have become a focal point of contemporary veterinary and medical research. Ticks induce several direct pathological responses in hosts, including anemia, irritability, and allergic reactions, that complicate the management of infestation. As obligate hematophages, ticks require host blood to transition

Sample collection

Samples were collected from naturally infested dogs in different districts of Kerala including Malappuram, Thrissur, Ernakulam and Thiruvananthapuram (northern latitudes 8º 18’ and 12º 48’ and eastern longitudes 74º 52’ and 77º 22’). Fully engorged female ticks were collected from kennel grounds and partially engorged female ticks attached to the host body were manually removed from naturally infested dogs at various sites including veterinary hospitals, pet grooming centers, animal birth

Resistance patterns of field isolates

The LC50 and LC95 values obtained for the susceptible isolate from Thiruvananthapuram were determined to be 131.7 ppm and 600 ppm, respectively and the calculated DD was 1200 ppm. A total of 35 field tick isolates (approximately 350 ticks) were screened with the LPT-DD, of which 18 isolates (51%) were found to be susceptible and 17 isolates (49%) were resistant. Among the 17 tick isolates that were resistant to amitraz, 13 were highly resistant (37%), one was moderately resistant (3%) and three

DISCUSSION

A meta-analysis of literature pertaining to acaricidal resistance in cattle ticks across the world revealed that 87.5% (77/88) of the studies reported the selection of resistance mostly to deltamethrin, cypermethrin, amitraz and ivermectin with resistance development prevalence estimates of 76.4, 74.1, 58.8 and 61 per cents, respectively. Both phenotypic and genotypic characterisation of amitraz resistance is widely studied in cattle ticks (Dzemo et al., 2022). Resistance against different

CONCLUSION

In conclusion, the study characterised the exon 3 of gene encoding the octopamine receptor of amitraz susceptible and resistant R. sanguineus s. l. ticks for the first time, reporting three novel non-synonymous mutations that could be linked to amitraz resistance. The presence of these mutations in the highly resistant and moderately resistant tick isolates emphasises that these SNPs could have conferred resistance. In addition, molecular docking of the mutated and unmutated receptor models

Uncited reference

Balabanidou et al., 2018; Corley et al., 2013; D’Alessandro et al., 2022; Ducornez et al., 2005; FAOFood and Agriculture Organization of the United Nations, 1971; Kemp, 1998; McTier et al., 2016; Raj et al., 2022; Rodríguez-Hidalgo et al., 2017; Sunkara et al., 2021; World Health Organization, 2006.

Declaration of Competing Interest

The authors report no declarations of interest.

Acknowledgements:

The authors are grateful to the Dean and Director of Research, College of Veterinary and Animal Sciences, KVASU for providing the facilities. Thanks are due to Staff, Undergraduate students (KVASU) and field veterinarians for rendering the help in sample collection.

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