Multiplex allele-specific polymerase chain reaction-based prevalence study of canine trapped neutrophil syndrome in Thailand.

Background and Aim: Trapped neutrophil syndrome (TNS) is an autosomal recessive genetic disorder found in Border Collies and is characterized by peripheral neutropenia and myeloid hyperplasia of bone marrow. The underlying cause of TNS is associated with a 4-base pair deletion mutation in the vacuolar protein sorting 13 homolog B (VPS13B) gene. In this study, we proposed and validated a novel multiplex allele specific-polymerase chain reaction (MAS-PCR) technique to assess the prevalence of TNS using VPS13B genotypes of Border Collies and Thai Ridgebacks in Thailand. Materials and Methods: We assessed the prevalence of TNS in 100 Border Collies and 30 Thai Ridgebacks using MAS-PCR-based allelic discrimination technique of the VPS13B gene. We then confirmed the VPS13B genotypes by direct DNA sequencing. Results: A total of 130 samples were successfully genotyped using MAS-PCR assays. Of the two dog breeds examined, the VPS13B mutation was present in Border Collies, whereas Thai Ridgebacks were unaffected by this mutation. In Border Collies, 96% of dogs tested had an intact VPS13B genotype, whereas the remaining individuals had a heterozygous mutation genotype, with prevalence and mutated VPS13B allele frequencies of 4% and 2%, respectively. Conclusion: Using a novel MAS-PCR assay targeting the VPS13B gene, this study demonstrates for the first time that carriers of TNS exist in Border Collies in Thailand. This assay is a reliable and cost-effective tool for diagnosing TNS based on VPS13B genotypes and is suitable for routine clinical practice.

A novel multiplex polymerase chain reaction assay for the genotypic survey of the non-homologous end-joining factor 1 gene associated with Collie eye anomaly in Thailand.

Background and Aim: Collie eye anomaly (CEA) is a hereditary and congenital ocular disorder, which affects several dog breeds, including Collies, Collie-related breeds, and other purebreds. An intronic deletion of 7799-bp in the non-homologous end-joining factor 1 (NHEJ1) gene has been identified as the genetic defect causing CEA. This study aimed to investigate the prevalence of CEA based on NHEJ1 genotyping assay in Thailand. Materials and Methods: We clarified the prevalence of CEA in 224 dogs from five purebred dog breeds using a novel multiplex polymerase chain reaction (PCR)-based technique and confirmed the genotypic status with direct DNA sequencing. Results: The highest frequency of the mutated NHEJ1 allele was 83.3% for Rough Collies, followed by 7.8% for Border Collies, 5.1% for Australian Shepherds, and 2.8% for Shetland Sheepdogs. The heterozygous mutated NHEJ1 genotype detected for Rough Collies, Border Collies, Australian Shepherds, and Shetland Sheepdogs was 33.3%, 15.6%, 10.3%, and 3.3%, respectively. The homozygous mutated NHEJ1 genotype was detected only in Rough Collies and Shetland Sheepdogs, accounting for 66.7% and 1.1%, respectively. Thai Ridgeback Dogs were not affected by this mutation. Conclusion: This study describes, for the 1st time, the genotypic survey of the NHEJ1 gene associated with CEA in dogs in Thailand. In addition, we successfully developed a new multiplex PCR assay with high accuracy, reproducibility, and cost-efficiency and validated its usefulness for determining NHEJ1 genotypes.

Prevalence of the MDR1 gene mutation in herding dog breeds and Thai Ridgebacks in Thailand.

BACKGROUND AND AIM: A canine multi-drug resistance 1 (MDR1) nt230(del4) is a well-known inherited disorder that primarily affects collies and various herding breeds. The most recognized clinical implication for affected dogs is associated with an increased risk of multiple drug toxicity. To date, MDR1 gene mutations have been identified globally, especially in dogs from the USA and European countries. This study aimed to investigate the prevalence of MDR1 nt230(del4) in herding dog breeds and Thai Ridgebacks in Thailand. MATERIALS AND METHODS: We clarified the prevalence of MDR1 nt230(del4) in 263 dogs of eight purebred dog breeds in Thailand using an allele-specific multiplex polymerase chain reaction method and direct DNA sequencing. RESULTS: Rough Collies, Australian Shepherds, Shetland Sheepdogs, and Old English Sheepdogs were affected by the mutation with mutant allelic frequencies of 57.14%, 12.82%, 11.28%, and 8.33%, respectively. Among these populations, the prevalence of the MDR1 (+/-) genotype was 57.14% (12/21) for Rough Collies, 25.64% (10/39) for Australian Shepherds, 16.13% (15/93) for Shetland Sheepdogs, and 16.67% (2/12) for Old English Sheepdogs, whereas the MDR1 (-/-) mutation was only identified in Rough Collies and Shetland Sheepdogs, with prevalences of 28.57% (6/21) and 3.22% (3/93), respectively. However, the MDR1 nt230(del4) was not identified in Border Collies, German Shepherds, White Swiss Shepherds, or Thai Ridgebacks. CONCLUSION: This study provides the current situation regarding MDR1 nt230(del4) in herding dog breeds in Thailand. In this survey, we investigated for the first time the status of MDR1 genotype in Thai Ridgebacks. These results are helpful for veterinarians managing effective therapeutic plans for commonly affected dog breeds, and these results will encourage all breeders to improve their selective breeding programs based on the MDR1 nt230(del4) status.