Comparison of ovine beta-globin haplotype sequences and a new multiplex PCR for identification

Author(s)

Okino, Cintia Hiromi ^[1] ; Frizoni Caminotto, Lara Cristina ^[1] ; Ikeda Kapritchkoff, Rafaela Tami ^[2] ; Meo Niciura, Simone Cristina ^[1] ; Esteves, Sergio Novita ^[1] ; dos Santos, Isabella Barbosa ^[2] ; Barbosa Toscano, Joao Henrique ^[2] ; Oliveira, M. C. S. ^[1] ; Chagas, A. C. S. ^[1]

Affiliation

^[1] Embrapa Pecuaria Sudeste, Rodovia Washington Luiz, Km 234 S-N, POB 339, BR-13560970 Sao Carlos, SP - Brazil

^[2] Univ Estadual Paulista UNESP, Fac Ciencias Agr & Vet, Via Acesso Prof Paulo Donato Castellane S-N, BR-14884900 Jaboticabal, SP - Brazil

• Document type: Journal article
• Source: Veterinary Parasitology; v. 300, DEC 2021.
• Web of Science Citations: 0

Abstract

Deletion of pre-adult beta(C)-globin in sheep harboring BB haplotype of beta-globin was associated to decreased tolerance to anemia and hypoxia, and consequently, reduced resistance to Haemonchus contortus infection, which is closely related to severe anemia. Recently, a qPCR using hydrolysis probe was successfully developed for beta-globin haplotype identification, and association between resistance against H. contortus and presence of beta(A) allele was observed in Morada Nova sheep. Thus, this study aimed to better investigate the differences between beta-globin haplotypes and to develop a conventional multiplex PCR, as an alternative to qPCR assay for beta-globin haplotype identification. A total of 333 Morada Nova lambs had their blood collected and tested by both qPCR and new multiplex PCR, and 100 % of agreement was observed between the results. Since different primers were designed for such assay development, including different target genes, high specificity of both methods may be also highlighted. Three A haplotype samples were submitted to DNA Sanger sequencing of beta-globin gene and compared to sequences previously deposited in Genbank. One nucleotide deletion in intronic region was observed only in AA haplotype of Morada Nova animals, while in BB animals the nucleotide remained present. To the best of our knowledge, this is the first report of multiplex conventional PCR for ovine beta-globin haplotype identification. The advantages of the developed conventional PCR are reduced reagents costs (less than a half price) and wider reachability, since even labs without real time PCR thermocyclers are able to offer this assay. Therefore, it may become an important tool for sheep producers to improve genetic selection of parasite resistant animals. (AU)

• FAPESP's process: 19/02929-3 - Target selective treatment based on liveweight gain as nematode control strategy in Morada Nova lamb
• Grantee: Ana Carolina de Souza Chagas
• Support Opportunities: Regular Research Grants
• FAPESP's process: 18/16070-1 - Study of the impact of gastrointestinal nematodes infection on the performance of Morada Nova Lambs for development of a targeted selective treatment
• Grantee: Isabella Barbosa dos Santos
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 17/00373-2 - Characterization of local immune responses associated with the parasitic resistance phenotype in morada nova lambs
• Grantee: João Henrique Barbosa Toscano
• Support Opportunities: Scholarships in Brazil - Master