Phenotypic characterization of recombinant Marek’s disease virus in live birds validates polymorphisms associated with virulence

Authors: Kim, Taejoong; Hearn, Cari; Mays, Jody; VELEZ-IRIZARRY, DEBORAH - Orise Fellow; REDDY, SANJAY - Texas A&M University; Spatz, Stephen; Cheng, Hans; Dunn, John

Submitted to: Viruses
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/14/2023
Publication Date: 11/16/2023
Citation: Kim, T.N., Hearn, C.J., Mays, J.K., Velez-Irizarry, D., Reddy, S.M., Spatz, S.J., Cheng, H.H., Dunn, J.R. 2023. Phenotypic characterization of recombinant Marek’s disease virus in live birds validates polymorphisms associated with virulence. Viruses. 15(11):2263. https://doi.org/10.3390/v15112263.
DOI: https://doi.org/10.3390/v15112263

Interpretive Summary: Marek’s disease virus (MDV) has evolved toward more virulent partly because the current vaccines cannot prevent MDV infection in vaccinated birds. With recent studies of MDV genomes with known virulence, we identified the nucleotide differences in a specific area of the MDV genome. Using the molecularly cloned highly virulent MDV strain 686, specific nucleotides were replaced with a less virulent MDV. A total of nine nucleotide replacements in eight genes within the MDV genome were introduced and verified by sequence analysis. The virulence of the nucleotide-replaced virus was reduced to that of the parental virus. The virulence rank determination with the comparison to the reference viruses indicates that the reduction of virulence rank is also confirmed. This study provides evidence that multiple nucleotide differences contributed to the virulence of MDV. The data will help to diagnose the virulence of new MDV isolates by rapid genome analysis and rational vaccine development by targeted modification of the virus genome.

Technical Abstract: Marek’s disease (MD) is a highly infectious lymphoproliferative disease in chickens with a significant economic impact. Mardivirus gallidalpha 2, also known as Marek’s disease virus (MDV), is the causative pathogen and has been categorized based on its virulence rank into four pathotypes: mild (m), virulent (v), very virulent (vv), and very virulent plus (vv+). A prior comparative genomics study suggested that several single-nucleotide polymorphisms (SNPs) and genes in the MDV genome are associated with virulence, including nonsynonymous (ns) SNPs in eight open reading frames (ORF): UL22, UL36, UL37, UL41, UL43, R-LORF8, R-LORF7, and ICP4. To validate the contribution of these nsSNPs to virulence, the vv+MDV strain 686 genome was modified by replacing nucleotides with those observed in the vMDV strains. Pathogenicity studies indicated that these substitutions reduced the MD incidence and increased the survival of challenged birds. Furthermore, using the best-fit pathotyping method to rank the virulence, the modified vv+MDV 686 viruses resulted in a pathotype similar to the vvMDV Md5 strain. Thus, these results support our hypothesis that SNPs in one or more of these ORFs are associated with virulence but, as a group, are not sufficient to result in a vMDV pathotype, suggesting that there are additional variants in the MDV genome associated with virulence, which is not surprising given this complex phenotype and our previous finding of additional variants and SNPs associated with virulence.