A reverse transcription-multiplex PCR strategy devised for concomitant detection and differentiation of foot and mouth disease virus serotypes O, A and Asia 1 in India.

Serotype identification occupies the central part of foot and mouth disease (FMD) diagnosis workflow and vaccination decision tree. In this study, a reverse transcription-multiplex PCR (RT-mPCR) strategy wherein three assays with unique combinations of serotype specific primers targeting the VP1 region was developed to differentiate FMD virus serotypes O, A and Asia 1 based on differential size of the PCR amplicons on agarose gel. Their diagnostic performance relative to the mPCR assay in use in India was evaluated on 169 clinical samples and 210 cell culture grown virus isolates. The relative diagnostic sensitivity was found to be 99.69%, 98.78% and 99.08% for primer combinations 1, 2 and 3, respectively. These assays proved their worth by detecting serotype in three FMD suspected specimens that went undiagnosed in the existing mPCR and also by identifying multiple serotypes in the same sample. Their detection limits varied from log10 2 to log10 4 viral RNA dilution and from 100 to 0.1 TCID50 virus depending on the serotype. The validated novel mPCR assays show promise to be included in the routine diagnostic tool-box to augment the efficiency of diagnosis of FMD virus serotypes that display extreme genetic diversity and a tendency of transboundary dispersal.

Foot-and-Mouth Disease Virus Serotype O Exhibits Phenomenal Genetic Lineage Diversity in India during 2018-2022.

In India, widespread foot-and-mouth disease (FMD) outbreaks occurred in 2021. The objective of this study was to identify genetic lineages and evaluate the antigenic relationships of FMD virus (FMDV) isolates gathered from outbreaks reported between 2019 and 2022. Our study shows that the lineages O/ME-SA/Ind2001e and the O/ME-SA/Cluster-2018 were both responsible for the FMD outbreaks on an epidemic scale during 2021. This observation is in contrast to earlier findings that suggested epidemic-scale FMD outbreaks in India are often connected to a single genetic lineage. Additionally, we report here the identification of the O/ME-SA/PanAsia-2/ANT10 sub-lineage in India for the first time, which was connected to two intermittent outbreaks in Jammu and Kashmir. The current study demonstrates that the O/ME-SA/ind2001e lineage has a strong presence outside of the Indian subcontinent. Furthermore, the O/ME-SA/Cluster-2018 was observed to have a wider geographic distribution than previously, and like the O/ME-SA/Ind2001d and O/ME-SA/Ind2001e lineages in the past, it may eventually spread outside of its geographic niche. For O/ME-SA/Ind2001e and O/ME-SA/Cluster-2018, the predicted substitution rate for the VP1 region was 6.737 × 10-3 and 8.257 × 10-3 nt substitutions per site per year, respectively. The time of the most recent common ancestor of the O/ME-SA/Ind2001e and O/ME-SA/Cluster-2018 strains suggests that the viruses possibly emerged during 2003-2011 and 2009-2017, respectively. Recent sightings of the O/ME-SA/PanAsia2/ANT10 virus in India and the O/ME-SA/Ind2001e virus in Pakistan point to possible cross-border transit of the viruses. The results of a two-dimensional viral neutralization test revealed that all of the field isolates were antigenically matched to the currently used Indian vaccine strain O INDR2/1975. These results suggest that the serotype O vaccine strain can protect against outbreaks brought on by all three circulating lineages.

Emergence of a novel genetic lineage 'A/ASIA/G-18/2019' of foot and mouth disease virus serotype A in India: A challenge to reckon with.

Foot and mouth disease (FMD) has engendered large scale socioeconomic crises on numerous occasions owing to its extreme contagiousness, transboundary nature, complicated epidemiology, negative impact on productivity, trade embargo, and need for intensive surveillance and expensive control measures. Emerging FMD virus variants have been predicted to have originated and spread from endemic Pool 2, native to South Asia, to other parts of the globe. In this study, 26 Indian serotype A isolates sampled between the year 2015 and 2022 were sequenced for the VP1 region. BLAST and maximum likelihood phylogeny suggest emergence of a novel genetic group within genotype 18, named here as 'A/ASIA/G-18/2019' lineage, that is restricted so far only to India and its eastern neighbour, Bangladesh. The lineage subsequent to its first appearance in 2019 seems to have displaced all other prevalent strains, in support of the phenomenon of 'genotype/lineage turnover'. It has diversified into two distinct sub-clusters, reflecting a phase of active evolution. The rate of evolution of the VP1 region for the Indian serotype A dataset was estimated to be 6.747 × 10-3 substitutions/site/year. India is implementing a vaccination centric FMD control programme. The novel lineage showed good antigenic match with the proposed vaccine candidate A IND 27/2011 when tested in virus neutralization test, while the existing vaccine strain A IND 40/2000 showed homology with only 31% of the isolates. Therefore, in order to combat this challenge of antigenic divergence, A IND 27/2011 could be the preferred strain in the Indian vaccine formulations.