ANTIMICROBIAL SUSCEPTIBILITY PROFILE OF METHICILLIN RESISTANT AND METHICILLIN SENSITIVE STAPHYLOCOCCUS AUREUS FROM BOVINE MILK IN THE STATE OF HARYANA, INDIA

Bovine mastitis is the principal leading cause of monetary losses to dairy farmers. This disease impacts the udder health as well as the quantitative and qualitative parameters of milk. The disease is multi-etiological, but Staphylococcus aureus (S. aureus) contributing to intramammary infections is the principal cause. Our work intended to go through the sensitivity pattern of S. aureus obtained from milk samples of bovines. The samples used in the study were received at College Central Laboratory, LUVAS, Hisar. The bovine milk samples were inoculated on blood agar to obtain bacterial isolates, followed by morphological and biochemical characterization. S. aureus was confirmed by phenotypic as well as molecular assays. Ninety-five staphylococci were preliminarily isolated from 381 quarter milk samples based on morphological features of a bacterial colony, Gram stain, catalase reaction, oxidase test, and HiStaph latexTM kit from bovines. Out of these, 86 S. aureus isolates were confirmed based on phenotypic (mannitol salt agar) as well as a molecular test (23S rRNA PCR). All these isolates were used for sensitivity profiling by Kirby Bauer disc diffusion method. Maximum sensitivity was observed for chloramphenicol and doxycycline, least against cloxacillin and methicillin. From 86 isolates, 62.79% were found to be multidrug-resistant (MDR). From MDR isolates, 11.11% were extensively drug-resistant (XDR) and none were pan-drug-resistant (PDR). The presence of a high percentage of MDR phenotypes among S. aureus isolates in this study draws our attention that treatment of animals must be carried out after the identification of pathogens, followed by patterns of sensitivity to antimicrobials

Detection of anti-3AB3 non-structural protein antibodies in foot-and-mouth disease virus vaccinated buffaloes at a semi-organized farm of Madhya Pradesh in India

Foot-and-mouth disease (FMD), one of the most important viral diseases of cloven-hoofed animals in India, is caused by FMD virus (FMDV) which belongs to Aphthovirus in the family Picornaviridae. There are three serotypes of FMDV (O, A and Asia 1) circulating amongst the livestock population of India. FMD is characterized by the formation of vesicles especially over the tongue and in between the interdigital space. FMD-affected animals with tongue lesions are reluctant to feed and subsequently yield less milk as well as affected animals never regain their production status causing huge economic losses to the animal owners. In the present study, random whole blood samples from FMD-vaccinated 38 adult buffaloes were collected in sterile containers of a semi-organized buffalo farm of Madhya Pradesh in India. Purified FMD vaccines only elicit antibodies (that are protective) against structural proteins of FMDV while natural FMDV infection invokes antibodies against both structural and non-structural proteins. Serum samples were employed in recombinant 3AB3 non-structural protein-based enzyme-linked immunosorbent assay (3AB3 NSP-ELISA kit provided by ICAR-Directorate of FMD, Mukteshwar) for differentiation of FMD-infected and vaccinated animals (DIVA). A total of 10.53% (4/38) serum samples tested positive in DIVA. Largely the vaccinated animals remained protected as no clinical signs of the disease were observed reiterating the importance of regular FMDV vaccination in animals at semi-organized dairy farms

VP1 region-based molecular characterization of foot-and-mouth disease virus serotype O from clinical cases in Haryana, India

Abstract Background Foot-and-mouth disease (FMD) is a highly transmissible viral infection affecting cloven-hoofed animals, significantly impacting livestock productivity and trade in India. The causative agent, FMD virus (FMDV), exhibits considerable genetic and antigenic diversity, with serotype O being most prevalent. Haryana, a northern Indian state with a large susceptible livestock population, has reported sporadic FMD cases despite ongoing vaccination efforts. This study aims to detect, type, and characterize FMDV from suspected animals in Haryana during 2024, with a focus on genetic variations in the VP1 region compared to the vaccine strain and contemporary isolates. Results Clinical samples from suspected cases were analyzed using RT-multiplex PCR, confirming the presence of FMDV O in five buffaloes from two districts. The partial P1 region was amplified and sequenced from five samples using 1C/2B gene-specific primers. Phylogenetic analysis classified all five samples (GenBank Accession Nos. PV068827–PV068830 & PV288326) within the Middle East-South Asia (ME-SA) topotype based on 1D gene sequences (639 nt). Four samples (PV068827–PV068830) exhibited 12.3–12.8% nucleotide divergence from the Indian vaccine strain FMDV O/IND/R2/1975 and clustered under the O/ME-SA/SA-2018 lineage, while one sample (PV288326), showing 14.3% divergence, belonged to the O/ME-SA/Ind2001e lineage. Amino acid analysis revealed 10–11 variations, including four common mutations. Antigenic sites remained conserved, except for an I144V substitution, which is common in all Indian serotype O isolates reported earlier. Conclusion These findings underscore the need for targeted interventions, such as enhanced biosecurity at livestock fairs and markets, restrictions on animal movement from endemic to disease-free zones, wildlife surveillance, and the inclusion of the role of less-monitored species, like small ruminants and wildlife in virus maintenance. Future studies should include antigenic relationship (r-value) tests to assess vaccine efficacy against circulating strains

Systematic follow-up investigation of NSP seroreactors and in-contact cattle and buffaloes for foot-and-mouth disease virus using probang sampling

Abstract Background Foot-and-mouth disease (FMD) is a highly contagious and economically devastating disease of cloven-hoofed animals. In India, the FMD Control Program has been ongoing for the last two decades. A 3AB3 nonstructural protein (NSP)-based indirect ELISA test is used for population serosurveys to differentiate between infected and vaccinated animals (DIVA). In the present study, a systematic follow-up investigation of the NSP seroreactors and in-contact bovines was carried out from rural cohorts as well as an organized farm in Haryana, India to identify the carrier or neoteric animals. No FMD outbreak was reported from Haryana, a Northern state of India in 2022 and NSP reactivity has also consistently been under 10% for the last five years (2018–2022). Results Bovines from ten villages of district Hisar, Haryana, demonstrated 5.3% (20/377) [cattle (11.3%; 12/106) and buffaloes (3.0%; 8/271)] FMDV 3AB3 NSP reactivity. Out of those 20 NSP reactors, nine months later, two buffaloes were randomly screened. Both were found negative for NSP reactivity as well as for FMDV in oropharyngeal fluid (OPF) by reverse transcription-multiplex polymerase chain reaction (RT-mPCR) using 1D/2B gene-specific primers. Further screening was done in a herd of regularly vaccinated cattle (n = 11) of an organized farm with no history of FMD outbreaks for more than a decade. All the susceptible animals were vaccinated with FMD + Haemorrhagic septicemia + Black Quarter combined oil adjuvanted vaccine. An NSP reactivity of 36.7% (4/11) in cattle calves 2–4 months after vaccination indicated either the exposure of animals to FMDV or the presence of residual NSPs in the vaccine. None of the OPF samples collected twice from these cattle at intervals of 36–44 days were found to be positive for FMDV with RT-mPCR. The observed NSP seropositivity could be linked to either false positive reactions or evidence of past exposure and virus elimination during OPF sampling. Nearly all animals exhibited protective antibody titers (≥ log10 1.65) against the structural proteins of FMDV serotypes O, A, and Asia-1 by Solid Phase Competitive ELISA (SPCE) indicating the effectiveness of vaccination. Conclusion The present study provided a preliminary follow-up investigation to assess the status of NSP seroreactors to establish the circulation of FMDV in the animal population, if any, so that the effectiveness of the ongoing vaccination program could be assessed and potential disease-free zones could be identified