Application: This study could be useful for the development of microbiome assay for Bovine Genital Campylobacteriosis in cattle.
Introduction: Bovine reproductive tract harbours multiple species of organisms that potentially play the key role in reproductive tract health, preventing pathogenic microorganisms’ growth as well as maintaining fertility and pregnancy. Campylobacter fetus subspecies venerealis (CFV) is a pathogenic bacterium transmitting to cows during breeding by asymptomatic carrier bulls causing Bovine Genital Campy-lobacteriosis (BGC). The objective of the present study is to characterise the bovine reproductive tract microbiome and potentially identify biomarker(s) for BGC immunity.
Materials and Methods: A total of 12 cycling heifers were selected, 6 were vaccinated once using a commercially available vaccine against BGC and six were left unvaccinated (control). Following oestrus synchronization (15 days post-vaccination) using two doses of PGF (Prostaglandin F2 alpha) 9 days apart, heifers were intravaginally challenged with a freshly prepared inoculum of pure live CFV bacteria (27x108 colony-forming units/heifer). Vaginal mucus samples were collected in phosphate buffer solution (PBS) at different timepoints, 12 times post-vaccination at 2-days intervals, and 8 times weekly post-challenge. The sample dates were aligned with the expected oestrus cycle phases and confirmed by ultrasonography. DNA was extracted from mucus sample and tested using qPCR and submitted for 16S rDNA amplicon Illumina sequencing (V5-V8 region). QIIME2 (v2020.11.1) bioinformatics pipeline was used for processing, analysing, and visualizing microbiome data.
Results: The mean abundance of vaginal microbiome population was associated with vaccination, oestrus, and challenge. Heifers were CFV qPCR negative before challenge and the percentage of heifers that were qPCR positive at days 2, 7, 14 and 28 after the challenge was 66.7%, 50%, 50% and 16.7% for nonvaccinated heifers and 0%, 33.33%, 66.67% and 16.67% for vaccinated heifers respectively. Vaccinated heifers eliminated the infection sooner than the nonvaccinated heifers (4 vs. 7 weeks). In both treatments, Firmicutes were more abundant than Bacteroides, Proteobacteria and Actinobacteria. Streptococcus spp., was higher in abundance and increased during oestrus than dioestrus in both treatment groups. In nonvaccinated heifers, Campylobacter spp. appeared 2 days following challenge compared to 14 days in the vaccinated heifers. In addition, Arthrobacter sp. (unassigned) were more abundant in oestrus compared to dioestrus in both treatments. Alternatively, Proteobacteria seems to increase after challenge, particularly Acinetobacter.
Conclusions: The microbiome profile of Proteobacteria and Actinobacteria changed depending on oestrus and dioestrus before and after challenge with CFV in vaccinated and nonvaccinated heifers.