Detection of Campylobacter fetus in the post-genomics era: A New target for an accurate molecular diagnosis

FARACE, PABLO; MORSELLA, CLAUDIA; PAOLICCHI, FERNANDO; CRAVERO, SILVIO; GIOFFRÉ, ANDREA

Congreso; V Congreso Argentino de Microbiología Agrícola y Ambiental; 2021

Reproductive diseases of livestock represent significant economic losses for the country. Campylobacter fetus is a fastidious microorganism and is considered a zoonotic pathogen. This bacteria is associated with infertility and abortion, a disease known as Bovine Genital Campylobacteriosis (BGC). C. fetus is a Gram-negative bacillus that comprises the mammalian subspecies C. fetus venerealis (Cfv) and C. fetus fetus (Cff). Several lines of evidence have questioned the need for identifying C. fetus subspecies. Additionally, current target genes employed for molecular diagnosis of C. fetus have been tested on a discrete number of samples or their target genes were not studied in depth. Thus, reliable molecular assays are still required to date. The availability of genomic sequences leaves open the possibility for searching and describing new target sequences to design novel and reliable diagnostic tools.One representative whole genome sequence of each Campylobacter species, four representative genomes of C. fetus and also three genomes of closely related genus were selected from Genbank. They were annotated with Prokka and were employed as input to perform a global multiple sequence alignment (MSA) with progressiveMauve. Potential C. fetus-specific regions from the MSA were corroborated by local alignment with BLAST and then were aligned with MUSCLE to identify the conserved regions for designing specific primers (SnapGene). The gene coding for a dihydrodipicolinate synthase family protein was selected. Thirty-four isolates of C. fetus (Cff, n: 22; Cfv, n: 4; Cfvi, n: 8) obtained from bovine clinical samples and four isolates of other Campylobacter species were evaluated by PCR. The presence of a 150 bp product was indicative of C. fetus. The wet-lab assay was positive for all the C. fetus isolates tested. The same result was obtained through in silico analysis (PrimerMap) of 263 publicly available whole genome sequences (GenBank).Herein we performed a study based on whole genome sequences data and global alignment to select a target sequence for an accurate molecular detection of C. fetus. We propose a novel PCR specific for C. fetus supported by genomic data and wet-lab assays.