Abstract
Six new Brucella species isolated from humans, wild animals and environmental sources, were added to the six classical Brucella species potentiating an additional threat for humans. Other atypical Brucella strains isolated from wild rodents, frogs and fishes, will likely be proposed as new species in the future. This work aims to implement a core-genome-based analytical pipeline, PORIFERA, for prediction of new markers to rapid identification of emerging Brucella species. The comparison of core-genome at greater resolution of closely related Brucella spp. may provide information on specific genetic markers, such as single-nucleotide polymorphism (SNP). Twenty-three Brucella genomes (10 B. melitensis, seven B. suis and six B. abortus) were sequenced using Illumina technology. Genome assemblies were performed using de novo assembler SPAdes. The analysis involved the 23 newly sequenced genomes and 25 Brucella spp. complete genomes publicly available from eight of the 12 recognized species B. suis, B. abortus, B.canis, B. ovis, B. microti, B. pinnipedialis and B. ceti. Two Brucella sp. strains isolated from amphibians (09RB8910; 09RB8471) were used as outgroup. The alignment of core-genome was performed using Parsnp and the evolutionary history was inferred using Neighbor-Joining and Maximum Likelihood methods available in the Harvest suite (vs 1.1.2). Functional annotation had been accomplished to each SNP, as well as exclusion of intragenic SNPs in order to identify novel discriminatory and informative biomarkers. The evolutionary history was inferred from a total of 256 667 putative SNPs shared among the 46 more closely related genomes and the genomes of the amphibians isolates 09RB8910 and 09RB8471. From these, a total of 31034 SNPs were significantly associated with B.canis (n=2127), B. ovis (n=2018), B. suis (n=1782), B. abortus (n=1466), B. suis (n=1365), B. ceti (n=1463), B. pinnipedialis (n=891) and B. microti (n=786). These data will be further used for development of novel molecular methods to identify, genotype or direct assignment of Brucella species.
