First isolation and characterization of Brucella microti from wild boar

Abstract

Background: Brucella microti was first isolated from common vole ( Microtus arvalis ) in the Czech Republic in Central Europe in 2007. As B. microti is the only Brucella species known to live in soil, its distribution, ecology, zoonotic potential, and genomic organization is of particular interest. The present paper is the first to report the isolation of B. microti from a wild boar ( Sus scrofa ), which is also the first isolation of this bacterial species in Hungary. Results: The B. microti isolate was cultured, after enrichment in Brucella -selective broth, from the submandibular lymph node of a female wild boar that was taken by hunters in Hungary near the Austrian border in September 2014. Histological and immunohistological examinations of the lymph node sections with B. abortus- , B. suis- and B. canis -specific sera gave negative results. The isolate did not require CO 2 for growth, was oxidase, catalase, and urease positive, H 2 S negative, grew well in the presence of 20 μ g/ml basic fuchsin and thionin, and had brownish pigmentation after three days of incubation. It gave strong positive agglutination with anti-A and anti-M but had a negative reaction with anti-R monospecific sera. The API 20 NE test identified it as Ochrobactrum anthropi with 99.9 % identity, and it showed B. microti -specific banding pattern in the Bruce- and Suis-ladder multiplex PCR systems. Whole genome re-sequencing id entified 30 SNPs in orthologous loci when compared to the B. microti reference genome available in GenBank, and the ML VA analysis yielded a unique profile. Conclusions: Given that the female wild boar did not develop any clinical disease, we hypothesize that this host species only harboured the bacterium, serving as a possible reservoir capable of maintaining and spreading this pathogen. The infectious source could have been either a rodent, a carcass that had been eaten or infection occurred via the boar rooting in soil. The low number of discovered SNPs suggests an unexpectedly high level of genetic homogeneity in this Brucella species. Keywords: Biochemistry, Brucella microti , Immunohistochemistry, MLVA, Morphology, Wild boar, Whole genome sequencing, Hungary