Brucella suis urease encoded by ure 1 but not ure 2 is necessary for intestinal infection of BALB/c mice

Background In prokaryotes, the ureases are multi-subunit, nickel-containing enzymes that catalyze the hydrolysis of urea to carbon dioxide and ammonia. The Brucella genomes contain two urease operons designated as ure 1 and ure 2. We investigated the role of the two Brucella suis urease operons on the infection, intracellular persistence, growth, and resistance to low-pH killing. Results The deduced amino acid sequence of urease-α subunits of operons-1 and -2 exhibited substantial identity with the structural ureases of alpha- and beta-proteobacteria, Gram-positive and Gram-negative bacteria, fungi, and higher plants. Four ure deficient strains were generated by deleting one or more of the genes encoding urease subunits of B. suis strain 1330 by allelic exchange: strain 1330Δ ure 1K (generated by deleting ureD and ureA in ure 1 operon), strain 1330Δ ure 2K ( ureB and ureC in ure 2 operon), strain 1330Δ ure 2C ( ureA , ureB , and ureC in ure 2 operon), and strain 1330Δ ure 1KΔ ure 2C ( ureD and ureA in ure 1 operon and ureA , ureB , and ureC in ure 2 operon). When grown in urease test broth, strains 1330, 1330Δ ure 2K and 1330Δ ure 2C displayed maximal urease enzyme activity within 24 hours, whereas, strains 1330Δ ure 1K and 1330Δ ure 1KΔ ure 2C exhibited zero urease activity even 96 h after inoculation. Strains 1330Δ ure 1K and 1330Δ ure 1KΔ ure 2C exhibited slower growth rates in tryptic soy broth relative to the wild type strain 1330. When the BALB/c mice were infected intraperitoneally with the strains, six weeks after inoculation, the splenic recovery of the ure deficient strains did not differ from the wild type. In contrast, when the mice were inoculated by gavage, one week after inoculation, strain 1330Δ ure 1KΔ ure 2C was cleared from livers and spleens while the wild type strain 1330 was still present. All B. suis strains were killed when they were incubated in-vitro at pH 2.0. When the strains were incubated at pH 2.0 supplemented with 10 mM urea, strain 1330Δ ure 1K was completely killed, ...