Molecular Cloning and Characterization of cgt , the Brucella abortus Cyclic β-1,2-Glucan Transporter Gene, and Its Role in Virulence

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Abstract
The animal pathogen Brucella abortus contains a gene cgt , which complemented Sinorhizobium meliloti nodule development ( ndvA ) and Agrobacterium tumefaciens chromosomal virulence ( chvA ) mutants. Complemented strains recovered the presence of anionic cyclic β-1,2-glucan, motility, tumor induction in A. tumefaciens , and nodule occupancy in S. meliloti , all traits strictly associated with the presence of cyclic β-1,2-glucan in the periplasm. Nucleotide sequencing revealed that B. abortus cgt contains a 1,797-bp open reading frame coding for a predicted membrane protein of 599 amino acids (65.9 kDa) that is 58.5 and 59.9% identical to S. meliloti NdvA and A. tumefaciens ChvA, respectively. Additionally, B. abortus cgt , like S. meliloti ndvA and A. tumefaciens chvA possesses ATP-binding motifs and the ABC signature domain features of a typical ABC transporter. Characterization of Cgt was carried out by the construction of null mutants in B. abortus 2308 and S19 backgrounds. Both mutants do not transport cyclic β-1,2-glucan to the periplasm, as shown by the absence of anionic cyclic glucan, and they display reduced virulence in mice and defective intracellular multiplication in HeLa cells. These results suggest that cyclic β-1,2-glucan must be transported into the periplasmatic space to exert its action as a virulence factor.