ORIGINAL PAPER Role of anionic charges of osmoregulated periplasmic glucans of Salmonella enterica serovar Typhimurium SL1344 in mice virulence Arvind A. Bhagwat • Porteen Kannan • Yi Ning Leow • Mahesh Dharne • Allen Smith Received: 21 September 2011 / Revised: 16 December 2011 / Accepted: 4 January 2012 / Published online: 26 January 2012 Ó Springer-Verlag (Outside the USA) 2012 Abstract opgB gene of Salmonella enterica serovar Typhimurium was identified earlier in a genome-wide screen for mice virulence (Valentine et al. in Infect Immun 66:3378–3383, 1998). Although mutation in opgB resulted in avirulent Salmonella strain, how this gene contributes to pathogenesis remains unclear. Based on DNA homology, opgB is predicted to be responsible for adding phospho- glycerate residues to osmoregulated periplasmic glucans (OPGs) giving them anionic characteristics. In Escherichia coli, yet another gene, opgC, is also reported to contribute to anionic characteristics of OPGs by adding succinic acid residues. We constructed opgB, opgC, and opgBC double mutants of S. enterica serovar Typhimurium strain SL1344. As predicted opgBC mutant synthesized neutral OPGs that were devoid of any anionic substituents. However, opgB, opgC, and opgBC mutations had no significant impact on mice virulence as well as on competitive organ coloniza- tion. In low osmotic conditions, opgB, opgC, and opgBC mutants exhibited delay in growth initiation in the presence of sodium deoxycholate. Anionic substituents of OPGs from Salmonella although appear to be needed to overcome resistance of deoxycholate in hypoosmotic growth media, no evidence was found for their role in mice virulence. Keywords Periplasmic glucans Á Low osmolarity Á Bile salt sensitivity Á Salmonellosis Á Food microbiology Á Microbial food safety Introduction Relatively little is known about how food borne enteric pathogens survive in nutrient deprived and low-osmolarity environments, such as irrigation and vegetable wash waters (Bhagwat 2006). The optimal growth of cells requires that the cytoplasm contains essential constituents with total osmolarity over 300 mosmol l -1 , while vegetable wash waters and irrigation waters osmolarity can be in the range of 80–40 mosmol l -1 (Kennedy 1996). In Escherichia coli, when cells are grown in media with low osmolarity, it has been proposed that the swelling and rupturing of cyto- plasmic membrane are prevented by the periplasmic anionic short glucose chains, termed ‘‘osmoregulated periplasmic glucans’’ (OPGs) (Kennedy 1996; Bohin 2000). OPGs are also thought to play an important role in host-microbe interactions involving specific plant and animal hosts, but the mechanisms are poorly understood (Bohin and Lacroix 2007). We recently demonstrated that OPGs of S. enterica serovar Typhimurium SL1344 are needed for optimal mice virulence and competitive organ colonization (Bhagwat et al. 2009; Liu et al. 2009). Interestingly, gene responsible for phosphoglycerate substitutions on OPGs, opgB, was identified in a genome- Communicated by Erko Stackebrandt. A. A. Bhagwat (&) Á P. Kannan Á Y. N. Leow Á M. Dharne Environmental Microbial and Food Safety Laboratory, Henry A. Wallace Beltsville Agricultural Research Center, Agricultural Research Service, USDA, 10300 Baltimore Avenue, Bldg. 173, BARC-E, Beltsville, MD 20705-2350, USA e-mail: arvind.bhagwat@ars.usda.gov Present Address: Y. N. Leow Ngee Ann Polytechnic, Singapore 599489, Singapore A. Smith Diet Genomics and Immunology Laboratory, Henry A. Wallace Beltsville Agricultural Research Center, Agricultural Research Service, USDA, 10300 Baltimore Avenue, Bldg. 173, BARC-E, Beltsville, MD 20705-2350, USA 123 Arch Microbiol (2012) 194:541–548 DOI 10.1007/s00203-012-0791-4