We have identified apuA in Streptococcus suis, which encodes a bifunctional amylopullulanase with conserved a-amylase and pullulanase substrate-binding domains and catalytic motifs. ApuA exhibited properties typical of a Gram-positive surface protein, with a putative signal sequence and LPKTGE cell-wall-anchoring motif. A recombinant protein containing the predicted N-terminal a-amylase domain of ApuA was shown to have a-(1,4) glycosidic activity. Additionally, an apuA mutant of S. suis lacked the pullulanase a-(1,6) glycosidic activity detected in a cell-surface protein extract of wild-type S. suis. ApuA was required for normal growth in complex medium containing pullulan as the major carbon source, suggesting that this enzyme plays a role in nutrient acquisition in vivo via the degradation of glycogen and food-derived starch in the nasopharyngeal and oral cavities. ApuA was shown to promote adhesion to porcine epithelium and mucus in vitro, highlighting a link between carbohydrate utilization and the ability of S. suis to colonize and infect the host.
INTRODUCTIONStreptococcus suis is a major porcine pathogen of significant commercial importance worldwide. In suckling and weaning pigs, it is the principal cause of acute meningitis, but can infect other organs leading to arthritis, serositis, endocarditis, otitis media and bronchopneumonia (Beineke et al., 2008;Madsen et al., 2002). Healthy pigs asymptomatically colonized with S. suis form a reservoir for this disease and play a major role in its epidemiology (Arends et al., 1984). To date, 33 different capsule serotypes of S. suis have been identified, but serotype 2 is most commonly associated with disease worldwide Lun et al., 2007). Serotype 2 strains were also associated with recent large outbreaks of severe human infections in China and elsewhere in Asia (Mai et al., 2008;Tang et al., 2006;Wertheim et al., 2009). The recently obtained genome sequences of two virulent Chinese S. suis serotype 2 strains (98HAH12 and 05ZYH33) and P1/7, the European reference strain (http://www.sanger.ac.uk/Projects/S_suis/), led to the identification of a large number of potential surface and secreted proteins that might play a role in virulence, including a number of putative carbohydratedegrading enzymes (Baums & Valentin-Weigand, 2009). Genes that encode carbohydrate-degrading enzymes are common in the genomes of other streptococcal pathogens and play a role in nutrient acquisition for growth and colonization on mucosal surfaces (Rollenhagen & Bumann, 2006;Shelburne et al., 2006Shelburne et al., , 2008a. Dietary sources of highly polymerized a-glycans such as starch and glycogen are abundant in the human colon (Levitt et al., 1987) and oropharynx (Mormann & Muhlemann, 1981;Shelburne et al., 2005 Shelburne et al., , 2007Virtaneva et al., 2005), as well as the epithelium of the vagina and lung (Gourlay et al., 2009;Gregoire et al., 1971; Santi et al., 2008;van Bueren et al., 2007). Degradation of starch and glycogen proceeds in most organisms via the action of amylases a...