The chemical and antigenic properties of the cell-surface lipopolysaccharides (LPSs) and capsular polysaccharides (CPSs) of seven representative strains of Actinobacillus suis from healthy and diseased pigs were investigated. Four strains produced a linear (1->6)-β-d-glucan homopolymer, β-d-Glcp-(1-[->6)-β-d-Glcp-(1-] n ->, as a LPS-O-chain (O1) and as a CPS (K1). Polyclonal antisera prepared against a (1->6)-β-d-glucan-containing strain showed a positive reaction against both LPSs and CPSs derived from the above strains (designated serotype O1/K1). One strain carried the (1->6)-β-d-glucan solely as a LPS-O-chain (serotype O1) and two strains did not express the (1->6)-β-d-glucan, but, instead, produced a different O-chain (designated serotype O2); these three strains expressed their own characteristic CPSs. (1->6)-β-d-Glucan structures are common cell wall components of yeast, fungi and lichens, but, to our knowledge, this is the first time a (1->6)-β-d-glucan has been described in a prokaryotic organism. Conformational and nuclear magnetic resonance analyses showed that the β-d-Glcp-(1->6)-β-d-Glcp linkage was flexible and two distinct glycosidic conformers are described. Cross-reactive antibodies to the A. suis (1->6)-β-d-glucan could be detected in sera from a variety of species and in sera from specific pathogen free pigs. This cross-reactivity may arise from immuno-stimulation of organisms present in the surrounding environment that contain (1->6)-β-d-glucan, which may also explain the high incidence of false positive results in previous serological tests for A. suis. In addition, these (1->6)-β-d-glucan background antibodies may be protective against A. suis infection. The characterization herein of (1->6)-β-d-glucan is the foundation for the development of a serotyping system for A. suis.