Running Title: Capsular transformation in group B streptococci

2 The polysaccharide capsule is a major antigenic factor in Streptococcus agalactiae (Lancefield group B streptococcus – GBS). Previous observations suggest that exchange of capsular loci is likely to occur rather frequently in GBS, even though GBS is not known to be naturally transformable. 5 We aimed at identifying and characterizing putative capsular switching events, by means of a combination of phenotypic and genotypic methods, including pulsed-field gel electrophoretic profiling, multilocus sequence typing and surface protein and pili gene profiling. We show that capsular switching by horizontal gene transfer is not as frequent as previously suggested. Serotyping errors may be the main reason behind 10 the overestimation of capsule switching, since phenotypic techniques are prone to errors of interpretation. The identified putative capsular transformants involved the acquisition of the entire capsular locus and were not restricted to the serotype-specific central genes, the previously suggested main mechanism underlying capsular switching. Our data, while questioning the frequency of capsular switching, provide clear evidence for 15 in vivo capsular transformation in S. agalactiae which may be of critical importance in planning future vaccination strategies against this pathogen.