To date, there are five subtypes of the sodium-dependent excitatory amino acid transporters that differ in their regional and cellular expression and ability to transport glutamate. Some systems appear to be functionally conserved in the pneumococcus, such as the well-characterized neuraminidase NanA or the Sus and Scr sucrose utilization systems, although there can be sequence variation between strains. There are also carbohydrate utilisation systems encoded on accessory regions that are not uniformly distributed amongst strains. McKessar and Hakenbeck have previously described an accessory region containing a cellobiose phosphotransferase system, whose components were also identified during a signature-tagged mutagenesis study using an otitis media model. In this study we examined an alternative accessory region harbouring a putative cellobiose phosphotransferase system, which has been previously described as being absent in D39 and TIGR4 and hypothesised to be involved in virulence. Despite its absence in D39 and TIGR4, this PTS was present in four distinct serotype 3 isolates, which were highly virulent in a mouse sepsis model, but absent in the significantly less virulent serotype 3 strain WU2. However, as with the cel locus, we found that this system is present in a wide range of pneumococcal isolates belonging to diverse serotypes, including unrelated avirulent isolates of serogroup 11. In contrast, sulfatases are highly conserved across eukaryotes and prokaryotes and are believed to be used by prokaryotes to hydrolyse sulphate ester groups from proteoglycans, glycosaminoglycans and choline sulphate. They are normally associated with a sulfatase-Telbivudine modifying factor, which is required to catalyse the post-translational activation of the sulfatase, an unusual feature for prokaryotic proteins. There are many other pneumococcal sugar transporters also associated with hydrolases, but this particular system is interesting in that some strains, including G54 and serotype 3 isolates of ST458, have a deletion removing the sulfatase, as well as parts of the sulfatase modifying factor and the last gene of the PTS operon. Due to the prevalence of this island and its interesting annotation, in this study we have constructed mutants of the island in two distinct S. pneumoniae genetic backgrounds and investigated the impact on phenotype in vivo. The orientation of the genes of the accessory region necessitated the inclusion of a promoter to allow expression of the erythromycin resistance gene, which is why the cassette was amplified from the mutant D39ABDe to include the capsule locus promoter. Following confirmation of the mutation by sequencing, opaque variants were selected for all experiments, except the biofilm assay and colonisation competition study for which transparent pneumococci were preferred. Initially, the mutant was examined for alterations in both planktonic and biofilm growth, as well as sugar fermentation. The ability of WCH206 DIsland to colonize the nasopharynx of mice was also examined. Mice were inoculated intranasally without anaesthesia and the nasopharynx was washed and nasopharyngeal tissue removed and homogenized post-inoculation. Data from the nasopharyngeal wash and homogenate were combined. The mutant was found to be significantly attenuated compared to wild-type WCH206 strain on day 1; log10CI values were less than 0 on days 2 and 4 as well, but this did not reach statistical Licochalcone-A significance. As serogroup 11 strains had previously been found to colonize at higher numbers on days 1 and 2 post-inoculation than serotype 3 strains in this model, we also constructed an island deletion replacement mutant in the serotype 11A carriage isolate Menzies5 using the same method.