Analysis of growth of the parental strain, Ev1 and their respective cg2937 disruptions in CGXII Neu5Ac

medium, revealed that disruption of cg2937 results in a complete loss of growth (Fig. 3a and b). The same phenotype was observed on solid media (Fig. 3d). We examined [14C]-Neu5Ac uptake using Ev1 and Ev1 cg2937::pDRIVE where uptake was also completely abolished in the strain disrupted in cg2937 (Fig. 3c). Hence, we conclude that the cg2937-40 genes encode the sole sialic acid transporter in C. glutamicum. Given the clear demonstration that the soil bacterium C. glutamicum has the ability to grow on sialic acid, we examined the distribution of the sialic acid transport and utilization genes within the genus Corynebacterium (Fig. 4). It is clear that the sialic

acid genes are not unique to C. glutamicum, but are present in a number of other members of the genus Corynebacterium find more particularly in organisms that cause diseases in human and animals where genome JQ1 sequences are available such as Corynebacterium diphtheriae (Cerdeno-Tarraga et al., 2003), Corynebacterium ulcerans (Trost et al., 2011) and Corynebacterium pseudotuberculosis (Trost et al., 2010). In every case, they have a SatABCD-like sialic acid transporter and the full set of genes needed for catabolism, namely nanA, nanE, nanK, nagA and nagB. While C. glutamicum, C. diphtheriae, C. pseudotuberculosis and C. ulcerans all encode a sialidase on their genome, the predicted sialidase in C. glutamicum (cg2935) is the only one encoded within the main nan-cluster and is not a clear orthologue of the nanH sialidase seen in the other three organisms (marked as nanH in Fig. 4). Sialic acid utilization has been well studied in a range of pathogens, and in this work, we demonstrate clearly that the soil bacterium C. glutamicum can transport and utilize Neu5Ac as a sole carbon source. Examination of the genome reveals what appears to be

a fairly canonical sialic acid cluster containing a full set of genes including an ABC transporter that we have demonstrated is essential for uptake (Fig. 4). It is not clear why the presence of sialic acid utilization genes was not recognized in a previous study (Almagro-Moreno & Boyd, 2009), looking at the distribution of the nanAEK genes in bacteria. The only Dipeptidyl peptidase member of the genus Corynebacterium, where sialic acid biology has been previously studied, is in C. diphtheriae. A sialidase was first isolated from this pathogen in 1963 (Warren & Spearing, 1963; Moriyama & Barksdale, 1967) and, remarkably, NanA activity was also identified shortly afterwards (Arden et al., 1972). Interestingly, the same study demonstrated that both sialidase and NanA (N-acetylneuraminate lyase) activities were also observed in C. ulcerans and Corynebacterium ovis (now C. pseudotuberculosis; Arden et al., 1972), which agrees with the presence of both nanH and nanA genes in all of these sequenced genomes (Fig. 4).