P023

Cobalt is an essential trace metal, incorporated in important metalloproteins, e.g. corronoid proteins. Best-known representatives of this group are cobalamins, such as vitamin B12. 80-90% of all bacteria contain vitamin B12 dependent enzymes, involved in several central metabolic processes and in host-pathogen interaction. Despite this essentiality, only 25-30% are able to produce vitamin B12 de novo, including many mycobacteria species. For the production of vitamin B12, cobalt uptake is essential. However, knowledge on cobalt uptake and regulation in Non-tuberculous mycobacteria (NTM) such as the opportunistic pathogen M. abscessus and M. avium is very limited.

Here we analyzed cobalt homeostasis of M. smegmatis (MSMEGwt). A regulator of the ArsR/SmtB family (msmeg_2606) associated with a putative metal transporter (msmeg_2607-2610) and genes of vitamin B12 synthesis (msmeg_2616-2618) was identified by genome mining. The regulon of msmeg_2606 was determined by transcriptome analyses of a MSMEG∆2606 deletion mutant. 30 genes were >5 fold differentially expressed in the mutant compared to MSMEGwt. Amongst these, expression of msmeg_2607-2610 was higher, indicating a role of MSMEG_2606 in regulation. BLAST and TMHMM analyses suggest that msmeg_2607-2610 encoded proteins are a CbiMNQO cobalt importer of the Energy-Coupling-Factor (ECF) family. In qRT-PCR experiments cbiM (msmeg_2607) expression was induced in MSMEGwt grown in cobalt-deficient medium, supporting the putative role of msmeg_2607-2610 in cobalt metabolism. A MSMEG∆cbiMN mutant will be used in future growth experiments and reporter studies to analyze regulation by MSMEG_2606 and the metals involved in regulation and transport. These findings might help identifying new antibacterial targets for treating NTM infection.