Bovine serum albumin as a resuscitation promoting factor for viable but non-culturable Mycobacterium tuberculosis via the activation of protein kinase A-dependent cellular processes

We investigated the mechanisms underlying the fetal bovine serum (FBS)-induced reactivation of the viable but non-culturable (VBNC) state of Mycobacterium tuberculosis (Mtb) using the H37Rv strain. We found that bovine serum albumin (BSA), a major component of FBS, could reactivate the VBNC state of Mtb cells without other reactivation-promoting agents. Human serum albumin also had a restorative effect similar to BSA, but mouse serum albumin and egg-white albumin (ovalbumin) did not, suggesting that a common protein structure between bovine and human serum albumin is essential for reactivation. In addition, antioxidative agents, such as N-acetyl-L-cysteine, showed no restorative effect, suggesting that the restoration of culturability might not be due to the antioxidative property of BSA. BSA-mediated reactivation is inhibited by H89 and staurosporine. These inhibitors are known to inhibit multiple protein kinases, including serine/threonine protein kinases, in Mtb, suggesting the involvement of mycobacterial protein kinases in cell division during reactivation. These findings provide new insights into the mechanisms of Mtb reactivation, and may contribute to the development of novel strategies for the treatment of latent tuberculosis. ImportanceReactivation of dormant, including viable but non-culturable (VBNC) Mycobacterium tuberculosis (Mtb) cells, represents a critical step in the progression from latent infection to active tuberculosis. However, the molecular mechanisms that trigger this transition are not yet fully understood. We herein report that bovine serum albumin (BSA), a major component of fetal bovine serum, can restore the culturability of VBNC Mtb cells. Human serum albumin showed a similar effect, whereas mouse and egg-white albumins did not, suggesting that the specific structural features shared between bovine and human albumins are essential for reactivation. The lack of reactivation by the antioxidant N-acetyl-L-cysteine indicates that the mechanism is not related to redox balance. Inhibition of BSA-induced reactivation by H89 and staurosporine implicates mycobacterial serine/threonine protein kinases in this process. These findings highlight the previously unrecognized role of albumin-kinase signaling in Mtb reactivation and suggest new molecular targets for preventing tuberculosis relapse.