Adding vitamin C as a nutritional supplement while treating drug-sensitive tuberculosis patients with first-line TB drugs will boost the efficiency of treatment, a study by Indian researchers shows. The increase in efficiency is not because vitamin C has antibacterial activity, as was suggested by a few researchers in 2013 from in vitro studies, but by doing the complete opposite — inducing dormancy in TB bacteria.
A team led by Prof. Jaya Sivaswami Tyagi from the Department of Biotechnology at AIIMS, New Delhi had first proposed the dormancy-inducing ability of vitamin C in TB bacilli in 2010 and has now reconfirmed it in a comprehensive study published in the journal Redox Biology.
The team found that vitamin C imposes multiple stresses on TB bacteria such as hypoxia, acid stress (where the pH is reduced to around 5.5), oxidative stress (through the generation of H2O2 and reactive oxygen species), reductive stress (due to cessation of aerobic respiration) and metabolic stress. “As a result of these stresses, there is slowing down of metabolism leading to dormancy and further progression to viable but non-culturable (VBNC) state,” says Prof. Tyagi. “Together, these stresses remarkably resemble the host environment that the bacteria would face.”
In the lab, TB bacteria already exposed to vitamin C displayed resistance to two first-line drugs — isoniazid and rifampicin — as it progressed to a dormant state. Unlike these two drugs, pyrazinamide drug is capable of killing TB bacteria even in a dormant state. “The addition of vitamin C increased the population of dormant bacteria and this led to an eight-fold increase in pyrazinamide’s ability to kill the bacteria. There was also a four-fold decrease in the minimum concentration (MIC) of pyrazinamide required to kill the bacteria even in an infection model,” says Dr. Kriti Sikri from the Department of Biotechnology, AIIMS and first author of the paper.
In the presence of pyrazinamide alone nearly 90% of the bacteria survived but when combined with vitamin C, the survival rate in vitro dropped sharply to less than 3%. “Vitamin C induces dormancy and enhances the population of slowly growing bacteria or bacteria that are not growing. And pyrazinamide drug targets these bacteria bringing about sharp reduction in the survival rate,” explains Prof. Tyagi. “The effect of vitamin C combined with TB drugs was reproduced in an intracellular model, which is akin to human infection. So, our findings acquire clinical relevance. When used along with other first-line drugs, vitamin C has the potential to shorten the treatment time.”
Screening novel drugs
Besides improving the efficacy of existing TB regimen, vitamin C can help in producing subclasses of bacteria to test new drugs. For instance, due to hypoxia, the metabolism gets slowed down leading to reductive stress. Lipids are formed as a compensatory mechanism and the breakdown of lipids produces energy. The AIIMS team used novel inhibitors to prevent the breakdown of lipids for energy purpose and this led to the death of TB bacteria.
Similarly, dormancy can be induced by adding vitamin C. Dormant bacteria are able to reduce the effectiveness of isoniazid and rifampicin by pumping them out using efflux pumps. The use of vitamin C can help in producing a population of dormant bacteria which can be used for screening drugs that inhibit efflux pumps, which the researchers were able to do. “We were able to reverse the tolerance and restore the bacterial sensitivity to these drugs by treating the bacteria with either of two efflux pump inhibitors — verapamil and piperine,” Prof. Tyagi says.
Likewise, vitamin C can be used for producing viable but non-culturable (VBNC) TB bacteria. Though VBNC bacteria are present in sputum samples they cannot be cultured and hence never identified leading to misdiagnosis. The VBNC bacteria can always reverse and cause disease. “Unlike reports that 100 days are required to produce VBNC bacteria, our model can generate them in just eight days, which can be used for screening drugs,” Prof. Tyagi says.
When used with first-line drugs, vitamin C can shorten the treatment time