Macrophage and mycobacterium
Macrophage phagocytosing mycobacteria

Sometimes the simple, obvious answer is right, and sometimes it’s completely backwards.

Tuberculosis was a terrifying, ubiquitous killer in the 19th century, but is relatively rare today (at least, in developed countries). The reason for the drop in Tb deaths isn’t entirely clear; it started with social factors probably including accidental or deliberate isolation of Tb patients, antibiotic treatment also knocked the disease back, and in some areas the vaccine (known as BCG) made a difference as well.

BCG is one of the oldest vaccines still in wide use; it was developed in the 1920s when a strain of Mycobacterium bovis (tuberculosis of cattle, contagious to humans) spontaneously lost virulence in culture. This avirulent strain of the bacterium was sent around the world and cultured independently, resulting in many distinct vaccine strains in different places and times. These strains are not only distinct genetically, but also phenotypically — they look different in culture, or grow differently, or whatever.

Over time, the vaccine has changed functionally, as well. Very early on the vaccine abruptly became even less virulent. More gradually, it seems that BCG has also become less effective; it’s no longer is able to protect against pulmonary Tb (although it’s still protective against other forms of the disease). Why is this?

At first glance this seems unsurprising. The bacterium has been grown in culture — outside of any animal host — for nearly 100 years. It’s had no selection to maintain its ability to grow in animals, or to avoid their immune responses, so of course it’s going to lose its ability to grow in animals.

But a recent paper1 suggests that exactly the opposite happened. Whether randomly, or because of some unexpected type of selection, the BCG strain has actually amplified an immune evasion function. This modern variant of the vaccine strain isn’t simply passively failing to induce an immune response; it’s actively suppressing the immune response.

Specifically, the authors argue that normal (wild, virulent) Mycobacterium secretes antioxidants as an immune evasion mechanism; that modern BCG also secretes lots of antioxidants; and that this is related to genomic duplications in some BCG strains:

Some BCG daughter strains exhibit genomic duplication of sigH, trxC (thioredoxin), trxB2 (thioredoxin reductase), whiB1, whiB7, and lpdA (Rv3303c) as well as increased expression of genes encoding other antioxidants including SodA, thiol peroxidase, alkylhydroperoxidases C and D, and other members of the whiB family of thioredoxin-like protein disulfide reductases.1

Further reading
Tb family trees
Conspicuous consumption
Life & Death, pre-vaccination

MycobacteriaIn other words, the long-term culture of BCG has yielded variants that are less immunogenic, because they are more actively suppressing the immune response. If their reasoning is correct, then reducing the antioxidant secretion from BCG should increase its immunogenicity. They took a BCG strain and deleted the duplicated antioxidant gene sigH (as well as the overexpressed SodA), and sure enough, the deleted version was more immunogenic and more protective in mice. “By reducing antioxidant activity and secretion in BCG to yield 3dBCG, we unmasked immune responses during vaccination with 3dBCG that were suppressed by the parent BCG vaccine.1

As a possible explanation, they note that their deletion variant also grows more slowly in culture than the “wild-type” BCG, and especially under certain culture conditions, and that this has led, coincidentally, to the reduced immunogenicity:

The practice of growing BCG aerobically with detergents to prevent clumping may have increased oxidant stress to cell wall structures and selected for increased antioxidant production. Then with each transfer the bacilli making more antioxidants represented a slightly greater proportion of the culture until they became dominant. In vivo, these mutations caused the vaccine to become less potent in activating host immunity. In effect, we believe that as BCG evolved it yielded daughter strains with an increased capacity for suppressing host immune responses. 1

If this turns out to be generally true, then there’s a relatively straightforward handle for converting BCG back into a more effective, and safer, vaccine; whereas if the reduced immunogenicity was because of over-attenuation, it’s not so simple — you’d be trying to make a vaccine more virulent, which is a tricky tightrope to walk.

Incidentally, I frequently complain about the terrible, terrible quality of press releases about scientific advances¬† (and therefore the terrible quality of much “science reporting”, which is basically regurgitating the terrible press releases) so I want to give props to the person at Vanderbilt University Medical Center who put together the release for this paper — it’s a clear, simple, interesting, and as far as I can tell accurate account of the finding, background, and observation.¬† It can be done well — I wish it was done this well more often.

  1. Sadagopal, S., Braunstein, M., Hager, C., Wei, J., Daniel, A., Bochan, M., Crozier, I., Smith, N., Gates, H., Barnett, L., Van Kaer, L., Price, J., Blackwell, T., Kalams, S., & Kernodle, D. (2009). Reducing the Activity and Secretion of Microbial Antioxidants Enhances the Immunogenicity of BCG PLoS ONE, 4 (5) DOI: 10.1371/journal.pone.0005531[][][][]