TITLE:
Identification of Growth Promoting Effect of rBCG/BCG Culture Supernatant and Its Potential Applications
AUTHORS:
Tom H. Jin, Tianli Qu, Anant Raina, Eric Tsao
KEYWORDS:
Resuscitation-Promoting Factors (Rpfs); rBCG/BCG; Culture
JOURNAL NAME:
World Journal of Vaccines,
Vol.3 No.2,
May
22,
2013
ABSTRACT: Background: Although BCG is the most widely
administered vaccine in the world, there have never been as many cases of TB as
there are now. Globally, more than 8.8 million people developed active TB and
1.4 million—many
of them—died in 2010. It
is estimated that half of pulmonary TB cases arise from latent Mtb infection, making the study of
latency and reactivation of utmost importance. Methods: Widely administered BCG vaccines and a gene modified recombinant BCG (rBCG) strain, AERAS-422, were used as models to investigate the
growth promoting function of resuscitation-promoting factors (Rpfs) in different bacilli culture phases.
Different supernatant fractions were prepared by ultrafiltration, and the
promoting function of each fraction containing secreted Rpf(s) was evaluated by
growth curve monitoring and colony counting on 7H10 agar plates. Results: The promoting effect of
culture supernatants was mainly associated with the high molecular weight
fraction (>30 kDa), which stimulated bacterial growth, but did not extend the exponential
phase of stimulated culture. Anti-RpfB antibody showed significant growth
restriction of the tested cultures. When comparing rBCG cultures containing 7H9 medium, the 10 - 30 kDa fraction, or the >30 kDa
fraction, only the >30 kDa fraction was displayed with down-regulation of
the secretion of RpfC, D and E. In colony counting tests, the plates containing
the >30 kDa fraction had total countable colony numbers 2 to 3 fold higher
than the plates with the 10 - 30 kDa fraction,
and colonies appeared one to two weeks earlier than on the regular plates. The
potential applications of the prepared supernatant fractions containing RpfA
and RpfB are discussed, which may include accelerating diagnosis of Mtb infection and future TB vaccine
development.