Mycobacterium tuberculosis (M. tuberculosis), the causative agent of tuberculosis (TB), is a
microbial pathogen which has infected about one third of the world’s population, with about
eight million new cases of TB reported annually of which almost two million are fatal. Those
coinfected with human immunodeficiency virus (HIV) are most vulnerable for development of
severe disease. The disease is acquired through the respiratory route, whereby M. tuberculosis
bacilli overcome the mechanical defences of the upper airways to reach the lungs where they
infect alveolar macrophages. Although considerable progress has been made in identifying
immune mechanisms which confer protection against M. tuberculosis, this has not resulted in
the development of an effective vaccine, underscoring the fact that novel insights into the
immunopathogenesis of M. tuberculosis infection are necessary. In this respect it is noteworthy
that almost nothing is known about the involvement of the major mycobacterial potassium (K+)
transporters in microbial virulence.
M. tuberculosis possesses two major K+-uptake systems, namely the Trk and Kdp systems. The
Trk seems to be functional when the extracellular K+ concentration is high, while the Kdp is an
inducible back-up system. The Trk system consists of two proteins, CeoB and CeoC, which are
encoded by the ceoB and ceoC genes, which have some degree of homology to the TrkA protein
of Escherichia coli. These proteins share the NAD+-binding motif, compatible with proton
motive force as the driver of cation uptake, suggesting that the M. tuberculosis Trk K+
transporter may operate as a K+ and protons (H+) symporter, raising the possibility that it may
antagonize vacuolar acidification, a critical event in the eradication of this intracellular
The possible involvement of the Trk system in the virulence of M. tuberculosis has been
addressed in the current study by investigating the intracellular survival of a trk-gene knockout
mutant of the microbial pathogen with that of the matched wild-type (WT) strain using human monocyte-derived macrophages. In addition, the cytokine profiles of macrophages infected by
both strains have also been investigated.
Macrophages were prepared from isolated human blood monocytes following sequential
differential adherence of CD14+ monocytes. These were matured into large monocytes-derived
macrophages co-expressing CD14+ / CD16+ following a 7 day incubation period. These cells
were then infected with either the WT (H37Rv) or trk-gene knockout strains of M. tuberculosis
at a 1:10 cell: bacteria ratio and intracellular survival as well as cytokine (IL-1b, IL-6, IL8, IL-
10, TNF-a) secretion profiles monitored over a 3 day period using a viable colony-counting
procedure and multiplex bead array technology, respectively.
No significant differences with respect to either intracellular survival or cytokine secretion
profiles were detected following infection of human monocyte-derived macrophages with the
WT or trk-gene knockout strains of M. tuberculosis. Although the observations are compatible
with lack of involvement of the Trk system in the intracellular survival and virulence of M.
tuberculosis, this study may lay groundwork for future studies which simultaneously encompass
both of the major K+ transporters of this microbial pathogen.