TITLE:
Evaluation of Recombinant Mycobacterium tuberculosis Antigens MPT64, CFP10, and ESAT6 for Delayed-Type Hypersensitivity Responses in Guinea Pigs
AUTHORS:
Wandee Yindeeyoungyeon, Kamolchanok Rukseree, Siriporn Tungsudjai, Tasanee Panichakul
KEYWORDS:
M. tuberculosis, M. bovis BCG Tokyo, Delayed-Type Hypersensitivity, DTH, MPT64, CFP10, ESAT6
JOURNAL NAME:
Advances in Microbiology,
Vol.5 No.8,
August
7,
2015
ABSTRACT: The tuberculin Purified Protein Derivative (PPD) skin test is widely
used; however, the results are often inaccurate. Positive results can be
observed in patients with active tuberculosis (TB) as well as in BCG-vaccinated
persons and individuals who are infected with mycobacteria but have not
developed the disease. MPT64, an antigen secreted from actively growing Mycobacterium
tuberculosis and some strains of M. bovis BCG such as BCG Tokyo and
BCG Russia, is immunogenic and elicits delayed-type hypersensitivity (DTH) in
guinea pigs and humans. This antigen has been used to develop a new skin test
for the diagnosis of active TB infection. Two of the antigens encoded by the M.
tuberculosis-specific region of difference 1 (RD1, deleted in M. bovis BCG strains), CFP10 (culture filtrate protein 10) and ESAT6 (early secreted
antigenic target-6), also induce M. tuberculosis-specific DTH responses.
The aim of this study was to investigate the DTH responses in guinea pigs
infected with M. tuberculosis or M. bovis BCG Tokyo elicited by
three purified recombinant proteins (rMPT64, rCFP10 and rESAT6) compared to
those elicited by PPD. In this study genes encoding MPT64, CFP10, and ESAT6
were cloned and expressed as recombinant proteins with the addition of a
C-terminal His6 tag for ease of purification by Immobilized Metal
ion Affinity Chromatography (IMAC). The recombinant proteins (rMPT64, rCFP10,
and rESAT6) were purified to homogeneity and were used to elicit DTH responses
in guinea pigs infected with M. tuberculosis or M. bovis BCG
Tokyo. The results showed that rMPT64 elicits a DTH response comparable to that
of PPD in M. bovis BCG Tokyo-vaccinated animals. However, M.
tuberculosis-infected animals show less reactivity to rMPT64 than they do
to PPD. Although single rCFP10 or rESAT6 did not readily elicit a DTH response
in M. tuberculosis-infected animals, combining these antigens
with rMPT64 led to an increased DTH response, thus enabling the detection of TB
infection.