[Frontier of mycobacterium research--host vs. mycobacterium]

During the past decade, we have observed advance in tuberculosis research including novel vaccines, innate immunity (TLR), SNIP analysis and molecular mechanism of drug resistance. Worldwide genome project enabled the whole genome sequence of host resistant against tuberculosis as well as the whole genome sequence of M. tuberculosis H37Rv. DNA technology has also provided a great impact on the development of novel vaccine against TB. In this symposium, we have invited leading researchers in the field of the frontier study of Mycobacterium research in order to provide general overview of the cutting edge of frontier research. Molecular mechanism of drug resistance of M. tuberculosis has been clarified. On the other hand, molecular mechanism of host-defence (insusceptibility of host) against M. tuberculosis has not yet elucidated. Dr. Taro Shirakawa (Kyoto University) reviewed the susceptibility genes of host in TB infection and presented candidate genes associated with multi-drug resistant tuberculosis. Dr. Naoto Keicho (International Medical Center of Japan) tried to identify host genetic factors involved in susceptibility to pulmonary Mycobacterium avium complex (MAC) infection by candidate gene approach and genome-wide approach. In Japan, Dr. Masaji Okada (National Hospital Organization Kinki-Chuo Chest Medical Center) has been engaged actively in the development of new tuberculosis vaccines (HVJ-liposome/Hsp65 DNA + IL-12 DNA vaccine and recombinant 72f BCG vaccine). He showed basic strategy for construction of new candidate vaccines and also showed significant efficacy on the protection of tuberculosis infection using cynomolgus monkeys, which are very similar to human tuberculosis. Dr. Hatsumi Taniguchi (University of Occupational and Environmental Health) presented that M. tuberculosis mIHF and the neighbor genes went into a dormacy-like state of M. smegmatis in J774 macrophage cells. This study might provide a weapon for elucidating the mechanism of dormacy of M. tuberculosis and the development of novel therapy. Dr. Chiyoji Abe (Nippon Becton Dickinson Co.) reviewed the molecular basis of the resistance to anti-tuberculosis drugs. Most cases of resistance are related to simple nucleotide substitutions rather than to acquisition of new elements. Dr. Kiyoshi Takeda (Kyushu University) showed interesting finding. He analyzed whether Toll-like receptor (TLR)-mediated activation of innate immunity in host defense against mycobacterial infection. MyD88/TRIF double defi-indicating that innate immunity is involved in anti-mycobacterial infection. (1) SNP (single nucleotide polymorphism) analysis in association with Mycobacterium tuberculosis: Taro SHIRAKAWA (Department of Health Promotion & Human Behavior, Kyoto University Medical School, and RIKEN SRC Center) Candidate gene approach was made on 18 SNPs in 11 genes in association with M. tuberculosis. Patients with multi-drug resistance against M. tuberculosis are also subjected. SNPs in NRAMP1 gene were associated with the disease, and drug resistance, its mechanisms remain unknown. (2) Search for genes susceptible to pulmonary Mycobacterium avium complex infection: Naoto KEICHO (Department of Respiratory Diseases, Research Institute, International Medical Center of Japan) Interaction among pathogens and host factors is important for development of infectious diseases. We are trying to identify host genetic factors involved in susceptibility to nonimmunocompromized pulmonary Mycobacterium avium complex (MAC) infection by candidate gene approach and genome-wide approach. Elucidation of functional significance of susceptibility gene polymorphisms will lead to a new strategy for control and prevention of the disease. (3) T cell immunity against Tuberculosis in host and the establishment of novel vaccine: Masaji OKADA (Clinical Research Center, National Hospital Organization Kinki-Chuo Chest Medical Center) T cell (CTL, Th1) immunity including granulysin play an important role in host defense against tuberculosis (TB) in human. Patients with TB or Multi-drug resistant TB showed suppression of all these immunities. HVJ-liposome/Hsp65 DNA + IL-12 DNA vaccination was 100 fold more efficient than BCG on the elimination of Mycobacterium tuberculosis (M.TB) in the BALB/c mice. Cytotoxic T cells activity against M. TB was augmented. By using these new vaccines (Hsp 65 DNA + IL-12 DNA, recombinant 72f BCG) and the cynomolgus monkey models which are very similar to human tuberculosis, the prophylactic effect of vaccines was observed. Thus, these novel vaccines should provide a useful tool for the prevention of human TB infection. (4) Mycobacterium tuberculosis mIHF and the neighbor genes go into a dormancy-like state of M. smegmatis J15CS in J774 cells: Hatsumi TANIGUCHI (Department of Microbiology, School of Medicine, University of Occupational and Environmental Health) Mycobacterium smegmatis J15CS transformants harboring the mIHF gene or the mIHF-gmk-Rv1390 genes showed no difference in in vitro growth and acid-fastness. However, transformants harboring mIHF-gmk-Rv1390 formed short-rod cell morphology and decreased acid-fastness in the mouse macrophage-like cell line J774 compared to those of the other transformants, and the nuclei of the infected J774 cells also changed. Nevertheless, the colony forming units were similar. These indicate that mIHF and the neighbor genes of M. tuberculosis might regulate a growth of mycobacteria in macrophages. (5) Molecular basis of the resistance to anti-tuberculosis drugs: Chiyoji ABE (Nippon Becton Dickinson Company, Ltd.) Considerable progress has been made toward understanding the molecular basis of the resistance to anti-tuberculosis drugs. Most cases of resistance are related usually to simple nucleotide substitutions rather than to acquisition of new elements. Multi-drug resistant isolates of Mycobacterium tuberculosis arise a consequence of sequential accumulation of mutation conferring resistance to single therapeutic agents. The basis of resistance is not able to be explained yet in a substantial percentage of strains for other anti-tuberculosis drugs than rifampin and pyrazinamide. Further studies are required to fully understand the molecular mechanisms of resistance. (6) Toll-like receptors in anti-mycobacterial immune responses: Kiyoshi TAKEDA (Department of Molecular Genetics, Medical Institute of Bioregulation, Kyushu University) Toll-like receptors (TLRs) play an essential role in the recognition of specific patterns of microbial components. TLRs mediate activation of innate immunity and further development of antigen-specific adaptive immunity. In TLR signaling pathways, Toll/IL-1 receptor (TIR) domain-containing adaptors, such as MyD88, TIRAP, TRIF, and TRAM, have been shown to play pivotal roles. Thus, the molecular mechanisms for TLR-mediated activation of innate immunity have been largely understood. We analyzed whether TLR-mediated activation of innate immunity is involved in host defense against mycobacterial infection. MyD88/TRIF double deficient mice, in which TLR-dependent activation of innate immunity is abolished, showed high sensitivity to mycobacterial infection, indicating that innate immunity is critically involved in anti-mycobacterial responses.