Early interactions between lung dendritic cells (LDCs) and Mycobacterium tuberculosis, the etiological agent of tuberculosis, are thought to be critical for mounting a protective anti-mycobacterial immune response and for determining the outcome of infection. However, these interactions are poorly understood, at least at the molecular level. Here we show that M. tuberculosis enters human monocyte-derived DCs after binding to the recently identified lectin DC-specific intercellular adhesion molecule-3 grabbing nonintegrin (DC-SIGN). By contrast, complement receptor (CR)3 and mannose receptor (MR), which are the main M. tuberculosis receptors on macrophages (Mφs), appeared to play a minor role, if any, in mycobacterial binding to DCs. The mycobacteria-specific lipoglycan lipoarabinomannan (LAM) was identified as a key ligand of DC-SIGN. Freshly isolated human LDCs were found to express DC-SIGN, and M. tuberculosis–derived material was detected in CD14−HLA-DR+DC-SIGN+ cells in lymph nodes (LNs) from patients with tuberculosis. Thus, as for human immunodeficiency virus (HIV), which is captured by the same receptor, DC-SIGN–mediated entry of M. tuberculosis in DCs in vivo is likely to influence bacterial persistence and host immunity.

Background Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), has the ability to persist in its human host for exceptionally long periods of time. However, little is known about the location of the bacilli in latently infected individuals. Long-term mycobacterial persistence in the lungs has been reported, but this may not sufficiently account for strictly extra-pulmonary TB, which represents 10–15% of the reactivation cases.Methodology/Principal FindingsWe applied in situ and conventional PCR to sections of adipose tissue samples of various anatomical origins from 19 individuals from Mexico and 20 from France who had died from causes other than TB. M. tuberculosis DNA could be detected by either or both techniques in fat tissue surrounding the kidneys, the stomach, the lymph nodes, the heart and the skin in 9/57 Mexican samples (6/19 individuals), and in 8/26 French samples (6/20 individuals). In addition, mycobacteria could be immuno-detected in perinodal adipose tissue of 1 out of 3 biopsy samples from individuals with active TB. In vitro, using a combination of adipose cell models, including the widely used murine adipose cell line 3T3-L1, as well as primary human adipocytes, we show that after binding to scavenger receptors, M. tuberculosis can enter within adipocytes, where it accumulates intracytoplasmic lipid inclusions and survives in a non-replicating state that is insensitive to the major anti-mycobacterial drug isoniazid.Conclusions/SignificanceGiven the abundance and the wide distribution of the adipose tissue throughout the body, our results suggest that this tissue, among others, might constitute a vast reservoir where the tubercle bacillus could persist for long periods of time, and avoid both killing by antimicrobials and recognition by the host immune system. In addition, M. tuberculosis-infected adipocytes might provide a new model to investigate dormancy and to evaluate new drugs for the treatment of persistent infection.

SummaryThe growth of Mycobacterium tuberculosis mutants unable to synthesize phthiocerol dimycocerosates (DIMs) was recently shown to be impaired in mouse lungs. However, the precise role of these molecules in the course of infection remained to be determined. Here, we provide evidence that the attenuation of a DIM-deficient strain takes place during the acute phase of infection in both lungs and spleen of mice, and that this attenuation results in part from the increased sensitivity of the mutant to the cidal activity of reactive nitrogen intermediates released by activated macrophages. We also show that the DIM-deficient mutant, the growth and survival of which were not impaired within resting macrophages and dendritic cells, induced these cells to secrete more tumour necrosis factor (TNF)-a a a a and interleukin (IL)-6 than the wild-type strain. Although purified DIM molecules by themselves had no effect on the activation of macrophages and dendritic cells in vitro , we found that the proper localization of DIMs in the cell envelope of M. tuberculosis is critical to their biological effects. Thus, our findings suggest that DIM production contributes to the initial growth of M. tuberculosis by protecting it from the nitric oxide-dependent killing of macrophages and modulating the early immune response to infection.

SummaryErp (exported repetitive protein), also known as P36, Pirg and Rv3810, is a member of a mycobacteriaspecific family of extracellular proteins. These proteins consist of three domains, the N-and C-terminal domains are similar in all mycobacterial species, however, the central domain contains a repeated PGLTS module and differs considerably between species. The erp knockout mutant of Mycobacterium tuberculosis displays very low levels of multiplication both in macrophage cell lines and in vivo in a mouse model of infection. The high interspecies variability of the central repeated region of the Erp protein led us to investigate whether these orthologous proteins were functionally equivalent in a mouse model of tuberculosis. We expressed a gene fusion with the erp gene of Mycobacterium smegmatis , Mycobacterium leprae or M. tuberculosis in trans in an erp -M. tuberculosis mutant and found that these three alleles restored multiplication to similar levels in the spleen of infected mice. However, these alleles gave different levels of colonization in the lung, for the early timepoints. Quantitative histological analyses of the lungs of infected animals showed that the nature of the erp allele strongly affected the number and the size of lung lesions, demonstrating the importance of surface determinants for virulence and tissue damage.

Erp (exported repetitive protein) is a member of a mycobacterium-specific family of extracellular proteins. A hydrophobic region that is localized at the C-terminal domain and that represents a quarter of the protein is highly conserved across species. Here we show that this hydrophobic region is not essential for restoring the virulence and tissue damage of an erp::aph mutant strain of M. tuberculosis as assessed by bacterial counts and lung histology analysis in a mouse model of tuberculosis.The genus Mycobacterium is very large and encompasses intra-and extracellular pathogens and environmental opportunistic pathogens as well as saprophytes (20). Mycobacterium tuberculosis, a facultative intracellular pathogen that persists within professional phagocytes, is responsible for more than 1.5 millions deaths per year (WHO information tuberculosis fact sheet, 2002; World Health Organization [http://www-.who.int/mediacentre/factsheets /who104/en/index.html]). The situation is worsened by the AIDS epidemic and the deteriorating socioeconomic conditions of increasing numbers of people. Moreover, the recent emergence of multidrug-resistant strains, together with the relative inefficacy of the M. tuberculosis BCG vaccine, has aggravated the situation.The development and the utilization of classical bacterial genetics in M. tuberculosis (19) have enabled the characterization of virulence factors (5, 8; for a review, see reference 11). To date, most of the characterized virulence factors have been products that are involved in the structure and function of the cell wall, a particularity of the Corynebacteria-MycobacteriaNocardia group of Actinomycetales (9, 20). The erp gene of M. tuberculosis has been characterized before completion of the genome sequence through the use of the phoA fusion method that helps to entrap genes coding for exported and secreted products (2, 15). Erp has been shown to be a crucial virulence factor. Indeed, disruption of the erp gene by insertion of an antibiotic element results in a marked decrease in virulence, with lower levels of survival and multiplication both in vitro in cell culture assays and in vivo in the mouse model of infection (3). The erp gene was later shown to be present not only in pathogenic mycobacteria but also in saprophytic and opportunistic pathogenic mycobacteria (17). However, no homologue has been found in other bacterial species, making Erp a mycobacterial signature.The Erp protein has a composite structure made of three domains. Both the amino (1 to 80)-and carboxy (176 to 284)-terminal domains are conserved, while the central domain is subject to high-level interspecies variability. It was recently shown that the nature of the erp allele strongly affects the number and the size of the lung lesions of infected animals (18). None of the three domains exhibit homology to other characterized proteins. The central domain consists of tandem repeats of five amino acids based on a PGLTS motif (variable both in the number and quality of repeats between mycobacterial species)...