Isolation unit for multidrug-resistant tuberculosis patients in a low endemic country, a step towards the World Health Organization End TB Strategy.

Tuberculosis (TB) remains a threat to public health and is the second cause of death due to a single infectious agent after HIV/AIDS. The worldwide distribution of TB is heterogeneous. The incidence is decreasing in most high-income regions, but the situation remains worrying in many parts of the world. The emergence of Mycobacterium tuberculosis strains resistant to key agents used in treatment (rifampin and isoniazid) contributes to TB transmission around the world. To achieve TB elimination, both high and low endemic countries must upscale their efforts to decrease disease transmission and improve cure rates. Management of drug-resistant TB is of particular importance. In this paper, we discuss the different models of care of multidrug-resistant TB (MDR-TB), the ethical considerations and the specific constraints present in high income countries. The management model chosen by the Belgian TB specialists in accordance with public health authorities as well as building of a specific MDR/XDR-TB isolation unit are also discussed.

Implementation of latent tuberculosis screening in HIV care centres: evaluation in a low tuberculosis incidence setting.

The screening and treatment of latent tuberculosis infection (LTBI) to prevent active tuberculosis (TB) is recommended by the WHO in all HIV-infected patients. The aim of this study was to evaluate its implementation within Belgium's HIV care. A multiple-choice questionnaire was sent to 55 physicians working in the country's AIDS reference centres. Response rate reached 62%. Only 20% screened all their HIV-infected patients for LTBI. Screening methods used and their interpretation vary from one physician to another. The main barriers to the implementation of LTBI screening and treatment, as perceived by the participants, are lack of sensitivity of screening tools, risks associated with polypharmacy and toxicity of treatment. The poor coverage of LTBI screening reported here and the inconsistency in methods used raises concern. However, this was not unexpected as, in low-TB incidence countries, who, when and how to screen for LTBI remains unclear and published guidelines show important disparities. Recently, a targeted approach in which only HIV-infected patients at highest risk of TB are screened has been suggested. Such a strategy would limit unnecessary exposure to LTBI treatment. This methodology was approved by 80% of the participants and could therefore achieve greater coverage. Its clinical validation is still pending.

[HIV infection : Reaching a zero risk of transmission]. / Transmission du VIH : Est-ce une fatalité ?

Despite a global reduction in the prevalence of HIV-infection, the HIV-epidemic is far from over. The prevention of HIV-transmission in all its forms (sexual, mother-to-child etc) must therefore remain a pillar in the fight against AIDS, and both potent and accessible prevention strategies are required. In addition to the classical and wellknown methods such as the condom, ant iretroviral therapy represents a potent prevention tool and the residual risk of transmission of correctly treated HIV-positive persons is virtually nihil. Antiretroviral therapy may and should be used in the prevention of HIV-transmission as Treatment as Prevention (TasP), Pre-Exposure Prophylaxis (PrEP), and Post- Exposure Prophylaxis (PEP). However, because of their exorbitant costs, the accessibility of these prevention strategies is limited, particularly for the most vulnerable populations.

Si l'infection par le VIH est globalement en diminution dans le monde, nous n'apercevons pas encore la fin de l'épidémie. Dès lors, nous avons besoin de moyens performants et accessibles pour tous pour diminuer la transmission du virus, essentiellement sexuelle mais aussi via la transmission de la mère à son enfant. En plus des moyens utilisés de façon répandue tel que le préservatif, le traitement antirétroviral représente à ce jour un outil très performant en termes de prévention, à travers la prophylaxie pré-exposition, la prophylaxie postexposition et le traitement de la personne infectée qui voit son risque de transmission virtuellement annulé. Néanmoins, l'accès à ces molécules coûteuses reste difficile pour les populations les plus défavorisées.

[Screening for tuberculosis]. / Le dépistage de la tuberculose.

Rapid identification using bacteriological methods and adequate treatment of active tuberculosis cases are the most important objective of any tuberculosis activity but, in order to eliminate the disease, another important component of tuberculosis control is to reduce the vast reservoir of latent tuberculosis infections. Tuberculin skin test and interferon-gamma release assays are designed to identify immune response against mycobacterial antigens. Both tests are accurate to detect latent but not active forms of tuberculosis. Interferon-gamma release assays have higher specificity than tuberculin skin testing in BCG-vaccinated populations particularly if BCG was administered after 1 year of age. Both tests perform poorly to predict risk for progression to active tuberculosis. Screening should therefore be limited to situations with a clear likelihood of transmission after contact, taking account of the infectiousness of the index case and the intensity of exposure, or to those with a great probability of developing tuberculosis: young children and immunocompromised persons.

Immunogenicity and protective efficacy of a tuberculosis DNA vaccine.

Tuberculosis is the most widespread and lethal infectious disease affecting humans. Immunization of mice with plasmid DNA constructs encoding one of the secreted components of Mycobacterium tuberculosis, antigen 85 (Ag85), induced substantial humoral and cell-mediated immune responses and conferred significant protection against challenge with live M. tuberculosis and M. bovis bacille Calmette-Guérin (BCG). These results indicate that immunization with DNA encoding a mycobacterial antigen provides an efficient and simple method for generating protective immunity and that this technique may be useful for defining the protective antigens of M. tuberculosis, leading to the development of a more effective vaccine.

[Which immunizations for which adults in 2011?]. / Quelles vaccinations pour quels adultes en 2011?

Immunizations are extremely efficient in prevention of diseases with a lethal potential. Healthy adults, pregnant women and patients suffering from chronic diseases may have a different benefit from vaccine available in our country. Numerous health problems need to be addressed during a short consultation, relegating immunization to a position of secondary importance. This paper will address the issue of immunization in special circumstances such as: healthy adults, pregnant women, HIV-infected patients, patients with end-stage renal disease, patients with chronic liver diseases and solid organ transplant candidates and recipients.

[Pulmonary tuberculosis]. / La tuberculose pulmonaire.

Tuberculosis (TB) is a global health problem driven by poverty, HIV infection, etc. In Europe, the problem of multidrug resistance (i.e., resistance to at least rifampin and isoniazid) (MR) develops. The cases come essentially from the former U.S.S.R. In Belgium, the incidence of tuberculosis continues to decline to 9.4/100,000 inhabitants in 2008. The percentage of MR germs is 2.8%. The distribution of cases is not uniform across the country. The incidence is much higher among people recently coming from high prevalence countries than among the Belgian native. The pulmonary forms of TB are more contagious and more common. The clinical signs are frequently non specific. The diagnosis is often mentioned up after performing a chest Xray and must always be confirmed by microbiological examination and culture of several sputum or other respiratory specimens. It is very important to identify the germ, M. tuberculosis complex and to test its sensitivity to anti-TB agents. Standard treatment consists of 4 drugs: isoniazid, rifampin, ethambutol and pyrazinamide for 2 months followed by rifampin and isoniazid for at least 4 additional months. In suspected cases of MR, 5 drugs are prescribed at the outset. Treatment and duration will be adjusted according to the results of susceptibility testing. The potential toxicities of second-line drugs should be well known by the physicians. Compliance of the patient is essential. Screening in the entourage is part of the therapeutic process.

Persistent T cell and B cell activities in the duodenal mucosa of AIDS patients.

OBJECTIVE: As HIV infection most commonly occurs via a mucosal surface, and as gastrointestinal symptoms are very frequent among HIV-infected patients, we investigated the functional properties of residual lymphocytes in the duodenal mucosa from HIV-infected individuals. DESIGN: Duodenal biopsies and blood samples were obtained from 19 HIV-infected patients [Centers for Disease Control and Prevention (CDC) stage III] and from 19 controls. METHODS: Phenotypic analysis of lymphocytes was performed by flow cytometry and/or immunocytochemistry. Interferon gamma (IFN-gamma), interleukin (IL) 4 and immunoglobulin secretions were analysed by enzyme-linked immunospot techniques. The phenotype of cytokine-producing cells was analysed by flow cytometry. RESULTS: The proportions of duodenal T lymphocytes from HIV-infected patients spontaneously secreting IFN-gamma or IL-4 were not lower than those from healthy controls. In patients with a high intestinal mucosal viral load, they were higher than in controls (P < 0.05). The proportions of immunoglobulin-secreting cells were significantly raised in HIV-infected patients for the three main isotypes. CONCLUSIONS: T- and B-cell populations of the intestinal mucosa remain functional or are even activated in patients with AIDS, even when the numbers of both mucosal and circulating CD4+ lymphocytes are strongly decreased.

Iron chelators inhibit the growth and induce the apoptosis of Kaposi's sarcoma cells and of their putative endothelial precursors.

Iron is suspected to be involved in the induction and/or progression of various human tumors. More particularly, iron may be involved in the pathogenesis of Kaposi's sarcoma, a tumor of probable vascular origin. This study was designed to investigate the effect of iron deprivation on Kaposi's sarcoma. The effects of iron chelators and iron deprivation associated with serum withdrawal were investigated on Kaposi's sarcoma-derived spindle cells, on a transformed Kaposi's sarcoma cell line (Kaposi's sarcoma Y-1) and on endothelial cells, which are the probable progenitors of Kaposi's sarcoma cells. Desferrioxamine and deferiprone, two chemically unrelated iron chelators, induced a time- and concentration-dependent inhibition of endothelial and Kaposi's sarcoma cell growth. The inhibition of cell growth was associated with a decrease in Ki-67 and in both stable and total proliferating cell nuclear antigen expression. Inhibition of the progression through the G1-phase of the cell cycle was further evidenced by decreased expression of cyclin D1 and of p34 cyclin-dependent kinase 4. Terminal deoxynucleotidyl transferase-mediated desoxyuridinetriphosphate nick end labeling assay, flow cytometry with annexin-V-fluorescein and morphologic analysis indicated that iron chelation also induced a time- and concentration-dependent apoptosis. This apoptotic effect was prevented by the addition of exogenous iron. Induction of iron deprivation in the culture medium by serum withdrawal led to similar cell cycle effects, which, however, could only be partly reverted by the addition of exogenous iron. In conclusion, these results show that iron deprivation inhibits the growth and induces the apoptosis of Kaposi's sarcoma cells and of their putative endothelial precursors. This suggests that iron chelators may represent a potential therapeutic approach for the treatment of Kaposi's sarcoma.

Early- and late-stage Kaposi's sarcoma-derived cells but not activated endothelial cells can invade de-epidermized dermis.

Whether Kaposi's sarcoma is a true neoplasm or a reactive endothelial cell outgrowth triggered by inflammatory cytokines remains unclear. In this study, we investigated the differential invasive properties of activated endothelial cells and Kaposi's sarcoma cells in a model of de-epidermized dermis, supplying the cells with matrix barriers similar to those found in vivo. Cells derived from early "patch-stage" and from late "nodular-stage" Kaposi's sarcoma lesions exhibited similar invasive properties, which indicates that cells with an invasive potential are present in the early stages of tumor development. Slow accumulation of the cells into the extracellular matrix, together with a low proliferation index and with expression of anti-apoptotic proteins, suggest that the progression of Kaposi's sarcoma may be related to escape from cell death rather than to increased proliferation. The Kaposi's sarcoma-Y1 cell line, which is tumorigenic in nude mice, also exhibited invasive properties. By contrast to the Kaposi's sarcoma-derived spindle cells, however, which were scattered between the collagen bundles, the Kaposi's sarcoma-Y1 cell population had a higher proliferation index and displayed a multilayer arrangement. Inflammatory cytokines and Kaposi's sarcoma cell supernatant could activate and stimulate the growth of human dermal microvascular endothelial cell, but could not induce their invasion in this model, showing that activated endothelial cells do not fit all the requirements to traverse the various barriers found in the dermal extracellular matrix. These results confer to Kaposi's sarcoma cells a tumor phenotype and suggest that the in vivo dominant endothelial cell population represents a reactive hyperplasia rather than the true tumor process.

Overexpression of Bcl-2 in Kaposi's sarcoma-derived cells.

The pathogenesis of Kaposi's sarcoma (KS), a tumor of probable vascular origin, remains an enigma. It is still unclear whether KS is a true malignancy or whether it represents a reactive polyclonal process. Using both an immunohistochemical and an immunoblot approach, we found that cells derived from KS lesions express significant levels of Bcl-2, a protein known to prolong cellular viability and to antagonize apoptosis. Bcl-2 expression was found in AIDS-related KS-derived cells, as well as in cells derived from iatrogenic and sporadic KS, indicating that Bcl-2 upregulation may be important in the pathogenesis of KS regardless of its epidemiologic form. By contrast, fibroblasts and dermal microvascular endothelial, cells which are the probable vascular progenitors of KS cells, expressed low levels of Bcl-2. The expression of Bcl-2 in KS-derived cells was associated with a long-term survival in serum-deprived conditions, a situation that has been shown to induce apoptosis in various cell types. Incubation of fibroblasts or of dermal microvascular endothelial cells with KS cell-free supernatants did not enhance Bcl-2 expression, suggesting that Bcl-2 expression is not mediated by an agent released by KS cells. Analogously, KS supernatants failed to promote the viability of fibroblasts and of dermal microvascular endothelial cells cultured in serum-free conditions. Our findings suggest that the spindle cells derived from KS have a survival advantage and may adequately represent the tumor cells of KS.

Immunological detection of mycobacterial antigens in infected fluids, cells and tissues by latex agglutination. Animal model and clinical application.

We devised an immunoassay for the detection of mycobacterial antigens in cell lysates and in tissue extracts which is based on the agglutination of latex particles coated with anti-Mycobacterium bovis F(ab')2, followed by counting of non-agglutinated particles. Mycobacterium bovis cell lysates were tested and a reference curve was established, having a lower limit of detection of 15-20 Mycobacteria. We were able to detect mycobacterial antigens in cell lysates from bronchoalveolar washings and in spleen and liver lysates obtained from experimentally infected rabbits. Antigens were also detected in ten out of 11 samples obtained from patients with proven tuberculous infection. These samples were readily distinguished from 32 negative control samples after pepsin treatment. In contrast, periodate treatment of samples to destroy carbohydrate, abolished all reactivity. Following gel filtration chromatography we identified three peaks with antigenic properties in samples of all types. The detection of mycobacterial carbohydrate antigens by latex agglutination and particle counting should be a useful adjunct in the diagnosis of tuberculosis.

An isoelectric focusing method for the study of the humoral response against the antigen 85 complex of Mycobacterium bovis BCG in the different forms of leprosy.

Isoelectric focusing was used to separate the three components of the antigen 85 complex of Mycobacterium bovis BCG. Antibody responses of leprosy patients against each Mycobacterium bovis BCG. Antibody responses of leprosy patients against each component were quantitated by densitometric analysis of immunoblot assays. The 85A component was recognized by 40% (8/20) of the lepromin positive and negative healthy subjects, by 76% (19/25) of the tuberculoid and by 96% (24/25) of the lepromatous leprosy sera. In contrast, the 85B component was not stained by the control sera, nor by the tuberculoid leprosy sera but by 64% (16/25) of the lepromatous leprosy sera. The results suggest that antigen 85B contains one or several epitopes that are specifically recognized by sera of lepromatous leprosy patients only.

A multidot immunobinding assay for the serodiagnosis of tuberculosis. Comparison with an enzyme-linked immunosorbent assay.

A simple dot immunobinding (dot blot) assay procedure has been developed for the detection of antibodies directed against a soluble mycobacterial antigen preparation. This technique was compared with the widely used ELISA, in a study of samples from tuberculous patients. Dot blots were read on a densitometer. The correlation between both assays was excellent (r = 0.91; P less than 0.001); 90% of sera from tuberculous patients were detected using both techniques and a serial two-fold dilution method. Assessments of the end-points of titration curves by reflectometry and simple visual interpretation gave similar results. The dot blot assay is easier to perform and appears to be a practical alternative to ELISA for the detection of anti-mycobacterial antibodies in tuberculous patients.

Detection of lytic antimycobacterial antibodies by immune lysis of liposomes sensitized to tuberculin.

Two procedures were used in order to incorporate purified protein derivative tuberculin (PPD) from M. tuberculosis, strain H37Rv, into calcein-containing liposomes: formation of multilamellar vesicles (MLV) in a PPD solution or exposure of preformed MLV to this solution. Immune lysis of these PPD-sensitized MLV was studied in the presence of a hyperimmune anti-M. tuberculosis sheep serum using a specific pathogen-free rabbit serum as a source of complement. A 50% release of encapsulated calcein was observed spectrofluorometrically after 30 min and remained unchanged up to 2 h. The release of calcein in the absence of complement or of anti-H37Rv serum or by liposomes which did not contain PPD never exceeded 1-2%. Liposomes formed in PPD solution were more sensitive to anti-H37Rv serum than preformed liposomes exposed to PPD. Trials with human sera from ten tuberculous patients revealed the presence of specific lytic immunoglobulins. In the presence of sera from skin test negative, non-tuberculous subjects, calcein release was significantly lower. This opens the way to a new method for the study of the humoral immunity in tuberculosis.

Antimycobacterial antibodies in pleural effusions.

We devised a dot blot assay to evaluate the IgG and IgA response to P32, a recently isolated antigen specific to mycobacteria. Pleural fluids and the corresponding sera were tested, obtained from five patients with pleural tuberculosis proven by direct examination and/or culture and from 14 patients with pleural effusions of other origins. We measured the total IgG and IgA levels in all samples and determined the anti-P32 titer after adjusting IgG and IgA respectively to the same levels in all samples. Those pleural fluids and sera from tuberculous patients contained a higher proportion of anti-P32 antibodies than samples obtained from nontuberculous control subjects; in those patients, the proportion of anti-P32 antibodies was generally higher in pleural effusion fluid than in serum.

Antibody levels to whole culture filtrate antigens and to purified P32 during treatment of smear-positive tuberculosis.

Antimycobacterial IgG levels were measured repeatedly during treatment in 12 patients with moderate or severe pulmonary tuberculous disease using a dot immunobinding assay. We used reflectance densitometry equipment to quantify the immunoperoxidase staining and a Mycobacterium bovis BCG culture filtrate and the purified P32 protein as antigens. Antibody response against whole culture filtrate and P32 antigen increased after a three-month period of treatment. After this antibiotherapy was completed, the estimated amount of antibodies directed against the P32 decreased while those against the whole culture filtrate remained at the same level. A serologic test using P32 as the antigen seems to allow a better discrimination between active and healed tuberculosis.

High-dose chemotherapy and autologous CD34-positive blood stem cell transplantation for multiple myeloma in an HIV carrier.

The epidemiology and clinical outcome of multiple myeloma in human immunodeficiency virus (HIV)-positive patients is poorly documented. There are uncertainties concerning the optimal management of this rare disorder. We report on the use of myeloablative chemotherapy with autologous stem cell transplantation in an HIV-positive patient with multiple myeloma.

Autologous stem cell infusion for acute myeloblastic leukemia in an HIV-1 carrier.

We present the case of an asymptomatic HIV carrier, who presented with acute myeloblastic leukemia in third relapse and successfully underwent autologous stem cell transplantation as a rescue treatment. This observation supports the conclusion that tolerance of autologous bone marrow or stem cell transplant in patients with HIV may correlate with a low viral burden and relatively good immune function.

Fifty cases of human immunodeficiency virus (HIV) infection: immunoultrastructural study of circulating lymphocytes.

The peripheral lymphocytes of 50 cases of human immunodeficiency virus (HIV) infection (13 of acquired immune deficiency syndrome (AIDS), 17 of AIDS related complex (ARC), and 20 healthy carriers) were studied immunoultrastructurally. The prevalence of "tubuloreticular structures" and "tubular confronting cisternae" increased with the progression of the disease. Numerous tubular confronting cisternae were noted in patients presenting with a high serum acid labile alpha-interferon values. The patients with depressed natural killer cell activity were characterised by circulating immature natural killer cells with abundant multivesicular bodies that were devoid of "parallel tubular arrays". With an immunogold staining technique the location of HIV antigen was detected ultrastructurally, both at the surface of "hand-mirror" natural killer cell lymphocytes and inside vacuolised cells, probably corresponding to infected T4 lymphocytes. These findings indicate the usefulness of electron microscopic techniques in evaluating the pathology and the pathogenetic outcome of AIDS.