Outbreak of West Nile virus causing severe neurological involvement in children, Nuba Mountains, Sudan, 2002.

An atypical outbreak of West Nile virus (WNV) occurred in Ngorban County, South Kordophan, Sudan, from May to August 2002. We investigated the epidemic and conducted a case-control study in the village of Limon. Blood samples were obtained for cases and controls. Patients with obvious sequelae underwent cerebrospinal fluid (CSF) sampling as well. We used enzyme-linked immunosorbent assay (ELISA) and neutralization tests for laboratory diagnosis and identified 31 cases with encephalitis, four of whom died. Median age was 36 months. Bivariate analysis did not reveal any significant association with the risk factors investigated. Laboratory analysis confirmed presence of IgM antibodies caused by WNV in eight of 13 cases, indicative of recent viral infection. The unique aspects of the WNW outbreak in Sudan, i.e. disease occurrence solely among children and the clinical domination of encephalitis, involving severe neurological sequelae, demonstrate the continuing evolution of WNV virulence. The spread of such a virus to other countries or continents cannot be excluded

T-cell and serological responses to Erp, an exported Mycobacterium tuberculosis protein, in tuberculosis patients and healthy individuals

<p>Abstract</p> <p>Background</p> <p>The identification of antigens able to differentiate tuberculosis (TB) disease from TB infection would be valuable. Cellular and humoral immune responses to Erp (Exported repetitive protein) – a recently identified <it>M. tuberculosis </it>protein – have not yet been investigated in humans and may contribute to this aim.</p> <p>Methods</p> <p>We analyzed the cellular and humoral immune responses to Erp, ESAT-6, Ag85B and PPD in TB patients, in BCG⁺individuals without infection, BCG⁺individuals with latent TB infection (LTBI) and BCG^-controls. We used lymphoproliferation, ELISpot IFN-γ, cytokine production assays and detection of specific human antibodies against recombinant <it>M. tuberculosis </it>proteins.</p> <p>Results</p> <p>We included 22 TB patients, 9 BCG⁺individuals without TB infection, 7 LTBI and 7 BCG^-controls. Erp-specific T cell counts were higher in LTBI than in the other groups. Erp-specific T cell counts were higher in LTBI subjects than TB patients (median positive frequency of 211 SFC/10⁶PBMC (range 118–2000) for LTBI subjects compared to 80 SFC/10⁶PBMC (range 50–191), p = 0.019); responses to PPD and ESAT-6 antigens did not differ between these groups. IFN-γ secretion after Erp stimulation differed between TB patients and LTBI subjects (p = 0.02). Moreover, LTBI subjects but not TB patients or healthy subjects produced IgG3 against Erp.</p> <p>Conclusion</p> <p>The frequencies of IFN-γ-producing specific T cells, the IFN-γ secretion and the production of IgG3 after Erp stimulation are higher in LTBI subjects than in TB patients, whereas PPD and ESAT-6 are not.</p

ESAT-6/CFP10 Skin Test Predicts Disease in M. tuberculosis-Infected Guinea Pigs

Background: Targeted preventive chemotherapy of individuals with progressive subclinical (incipient) disease before it becomes contagious would break the chain of tuberculosis transmission in high endemic regions. We have studied the ability of a skin test response to ESAT-6 and CFP10 (E6/C10) to predict later development of tuberculosis disease in the guinea pig model. Methods and Findings: Guinea pigs, either vaccinated with BCG or unvaccinated, were infected with a low dose of Mycobacterium tuberculosis by the aerosol route and the development of delayed type hypersensitivity responses to E6/C10 and to purified protein derivative (PPD) were followed until the onset of clinical disease. We demonstrated a negative correlation between the size of the skin test response and the time to the onset of clinical disease; a large E6/C10 skin test response correlated to a shorter survival time post skin testing, while a small E6/C10 skin test reaction correlated with a longer survival time (r = 20.6 and P,0.0001). No correlation was found using PPD. Conclusions: Our data suggest that it may be possible to develop a prognostic skin test based on E6/C10 that will allow the identification of individuals with incipient disease, who have the highest risk of developing active tuberculosis in the near future

Surprisingly High Specificity of the PPD Skin Test for M. tuberculosis Infection from Recent Exposure in The Gambia

BACKGROUND: Options for intervention against Mycobacterium tuberculosis infection are limited by the diagnostic tools available. The Purified Protein Derivative (PPD) skin test is thought to be non-specific, especially in tropical settings. We compared the PPD skin test with an ELISPOT test in The Gambia. METHODOLOGY/PRINCIPAL FINDINGS: Household contacts over six months of age of sputum smear positive TB cases and community controls were recruited. They underwent a PPD skin test and an ELISPOT test for the T cell response to PPD and ESAT-6/CFP10 antigens. Responsiveness to M. tuberculosis exposure was analysed according to sleeping proximity to an index case using logistic regression. 615 household contacts and 105 community controls were recruited. All three tests assessed increased significantly in positivity with increasing M. tuberculosis exposure, the PPD skin test most dramatically (OR 15.7; 95% CI 6.6–35.3). While the PPD skin test positivity continued to trend downwards in the community with increasing distance from a known case (61.9% to 14.3%), the PPD and ESAT-6/CFP-10 ELISPOT positivity did not. The PPD skin test was more in agreement with ESAT-6/CFP-10 ELISPOT (75%, p = 0.01) than the PPD ELISPOT (53%, p<0.0001). With increasing M. tuberculosis exposure, the proportion of ESAT-6/CFP-10 positive contacts who were PPD skin test positive increased (p<0.0001), and the proportion of ESAT-6/CFP-10 negative contacts that were PPD skin test negative decreased (p<0.0001); the converse did not occur. CONCLUSIONS/SIGNIFICANCE: The PPD skin test has surprisingly high specificity for M. tuberculosis infection from recent exposure in The Gambia. In this setting, anti-tuberculous prophylaxis in PPD skin test positive individuals should be revisited

Utility of interferon-γ ELISPOT assay responses in highly tuberculosis-exposed patients with advanced HIV infection in South Africa

BACKGROUND: Interferon-gamma (IFN-gamma) ELISPOT assays incorporating Mycobacterium tuberculosis-specific antigens are useful in the diagnosis of tuberculosis (TB) or latent infection. However, their utility in patients with advanced HIV is unknown. We studied determinants of ELISPOT responses among patients with advanced HIV infection (but without active TB) living in a South African community with very high TB notification rates. METHODS: IFN-gamma responses to ESAT-6 and CFP-10 in overnight ELISPOT assays and in 7-day whole blood assays (WBA) were compared in HIV-infected patients (HIV+, n = 40) and healthy HIV-negative controls (HIV-, n = 30) without active TB. Tuberculin skin tests (TSTs) were also done. RESULTS: ELISPOTs, WBAs and TSTs were each positive in >70% of HIV- controls, reflecting very high community exposure to M. tuberculosis. Among HIV+ patients, quantitative WBA responses and TSTs (but not the proportion of positive ELISPOT responses) were significantly impaired in those with CD4 cell counts <100 cells/mul compared to those with higher counts. In contrast, ELISPOT responses (but not WBA or TST) were strongly related to history of TB treatment; a much lower proportion of HIV+ patients who had recently completed treatment for TB (n = 19) had positive responses compared to those who had not been treated (11% versus 62%, respectively; P < 0.001). Multivariate analysis confirmed that ELISPOT responses had a strong inverse association with a history of recent TB treatment (adjusted OR = 0.06, 95%CI = 0.10-0.40, P < 0.01) and that they were independent of CD4 cell count and viral load. Among HIV+ individuals who had not received TB treatment both the magnitude and proportion of positive ELISPOT responses (but not TST or WBA) were similar to those of HIV-negative controls. CONCLUSION: The proportion of positive ELISPOT responses in patients with advanced HIV infection was independent of CD4 cell count but had a strong inverse association with history of TB treatment. This concurs with the previously documented low TB risk among patients in this cohort with a history of recent treatment for TB. These data suggest ELISPOT assays may be useful for patient assessment and as an immuno-epidemiological research tool among patients with advanced HIV and warrant larger scale prospective evaluation

Safety and immunogenicity of the candidate tuberculosis vaccine MVA85A in West Africa.

BACKGROUND: Vaccination with a recombinant modified vaccinia Ankara expressing antigen 85A from Mycobacterium tuberculosis, MVA85A, induces high levels of cellular immune responses in UK volunteers. We assessed the safety and immunogenicity of this new vaccine in West African volunteers. METHODS AND FINDINGS: We vaccinated 21 healthy adult male subjects (11 BCG scar negative and 10 BCG scar positive) with MVA85A after screening for evidence of prior exposure to mycobacteria. We monitored them over six months, observing for clinical, haematological and biochemical adverse events, together with assessment of the vaccine induced cellular immune response using ELISPOT and flow cytometry. MVA85A was well tolerated with no significant adverse events. Mild local and systemic adverse events were consistent with previous UK trials. Marked immunogenicity was found whether individuals had a previous BCG scar or not. There was not enhanced immunogenicity in those with a BCG scar, and induced T cell responses were better maintained in apparently BCG-naïve Gambians than previously studied BCG-naïve UK vaccinees. Although responses were predominantly attributable to CD4+ T cells, we also identified antigen specific CD8+ T cell responses, in subjects who were HLA B-35 and in whom enough blood was available for more detailed immunological analysis. CONCLUSIONS: These data on the safety and immunogenicity of MVA85A in West Africa support its accelerated development as a promising booster vaccine for tuberculosis. TRIAL REGISTRATION: ClinicalTrials.gov NCT00423839

Glutamate mediated metabolic neutralization mitigates propionate toxicity in intracellular Mycobacterium tuberculosis

Metabolic networks in biological systems are interconnected, such that malfunctioning parts can be corrected by other parts within the network, a process termed adaptive metabolism. Unlike Bacillus Calmette-Guérin (BCG), Mycobacterium tuberculosis (Mtb) better manages its intracellular lifestyle by executing adaptive metabolism. Here, we used metabolomics and identified glutamate synthase (GltB/D) that converts glutamine to glutamate (Q → E) as a metabolic effort used to neutralize cytoplasmic pH that is acidified while consuming host propionate carbon through the methylcitrate cycle (MCC). Methylisocitrate lyase, the last step of the MCC, is intrinsically downregulated in BCG, leading to obstruction of carbon flux toward central carbon metabolism, accumulation of MCC intermediates, and interference with GltB/D mediated neutralizing activity against propionate toxicity. Indeed, vitamin B12 mediated bypass MCC and additional supplement of glutamate led to selectively correct the phenotypic attenuation in BCG and restore the adaptive capacity of BCG to the similar level of Mtb phenotype. Collectively, a defective crosstalk between MCC and Q → E contributes to attenuation of intracellular BCG. Furthermore, GltB/D inhibition enhances the level of propionate toxicity in Mtb. Thus, these findings revealed a new adaptive metabolism and propose GltB/D as a synergistic target to improve the antimicrobial outcomes of MCC inhibition in Mtb

Systematic Genetic Nomenclature for Type VII Secretion Systems

CITATION: Bitter, W., et al. 2009. Systematic genetic nomenclature for type VII secretion systems. PLoS Pathogens, 5(10): 1-6, doi: 10.1371/journal.ppat.1000507.The original publication is available at http://journals.plos.org/plospathogensMycobacteria, such as the etiological agent of human tuberculosis, Mycobacterium tuberculosis, are protected by an impermeable cell envelope composed of an inner cytoplasmic membrane, a peptidoglycan layer, an arabinogalactan layer, and an outer membrane. This second membrane consists of covalently linked, tightly packed long-chain mycolic acids [1,2] and noncovalently bound shorter lipids involved in pathogenicity [3–5]. To ensure protein transport across this complex cell envelope, mycobacteria use various secretion pathways, such as the SecA1-mediated general secretory pathway [6,7], an alternative SecA2-operated pathway [8], a twin-arginine translocation system [9,10], and a specialized secretion pathway variously named ESAT-6-, SNM-, ESX-, or type VII secretion [11–16]. The latter pathway, hereafter referred to as type VII secretion (T7S), has recently become a large and competitive research topic that is closely linked to studies of host–pathogen interactions of M. tuberculosis [17] and other pathogenic mycobacteria [16]. Molecular details are just beginning to be revealed [18–22] showing that T7S systems are complex machineries with multiple components and multiple substrates. Despite their biological importance, there has been a lack of a clear naming policy for the components and substrates of these systems. As there are multiple paralogous T7S systems within the Mycobacteria and orthologous systems in related bacteria, we are concerned that, without a unified nomenclature system, a multitude of redundant and obscure gene names will be used that will inevitably lead to confusion and hinder future progress. In this opinion piece we will therefore propose and introduce a systematic nomenclature with guidelines for name selection of new components that will greatly facilitate communication and understanding in this rapidly developing field of research.http://journals.plos.org/plospathogens/article?id=10.1371%2Fjournal.ppat.1000507Publisher's versio

New approaches in the diagnosis and treatment of latent tuberculosis infection

With nearly 9 million new active disease cases and 2 million deaths occurring worldwide every year, tuberculosis continues to remain a major public health problem. Exposure to Mycobacterium tuberculosis leads to active disease in only ~10% people. An effective immune response in remaining individuals stops M. tuberculosis multiplication. However, the pathogen is completely eradicated in ~10% people while others only succeed in containment of infection as some bacilli escape killing and remain in non-replicating (dormant) state (latent tuberculosis infection) in old lesions. The dormant bacilli can resuscitate and cause active disease if a disruption of immune response occurs. Nearly one-third of world population is latently infected with M. tuberculosis and 5%-10% of infected individuals will develop active disease during their life time. However, the risk of developing active disease is greatly increased (5%-15% every year and ~50% over lifetime) by human immunodeficiency virus-coinfection. While active transmission is a significant contributor of active disease cases in high tuberculosis burden countries, most active disease cases in low tuberculosis incidence countries arise from this pool of latently infected individuals. A positive tuberculin skin test or a more recent and specific interferon-gamma release assay in a person without overt signs of active disease indicates latent tuberculosis infection. Two commercial interferon-gamma release assays, QFT-G-IT and T-SPOT.TB have been developed. The standard treatment for latent tuberculosis infection is daily therapy with isoniazid for nine months. Other options include therapy with rifampicin for 4 months or isoniazid + rifampicin for 3 months or rifampicin + pyrazinamide for 2 months or isoniazid + rifapentine for 3 months. Identification of latently infected individuals and their treatment has lowered tuberculosis incidence in rich, advanced countries. Similar approaches also hold great promise for other countries with low-intermediate rates of tuberculosis incidence