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1.  Functional Activity of Monocytes and Macrophages in HTLV-1 Infected Subjects 
The Human T lymphotropic virus type-1 (HTLV-1) infects predominantly T cells, inducing proliferation and lymphocyte activation. Additionally, HTLV-1 infected subjects are more susceptible to other infections caused by other intracellular agents. Monocytes/macrophages are important cells in the defense against intracellular pathogens. Our aims were to determine the frequency of monocytes subsets, expression of co-stimulatory molecules in these cells and to evaluate microbicidal ability and cytokine and chemokine production by macrophages from HTLV-1 infected subjects. Participants were 23 HTLV-1 carriers (HC), 22 HAM/TSP patients and 22 healthy subjects (HS) not infected with HTLV-1. The frequencies of monocyte subsets and expression of co-stimulatory molecules were determined by flow cytometry. Macrophages were infected with L. braziliensis or stimulated with LPS. Microbicidal activity of macrophages was determined by optic microscopy. Cytokines/chemokines from macrophage supernatants were measured by ELISA. HAM/TSP patients showed an increase frequency of intermediate monocytes, but expression of co-stimulatory molecules was similar between the groups. Macrophages from HTLV-1 infected individuals were infected with L. braziliensis at the same ratio than macrophages from HS, and all the groups had the same ability to kill Leishmania parasites. However, macrophages from HTLV-1 infected subjects produced more CXCL9 and CCL5, and less IL-10 than cells from HS. While there was no correlation between IFN-γ and cytokine/chemokine production by macrophages, there was a correlation between proviral load and TNF and CXCL10. These data showed a dissociation between the inflammatory response and microbicidal ability of macrophages from HTLV-1 infected subjects. While macrophages ability to kill an intracellular pathogen did not differ among HTLV-1 infected subjects, these cells secreted high amount of chemokines even in unstimulated cultures. Moreover the increasing inflammatory activity of macrophages was similar in HAM/TSP patients and HC and it was related to HTLV-1 proviral load rather than the high IFN-γ production observed in these subjects.
Author Summary
HTLV-1 predominantly infects T cells, inducing cell proliferation and activation. While there is a larger amount of studies regarding T cells functions in HTLV-1 infected subjects, little is known about innate immunity. We evaluated monocyte and macrophage functions in HTLV-1 infected subjects. We observed that HAM/TSP patients have an increased frequency of intermediate monocytes, but expression of co-stimulatory molecules in these cells was similar between HTLV-1 infected subjects and healthy subjects (HS). Additionally, the microbicidal ability of macrophages from HTLV-1 infected subjects to kill Leishmania braziliensis is preserved, and these cells showed inflammatory profile, producing more CXCL9 and CCL5, and less IL-10 than macrophages from HS. It was important to determine if the exacerbated ability of macrophages to secrete cytokine was due to IFN-γ production. While there was no correlation between IFN-γ levels by PBMCs and cytokine/chemokine production by macrophages, there was a direct correlation between proviral load and TNF and CXCL10 levels. Our data indicate that despite the high production of proinflammatory mediators, macrophages from HTLV-1 infected subjects kill an intracellular pathogen in similar levels than cells from HS and pointed out for the role of viral factors inducing the inflammatory response in these cells.
doi:10.1371/journal.pntd.0003399
PMCID: PMC4270688  PMID: 25521499
2.  Broad Adaptive Immune Responses to M. tuberculosis Antigens Precede TST Conversion in Tuberculosis Exposed Household Contacts in a TB-Endemic Setting 
PLoS ONE  2014;9(12):e116268.
Background
The identification of Mycobacterium-tuberculosis (Mtb) infected individuals remains a challenge due to an insufficient understanding of immune responses detected with the current diagnostic tests for latent tuberculosis i.e. the tuberculin skin test (TST) or IFN–γ release assays (IGRAs) and an inability to distinguish infection stages with current immunologic assays. Further classification based on markers other than IFN–γ may help to define markers of early Mtb infection.
Methods
We assessed the TST status of Mtb-exposed household contacts at baseline and at 6 months. Contacts were classified into those with initial positive TST (TST+); those with baseline negative TST but TST conversion at 6 months (TST converters, TSTC) and those with persistently negative TST (PTST−). We assessed their short- and long-term immune responses to PPD and ESAT–6/CFP–10 (EC) via IFN–γ ELISPOT and a multiplex cytokine array in relation to TST status and compared them to those of TB cases to identify immune profiles associated with a spectrum of infection stages.
Results
After 1 and 6 days stimulation with EC, 12 cytokines (IFN–γ, IL–2, IP–10, TNF–α, IL–13, IL–17, IL–10, GMCSF, MIP–1β, MCP–3, IL–2RA and IL–1A) were not different in TSTC compared to TST+ suggesting that robust adaptive Mtb-specific immune responses precede TST conversion. Stratifying contacts by baseline IFN–γ ELISPOT to EC in combination with TST results revealed that IP–10 and IL–17 were highest in the group of TST converters with positive baseline ELISPOT, suggesting they might be markers for recent infection.
Conclusion
We describe a detailed analysis of Mtb-specific biomarker profiles in exposed household contacts in a TB endemic area that provides insights into the dynamic immune responses to Mtb infection and may help to identify biomarkers for ‘at-risk’ populations beyond TST and IGRA.
doi:10.1371/journal.pone.0116268
PMCID: PMC4280211  PMID: 25549338
3.  Evaluation of Gamma Interferon Release Assays Using Mycobacterium tuberculosis Antigens for Diagnosis of Latent and Active Tuberculosis in Mycobacterium bovis BCG-Vaccinated Populations▿  
Clinical and Vaccine Immunology : CVI  2010;17(12):1985-1990.
T-cell-based gamma interferon (IFN-γ) release assays (IGRAs) using Mycobacterium tuberculosis-specific antigens have shown higher sensitivity and specificity than the routine tuberculin skin test (TST). However, the effects of Mycobacterium bovis BCG vaccination and anti-tuberculosis (TB) treatment on dynamic T-cell responses to M. tuberculosis-specific antigens in active TB cases have rarely been investigated in regions where TB is endemic. Eighty-nine patients with active pulmonary TB (ATB) and 57 healthy controls (HC) from China were recruited and tested by sputum smear and culture, TSTs, and IGRAs with M. tuberculosis-specific antigens ESAT-6 and CFP-10 (T-SPOT.TB) as well as purified protein derivative (PPD) stimulation. All 146 participants were screened by the T-SPOT.TB assay at recruitment. T-SPOT.TB-positive rates in ATB and HC groups were 87.6% (78/89) and 21.1% (12/57), respectively. Of 38 ATB patients who were both TST and T-SPOT.TB tested, the positive rates were 73.7% (28/38) and 94.7% (36/38), respectively (P = 0.0215), and those in the HC group were 62.3% (33/53) and 18.9% (10/53), respectively (P < 0.0001). The T-SPOT.TB-positive rates declined during TB treatment and were 94.4% (51/54), 86.4% (19/22), and 61.5% (8/13) for ATB patients receiving 0- to 1-month, 1- to 3-month, and 3- to 6-month anti-TB treatment, respectively. The IGRA is a most promising test for both active TB and latent TB infection (LTBI) diagnosis due to the improvement of its specificity and convenience, especially in the Mycobacterium bovis BCG-vaccinated population. Furthermore, the T-SPOT.TB assay using ESAT-6 and CFP-10 in ATB patients during anti-TB treatment could serve as a potential predictor of therapeutic efficacy.
doi:10.1128/CVI.00294-10
PMCID: PMC3008201  PMID: 20943878
4.  Regulatory T Cell Expansion in HTLV-1 and Strongyloidiasis Co-infection Is Associated with Reduced IL-5 Responses to Strongyloides stercoralis Antigen 
Background
Human strongyloidiasis varies from a chronic but limited infection in normal hosts to hyperinfection in patients treated with corticosteroids or with HTLV-1 co-infection. Regulatory T cells dampen immune responses to infections. How human strongyloidiasis is controlled and how HTLV-1 infection affects this control are not clear. We hypothesize that HTLV-1 leads to dissemination of Strongyloides stercoralis infection by augmenting regulatory T cell numbers, which in turn down regulate the immune response to the parasite.
Objective
To measure peripheral blood T regulatory cells and Strongyloides stercoralis larval antigen-specific cytokine responses in strongyloidiasis patients with or without HTLV-1 co-infection.
Methods
Peripheral blood mononuclear cells (PBMCs) were isolated from newly diagnosed strongyloidiasis patients with or without HTLV-1 co-infection. Regulatory T cells were characterized by flow cytometry using intracellular staining for CD4, CD25 and FoxP3. PBMCs were also cultured with and without Strongyloides larval antigens. Supernatants were analyzed for IL-5 production.
Results
Patients with HTLV-1 and Strongyloides co-infection had higher parasite burdens. Eosinophil counts were decreased in the HTLV-1 and Strongyloides co-infected subjects compared to strongyloidiasis-only patients (70.0 vs. 502.5 cells/mm3, p = 0.09, Mann-Whitney test). The proportion of regulatory T cells was increased in HTLV-1 positive subjects co-infected with strongyloidiasis compared to patients with only strongyloidiasis or asymptomatic HTLV-1 carriers (median = 17.9% vs. 4.3% vs. 5.9 p<0.05, One-way ANOVA). Strongyloides antigen-specific IL-5 responses were reduced in strongyloidiasis/HTLV-1 co-infected patients (5.0 vs. 187.5 pg/ml, p = 0.03, Mann-Whitney test). Reduced IL-5 responses and eosinophil counts were inversely correlated to the number of CD4+CD25+FoxP3+ cells.
Conclusions
Regulatory T cell counts are increased in patients with HTLV-1 and Strongyloides stercoralis co-infection and correlate with both low circulating eosinophil counts and reduced antigen-driven IL-5 production. These findings suggest a role for regulatory T cells in susceptibility to Strongyloides hyperinfection.
Author Summary
Human strongyloidiasis varies from a mild, controlled infection to a severe frequently fatal disseminated infection depending on the hosts. Patients infected with the retrovirus HTLV-1 have more frequent and more severe forms of strongyloidiasis. It is not clear how human strongyloidiasis is controlled by the immune system and how HTLV-1 infection affects this control. We hypothesize that HTLV-1 leads to dissemination of Strongyloides stercoralis by augmenting regulatory T cell numbers, which in turn down regulate the immune response to the parasite. In our study, patients with HTLV-1 and Strongyloides co-infection had higher parasite burdens than patients with only strongyloidiasis. Eosinophils play an essential role in control of strongyloidiasis in animal models, and eosinophil counts were decreased in the HTLV-1 and Strongyloides stercoralis co-infected subjects compared to patients with only strongyloidiasis. The proportion of T cells with a regulatory cell phenotype was increased in HTLV-1 positive subjects co-infected with strongyloidiasis compared to patients with only strongyloidiasis. IL-5 is a key host molecule in stimulating eosinophil production and activation, and Strongyloides stercoralis antigen-specific IL-5 responses were reduced in strongyloidiasis/HTLV-1 co-infected patients. Reduced IL-5 responses and eosinophil counts were inversely correlated to the number of regulatory T cells. These findings suggest a role for regulatory T cells in susceptibility to Strongyloides hyperinfection.
doi:10.1371/journal.pntd.0000456
PMCID: PMC2686100  PMID: 19513105
5.  PREVALENCE OF HTLV-1 INFECTION IN HOSPITALIZED PATIENTS WITH TUBERCULOSIS 
SUMMARY
Objective
A few reports have suggested that HTLV-1 may influence immunological response and therefore, clinical course of tuberculosis in co-infected individuals. We wished to determine the prevalence of HTLV-1 infection among hospitalized patients in Salvador, Brazil, a region endemic for both HTLV-1 infection and latent tuberculosis infection.
Design
A cross-sectional study was conducted at a pulmonary disease hospital between September 1st of 2006 to August 31st of 2007. Study participants were interviewed and tested for HTLV-1 infection and current or past episode of tuberculosis.
Results
Of 607 participants recruited into the study, 360 (59.3%) had current or past history of tuberculosis and 50 (8.2%) had HTLV-1 infection; 39 (6.4%) had both. After controlling for confounding variables, we found that the odds of patients with a positive HTLV-1 test having tuberculosis were 2.57 times the odds (95%, CI: 1,23, 5:35) in those who tested negative for HTLV-1 infection.
Conclusion
In a region endemic for both tuberculosis and HTLV-1 infection, HTLV-1 infection increases the risk of Mycobacterium tuberculosis infection. Such a risk may influence tuberculosis transmission and therefore epidemiology of the disease in this community.
PMCID: PMC2963180  PMID: 19919770
HTLV-1; Tuberculosis; Mycobacterium tuberculosis
6.  Serological Testing Versus Other Strategies for Diagnosis of Active Tuberculosis in India: A Cost-Effectiveness Analysis 
PLoS Medicine  2011;8(8):e1001074.
This cost-effectiveness study shows that sputum smear microscopy is the most cost-effective test for active tuberculosis (TB) in India, and liquid culture plus microscopy is more cost-effective for TB diagnosis than serological tests.
Background
Undiagnosed and misdiagnosed tuberculosis (TB) drives the epidemic in India. Serological (antibody detection) TB tests are not recommended by any agency, but widely used in many countries, including the Indian private sector. The cost and impact of using serology compared with other diagnostic techniques is unknown.
Methods and Findings
Taking a patient cohort conservatively equal to the annual number of serological tests done in India (1.5 million adults suspected of having active TB), we used decision analysis to estimate costs and effectiveness of sputum smear microscopy (US$3.62 for two smears), microscopy plus automated liquid culture (mycobacterium growth indicator tube [MGIT], US$20/test), and serological testing (anda-tb ELISA, US$20/test). Data on test accuracy and costs were obtained from published literature. We adopted the perspective of the Indian TB control sector and an analysis frame of 1 year. Our primary outcome was the incremental cost per disability-adjusted life year (DALY) averted. We performed one-way sensitivity analysis on all model parameters, with multiway sensitivity analysis on variables to which the model was most sensitive.
If used instead of sputum microscopy, serology generated an estimated 14,000 more TB diagnoses, but also 121,000 more false-positive diagnoses, 102,000 fewer DALYs averted, and 32,000 more secondary TB cases than microscopy, at approximately four times the incremental cost (US$47.5 million versus US$11.9 million). When added to high-quality sputum smears, MGIT culture was estimated to avert 130,000 incremental DALYs at an incremental cost of US$213 per DALY averted. Serology was dominated by (i.e., more costly and less effective than) MGIT culture and remained less economically favorable than sputum smear or TB culture in one-way and multiway sensitivity analyses.
Conclusions
In India, sputum smear microscopy remains the most cost-effective diagnostic test available for active TB; efforts to increase access to quality-assured microscopy should take priority. In areas where high-quality microscopy exists and resources are sufficient, MGIT culture is more cost-effective than serology as an additional diagnostic test for TB. These data informed a recently published World Health Organization policy statement against serological tests.
Please see later in the article for the Editors' Summary
Editors' Summary
Background
Every year, about 2 million people develop tuberculosis in India—a fifth of the global incidence of this highly contagious bacterial infection. Mycobacterium tuberculosis, the bacterium that causes tuberculosis, is spread in airborne droplets when people with the disease cough or sneeze and usually infects the lungs although it can also infect other organs. The characteristic symptoms of tuberculosis are a persistent cough, weight loss, and night sweats. Diagnostic tests for tuberculosis include sputum smear microscopy (microscopic analysis of mucus brought up from the lungs by coughing), culture (growth) of M. tuberculosis from sputum samples in liquid media (using, for example, a commercial product called the mycobacteria growth indicator tube or MGIT), and nucleic acid amplification tests (which detect the bacterium's genome in patient samples) such as the Xpert MTB/RIF system. Tuberculosis can usually be cured by taking several powerful antibiotics daily for at least 6 months.
Why Was This Study Done?
In India, as elsewhere, undiagnosed and misdiagnosed tuberculosis drives the tuberculosis epidemic by increasing the transmission of M. tuberculosis. Unfortunately, sputum smear microscopy, the current mainstay of tuberculosis diagnosis worldwide, detects only half of tuberculosis cases, mycobacterial culture can take weeks to provide a diagnosis, and rapid techniques such as nucleic acid amplification require infrastructure that is often not available in developing countries. Consequently, in India and other developing countries, serological tests are widely used for the diagnosis of tuberculosis. Serological tests detect antibodies against M. tuberculosis in the blood (antibodies are proteins made by the immune system in response to infections). Serological tests are fast and simple to perform, but they are not recommended for clinical use, and the available evidence suggests that they do not diagnose tuberculosis accurately. Even so, and in the absence of information about the cost and impact (cost-effectiveness) of serological testing, about 1.5 million serological tests for tuberculosis are conducted every year in India at a cost of more than US$15 million. Here, the researchers analyze the cost-effectiveness of serological tests compared to other diagnostic tests from the perspective of tuberculosis control in India.
What Did the Researchers Do and Find?
The researchers used “decision analysis” to estimate the cost-effectiveness of sputum smear microscopy, microscopy plus liquid culture using the MGIT system, and serological testing using the widely used anda-tb ELISA commercial test in a hypothetical group of 1.5 million people suspected of having tuberculosis. Decision analysis formally assesses the decision-making process by using models that evaluate outcomes under different scenarios. By feeding data on the costs and accuracy of different diagnostic tests into their decision-analysis model, the researchers estimate that, over a year, serology would generate 14,000 more tuberculosis diagnoses than sputum microscopy. However, it would also generate 121,000 more false-positive diagnoses and 32,000 more tuberculosis transmissions to other people (secondary transmissions), and avert 102,000 fewer disability-adjusted life years (DALYs; a DALY is a year of healthy life lost because of premature death or disability) at four times the incremental cost of sputum microscopy. MGIT culture added to sputum smear microscopy would avert 130,000 DALYs at an incremental cost of US$213 per DALY averted. Finally, sensitivity analyses (reruns of the decision-analysis model using different values for test costs and accuracy) identified no scenario in which serology was either less costly or more effective than sputum smear microscopy alone or in which serology plus sputum microscopy was more cost-effective than MGIT culture plus sputum microscopy.
What Do These Findings Mean?
These findings identify sputum smear microscopy as the most cost-effective existing diagnostic test for tuberculosis in India. Moreover, they suggest that in areas where high-quality microscopy is available, resources are sufficient, and infrastructure to effectively use culture exists, the addition of MGIT culture to sputum smear microscopy would be more cost-effective than the addition of serology. Importantly, these findings suggest that, if used as an initial test for tuberculosis in India, serology would result in more DALYs, more secondary infections, and more false-positive diagnoses than sputum smear microscopy while increasing per-patient costs to the Indian tuberculosis control sector. Given these findings and the results of a recent updated systematic review on the accuracy of serological tests, the World Health Organization's Strategic and Technical Advisory Group for Tuberculosis recently advised against the use of currently available serological tests for the diagnosis of tuberculosis. The WHO negative policy against serological tests must now be implemented in India.
Additional Information
Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001074.
Details of the recent systematic review of serological tests for tuberculosis diagnosis are available in a PLoS Medicine Research Article by Steingart et al.
The World Health Organization provides information on all aspects of tuberculosis, including tuberculosis diagnostics and the Stop TB Partnership (some information is in several languages); its Strategic and Technical Advisory Group for Tuberculosis recommendations on tuberculosis diagnosis are available
The Evidence-based TB Diagnosis Web site by the Stop TB Partnership's New Diagnostics Working Group provides evidence syntheses on various TB tests, along with guidelines, resources, and training materials
The US Centers for Disease Control and Prevention has information about tuberculosis, including information on the diagnosis of tuberculosis disease
The US National Institute of Allergy and Infectious Diseases also has information on all aspects of tuberculosis
MedlinePlus has links to further information about tuberculosis (in English and Spanish)
doi:10.1371/journal.pmed.1001074
PMCID: PMC3153451  PMID: 21857810
7.  Immunological Profile of HTLV-1-Infected Patients Associated with Infectious or Autoimmune Dermatological Disorders 
In the present study, the frequency, the activation and the cytokine and chemokine profile of HTLV-1 carriers with or without dermatological lesions were thoroughly described and compared. The results indicated that HTLV-1-infected patients with dermatological lesions have distinct frequency and activation status when compared to asymptomatic carriers. Alterations in the CD4+HLA-DR+, CD8+ T cell, macrophage-like and NKT subsets as well as in the serum chemokines CCL5, CXCL8, CXCL9 and CXCL10 were observed in the HTLV-1-infected group with skin lesions. Additionally, HTLV-1 carriers with dermatological skin lesions showed more frequently high proviral load as compared to asymptomatic carriers. The elevated proviral load in HTLV-1 patients with infectious skin lesions correlated significantly with TNF-α/IL-10 ratio, while the same significant correlation was found for the IL-12/IL-10 ratio and the high proviral load in HTLV-1-infected patients with autoimmune skin lesions. All in all, these results suggest a distinct and unique immunological profile in the peripheral blood of HTLV-1-infected patients with skin disorders, and the different nature of skin lesion observed in these patients may be an outcome of a distinct unbalance of the systemic inflammatory response upon HTLV-1 infection.
Author Summary
In the present study, the immunological profiles of HTLV-1 carriers with or without dermatological lesions were thoroughly described and compared. The results indicated that HTLV-1-infected patients with dermatological lesions have distinct frequency and activation status than asymptomatic carriers. Alterations in cells and molecules that are important for immune cell function were observed in the HTLV-1-infected group with skin lesions. Additionally, HTLV-1 carriers with dermatological skin lesions have elevated frequency of high proviral load as compared to asymptomatic carriers, which indicates that the virus may be present in higher frequency in those patients. Patients with different skin lesions, autoimmune or infectious, also demonstrated differences in their immunological profile. All in all, these results suggest a distinct and unique immunological profile in the blood of HTLV-1-infected patients with skin disorders, and the different nature of skin lesion observed in these patients may be an outcome of a distinct unbalance of the systemic inflammatory response upon HTLV-1 infection.
doi:10.1371/journal.pntd.0002328
PMCID: PMC3723575  PMID: 23936564
8.  Genetic Characterization of Human T-Cell Lymphotropic Virus Type 1 in Mozambique: Transcontinental Lineages Drive the HTLV-1 Endemic 
Background
Human T-Cell Lymphotropic Virus Type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). It has been estimated that 10–20 million people are infected worldwide, but no successful treatment is available. Recently, the epidemiology of this virus was addressed in blood donors from Maputo, showing rates from 0.9 to 1.2%. However, the origin and impact of HTLV endemic in this population is unknown.
Objective
To assess the HTLV-1 molecular epidemiology in Mozambique and to investigate their relationship with HTLV-1 lineages circulating worldwide.
Methods
Blood donors and HIV patients were screened for HTLV antibodies by using enzyme immunoassay, followed by Western Blot. PCR and sequencing of HTLV-1 LTR region were applied and genetic HTLV-1 subtypes were assigned by the neighbor-joining method. The mean genetic distance of Mozambican HTLV-1 lineages among the genetic clusters were determined. Human mitochondrial (mt) DNA analysis was performed and individuals classified in mtDNA haplogroups.
Results
LTR HTLV-1 analysis demonstrated that all isolates belong to the Transcontinental subgroup of the Cosmopolitan subtype. Mozambican HTLV-1 sequences had a high inter-strain genetic distance, reflecting in three major clusters. One cluster is associated with the South Africa sequences, one is related with Middle East and India strains and the third is a specific Mozambican cluster. Interestingly, 83.3% of HIV/HTLV-1 co-infection was observed in the Mozambican cluster. The human mtDNA haplotypes revealed that all belong to the African macrohaplogroup L with frequencies representatives of the country.
Conclusions
The Mozambican HTLV-1 genetic diversity detected in this study reveals that although the strains belong to the most prevalent and worldwide distributed Transcontinental subgroup of the Cosmopolitan subtype, there is a high HTLV diversity that could be correlated with at least 3 different HTLV-1 introductions in the country. The significant rate of HTLV-1a/HIV-1C co-infection, particularly in the Mozambican cluster, has important implications for the controls programs of both viruses.
Author Summary
Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of Adult T-Cell Leukemia/Lymphoma (ATL), the Tropical Spastic Paraparesis/HTLV-1-associated Myelopathy (TSP/HAM) and other inflammatory diseases, including dermatitis, uveitis, and myositis. It is estimated that 2–8% of the infected persons will develop a HTLV-1-associated disease during their lifetimes, frequently TSP/HAM. Thus far, there is not a specific treatment to this progressive and chronic disease. HTLV-1 has means of three transmission: (i) from mother to child during prolonged breastfeeding, (ii) between sexual partners and (iii) through blood transfusion. HTLV-1 has been characterized in 7 subtypes and the geographical distribution and the clinical impact of this infection is not well known, mainly in African population. HTLV-1 is endemic in sub-Saharan Africa. Mozambique is a country of southeastern Africa where TSP/HAM cases were reported. Recently, our group estimated the HTLV prevalence among Mozambican blood donors as 0.9%. In this work we performed a genetic analysis of HTLV-1 in blood donors and HIV/HTLV co-infected patients from Maputo, Mozambique. Our results showed the presence of three HTLV-1 clusters within the Cosmopolitan/Transcontinental subtype/subgroup. The differential rates of HIV-1/HTLV-1 co-infection in the three HTLV-1 clusters demonstrated the dynamic of the two viruses and the need for implementation of control measures focusing on both retroviruses.
doi:10.1371/journal.pntd.0001038
PMCID: PMC3075232  PMID: 21532745
9.  HTLV-1 Evades Type I Interferon Antiviral Signaling by Inducing the Suppressor of Cytokine Signaling 1 (SOCS1) 
PLoS Pathogens  2010;6(11):e1001177.
Human T cell leukemia virus type 1 (HTLV-1) is the etiologic agent of Adult T cell Leukemia (ATL) and the neurological disorder HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the majority of HTLV-1–infected individuals remain asymptomatic carriers (AC) during their lifetime, 2–5% will develop either ATL or HAM/TSP, but never both. To better understand the gene expression changes in HTLV-1-associated diseases, we examined the mRNA profiles of CD4+ T cells isolated from 7 ATL, 12 HAM/TSP, 11 AC and 8 non-infected controls. Using genomic approaches followed by bioinformatic analysis, we identified gene expression pattern characteristic of HTLV-1 infected individuals and particular disease states. Of particular interest, the suppressor of cytokine signaling 1—SOCS1—was upregulated in HAM/TSP and AC patients but not in ATL. Moreover, SOCS1 was positively correlated with the expression of HTLV-1 mRNA in HAM/TSP patient samples. In primary PBMCs transfected with a HTLV-1 proviral clone and in HTLV-1-transformed MT-2 cells, HTLV-1 replication correlated with induction of SOCS1 and inhibition of IFN-α/β and IFN-stimulated gene expression. Targeting SOCS1 with siRNA restored type I IFN production and reduced HTLV-1 replication in MT-2 cells. Conversely, exogenous expression of SOCS1 resulted in enhanced HTLV-1 mRNA synthesis. In addition to inhibiting signaling downstream of the IFN receptor, SOCS1 inhibited IFN-β production by targeting IRF3 for ubiquitination and proteasomal degradation. These observations identify a novel SOCS1 driven mechanism of evasion of the type I IFN antiviral response against HTLV-1.
Author Summary
Infection with HTLV-1 leads to the development of Adult T cell Leukemia (ATL) or the neurological disorder HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Although the majority of HTLV-1–infected individuals remain asymptomatic carriers (AC) during their lifetime, 2–5% will develop either ATL or HAM/TSP. Using gene expression profiling of CD4+ T lymphocytes from HTLV-1 infected patients, we identified Suppressor of cytokine signaling 1 (SOCS1) as being highly expressed in HAM/TSP and AC patients. SOCS1 expression positively correlated with the high HTLV-1 mRNA load that is characteristic of HAM/TSP patients. SOCS1 inhibited cellular antiviral signaling during HTLV-1 infection by degrading IRF3, an essential transcription factor in the interferon pathway. Our study reveals a novel evasion mechanism utilized by HTLV-1 that leads to increased retroviral replication, without triggering the innate immune response.
doi:10.1371/journal.ppat.1001177
PMCID: PMC2973829  PMID: 21079688
10.  Latent tuberculosis infection, tuberculin skin test and vitamin D status in contacts of tuberculosis patients: a cross-sectional and case-control study 
BMC Infectious Diseases  2011;11:349.
Background
Deficient serum vitamin D levels have been associated with incidence of tuberculosis (TB), and latent tuberculosis infection (LTBI). However, to our knowledge, no studies on vitamin D status and tuberculin skin test (TST) conversion have been published to date. The aim of this study was to estimate the associations of serum 25-hydroxyvitamin D3 (25[OH]D) status with LTBI prevalence and TST conversion in contacts of active TB in Castellon (Spain).
Methods
The study was designed in two phases: cross-sectional and case-control. From November 2009 to October 2010, contacts of 42 TB patients (36 pulmonary, and 6 extra-pulmonary) were studied in order to screen for TB. LTBI and TST conversion cases were defined following TST, clinical, analytic and radiographic examinations. Serum 25(OH)D levels were measured by electrochemiluminescence immunoassay (ECLIA) on a COBAS® 410 ROCHE® analyzer. Logistic regression models were used in the statistical analysis.
Results
The study comprised 202 people with a participation rate of 60.1%. Only 20.3% of the participants had a sufficient serum 25(OH)D (≥ 30 ng/ml) level. In the cross-sectional phase, 50 participants had LTBI and no association between LTBI status and serum 25(OH)D was found. After 2 months, 11 out of 93 negative LTBI participants, without primary prophylaxis, presented TST conversion with initial serum 25(OH)D levels: a:19.4% (7/36): < 20 ng/ml, b:12.5% (4/32):20-29 ng/ml, and c:0%(0/25) ≥ 30 ng/ml. A sufficient serum 25(OH)D level was a protector against TST conversion a: Odds Ratio (OR) = 1.00; b: OR = 0.49 (95% confidence interval (CI) 0.07-2.66); and c: OR = 0.10 (95% CI 0.00-0.76), trends p = 0.019, adjusted for high exposure and sputum acid-fast bacilli positive index cases. The mean of serum level 25(OH)D in TST conversion cases was lower than controls,17.5 ± 5.6 ng/ml versus 25.9 ± 13.7 ng/ml (p = 0.041).
Conclusions
The results suggest that sufficient serum 25(OH)D levels protect against TST conversion.
doi:10.1186/1471-2334-11-349
PMCID: PMC3292546  PMID: 22171844
Tuberculosis; Vitamin D; Latent tuberculosis infection; Tuberculin skin test conversion; Case-control study
11.  Functional Signatures of Human CD4 and CD8 T Cell Responses to Mycobacterium tuberculosis 
With 1.4 million deaths and 8.7 million new cases in 2011, tuberculosis (TB) remains a global health care problem and together with HIV and Malaria represents one of the three infectious diseases world-wide. Control of the global TB epidemic has been impaired by the lack of an effective vaccine, by the emergence of drug-resistant forms of Mycobacterium tuberculosis (Mtb) and by the lack of sensitive and rapid diagnostics. It is estimated, by epidemiological reports, that one third of the world’s population is latently infected with Mtb, but the majority of infected individuals develop long-lived protective immunity, which controls and contains Mtb in a T cell-dependent manner. Development of TB disease results from interactions among the environment, the host, and the pathogen, and known risk factors include HIV co-infection, immunodeficiency, diabetes mellitus, overcrowding, malnutrition, and general poverty; therefore, an effective T cell response determines whether the infection resolves or develops into clinically evident disease. Consequently, there is great interest in determining which T cells subsets mediate anti-mycobacterial immunity, delineating their effector functions. On the other hand, many aspects remain unsolved in understanding why some individuals are protected from Mtb infection while others go on to develop disease. Several studies have demonstrated that CD4+ T cells are involved in protection against Mtb, as supported by the evidence that CD4+ T cell depletion is responsible for Mtb reactivation in HIV-infected individuals. There are many subsets of CD4+ T cells, such as T-helper 1 (Th1), Th2, Th17, and regulatory T cells (Tregs), and all these subsets co-operate or interfere with each other to control infection; the dominant subset may differ between active and latent Mtb infection cases. Mtb-specific-CD4+ Th1 cell response is considered to have a protective role for the ability to produce cytokines such as IFN-γ or TNF-α that contribute to the recruitment and activation of innate immune cells, like monocytes and granulocytes. Thus, while other antigen (Ag)-specific T cells such as CD8+ T cells, natural killer (NK) cells, γδ T cells, and CD1-restricted T cells can also produce IFN-γ during Mtb infection, they cannot compensate for the lack of CD4+ T cells. The detection of Ag-specific cytokine production by intracellular cytokine staining (ICS) and the use of flow cytometry techniques are a common routine that supports the studies aimed at focusing the role of the immune system in infectious diseases. Flow cytometry permits to evaluate simultaneously the presence of different cytokines that can delineate different subsets of cells as having “multifunctional/polyfunctional” profile. It has been proposed that polyfunctional T cells, are associated with protective immunity toward Mtb, in particular it has been highlighted that the number of Mtb-specific T cells producing a combination of IFN-γ, IL-2, and/or TNF-α may be correlated with the mycobacterial load, while other studies have associated the presence of this particular functional profile as marker of TB disease activity. Although the role of CD8 T cells in TB is less clear than CD4 T cells, they are generally considered to contribute to optimal immunity and protection. CD8 T cells possess a number of anti-microbial effector mechanisms that are less prominent or absent in CD4 Th1 and Th17 T cells. The interest in studying CD8 T cells that are either MHC-class Ia or MHC-class Ib-restricted, has gained more attention. These studies include the role of HLA-E-restricted cells, lung mucosal-associated invariant T-cells (MAIT), and CD1-restricted cells. Nevertheless, the knowledge about the role of CD8+ T cells in Mtb infection is relatively new and recent studies have delineated that CD8 T cells, which display a functional profile termed “multifunctional,” can be a better marker of protection in TB than CD4+ T cells. Their effector mechanisms could contribute to control Mtb infection, as upon activation, CD8 T cells release cytokines or cytotoxic molecules, which cause apoptosis of target cells. Taken together, the balance of the immune response in the control of infection and possibly bacterial eradication is important in understanding whether the host immune response will be appropriate in contrasting the infection or not, and, consequently, the inability of the immune response, will determine the dissemination and the transmission of bacilli to new subjects. In conclusion, the recent highlights on the role of different functional signatures of T cell subsets in the immune response toward Mtb infection will be discerned in this review, in order to summarize what is known about the immune response in human TB. In particular, we will discuss the role of CD4 and CD8 T cells in contrasting the advance of the intracellular pathogen in already infected people or the progression to active disease in subjects with latent infection. All the information will be aimed at increasing the knowledge of this complex disease in order to improve diagnosis, prognosis, drug treatment, and vaccination.
doi:10.3389/fimmu.2014.00180
PMCID: PMC4001014  PMID: 24795723
M. tuberculosis; cytokines; human memory T cells; disease; infection
12.  Comparison of Xpert MTB/RIF with Other Nucleic Acid Technologies for Diagnosing Pulmonary Tuberculosis in a High HIV Prevalence Setting: A Prospective Study 
PLoS Medicine  2011;8(7):e1001061.
In this prospective, real-world cohort study nested within a national screening program for tuberculosis, Lesley Scott and colleagues compare the performance of Xpert MTB/RIF on a single sputum sample with different TB sputum detection technologies.
Background
The Xpert MTB/RIF (Cepheid) non-laboratory-based molecular assay has potential to improve the diagnosis of tuberculosis (TB), especially in HIV-infected populations, through increased sensitivity, reduced turnaround time (2 h), and immediate identification of rifampicin (RIF) resistance. In a prospective clinical validation study we compared the performance of Xpert MTB/RIF, MTBDRplus (Hain Lifescience), LightCycler Mycobacterium Detection (LCTB) (Roche), with acid fast bacilli (AFB) smear microscopy and liquid culture on a single sputum specimen.
Methods and Findings
Consecutive adults with suspected TB attending a primary health care clinic in Johannesburg, South Africa, were prospectively enrolled and evaluated for TB according to the guidelines of the National TB Control Programme, including assessment for smear-negative TB by chest X-ray, clinical evaluation, and HIV testing. A single sputum sample underwent routine decontamination, AFB smear microscopy, liquid culture, and phenotypic drug susceptibility testing. Residual sample was batched for molecular testing. For the 311 participants, the HIV prevalence was 70% (n = 215), with 120 (38.5%) culture-positive TB cases. Compared to liquid culture, the sensitivities of all the test methodologies, determined with a limited and potentially underpowered sample size (n = 177), were 59% (47%–71%) for smear microscopy, 76% (64%–85%) for MTBDRplus, 76% (64%–85%) for LCTB, and 86% (76%–93%) for Xpert MTB/RIF, with specificities all >97%. Among HIV+ individuals, the sensitivity of the Xpert MTB/RIF test was 84% (69%–93%), while the other molecular tests had sensitivities reduced by 6%. TB detection among smear-negative, culture-positive samples was 28% (5/18) for MTBDRplus, 22% (4/18) for LCTB, and 61% (11/18) for Xpert MTB/RIF. A few (n = 5) RIF-resistant cases were detected using the phenotypic drug susceptibility testing methodology. Xpert MTB/RIF detected four of these five cases (fifth case not tested) and two additional phenotypically sensitive cases.
Conclusions
The Xpert MTB/RIF test has superior performance for rapid diagnosis of Mycobacterium tuberculosis over existing AFB smear microscopy and other molecular methodologies in an HIV- and TB-endemic region. Its place in the clinical diagnostic algorithm in national health programs needs exploration.
Please see later in the article for the Editors' Summary
Editors' Summary
Background
Tuberculosis (TB)—a contagious bacterial infection that mainly affects the lungs—is a global public health problem. In 2009, 9.4 million people developed TB, and 1.7 million people died from the disease; a quarter of these deaths were in HIV-positive individuals. People who are infected with HIV, the virus that causes AIDS, are particularly susceptible to TB because of their weakened immune system. Consequently, TB is a leading cause of illness and death among people living with HIV. TB is caused by Mycobacterium tuberculosis, which is spread in airborne droplets when people with the disease cough or sneeze. Its characteristic symptoms are a persistent cough, night sweats, and weight loss. Diagnostic tests for TB include sputum smear analysis (the microscopic examination of mucus brought up from the lungs by coughing for the presence of M. tuberculosis) and mycobacterial liquid culture (in which bacteriologists try to grow M. tuberculosis from sputum samples and test its drug sensitivity). TB can usually be cured by taking several powerful drugs daily for at least six months.
Why Was This Study Done?
Mycobacterial culture is a sensitive but slow way to diagnose TB. To halt the disease's spread, it is essential that TB—particularly TB that is resistant to several treatment drugs (multidrug-resistant, or MDR, TB)—is diagnosed quickly. Recently, several nucleic acid amplification technology (NAAT) tests have been developed that rapidly detect M. tuberculosis DNA in patient samples and look for DNA changes that make M. tuberculosis drug-resistant. In December 2010, the World Health Organization (WHO) endorsed Xpert MTB/RIF—an automated DNA test that detects M. tuberculosis and rifampicin resistance (an indicator of MDR TB) within two hours—for the investigation of patients who might have TB, especially in regions where MDR TB and HIV infection are common. TB diagnosis in HIV-positive people can be difficult because they are more likely to have smear-negative TB than HIV-negative individuals. In this prospective study, the researchers compare the performance of Xpert MTB/RIF on a single sputum sample with that of smear microscopy, liquid culture, and two other NAAT tests (MTBDRplus and LightCycler Mycobacterium Detection) in adults who might have TB in Johannesburg (South Africa), a region where many adults are HIV-positive.
What Did the Researchers Do and Find?
The researchers evaluated adults with potential TB attending a primary health care clinic for TB according to national guidelines and determined their HIV status. A sputum sample from 311 participants underwent smear microscopy, liquid culture, and drug susceptibility testing; 177 samples were also tested for TB using NAAT tests. They found that 70% of the participants were HIV-positive and 38.5% had culture-positive TB. Compared to liquid culture, smear microscopy, MTBDRplus, LightCycler Mycobacterium Detection, and Xpert MTB/RIF had sensitivities of 59%, 76%, 76%, and 86%, respectively. That is, assuming that liquid culture detected everyone with TB, Xpert MTB/RIF detected 86% of the cases. The specificity of all the tests compared to liquid culture was greater than 97%. That is, they all had a low false-positive rate. Among people who were HIV-positive, the sensitivity of Xpert MTB/RIF was 84%; the sensitivities of the other NAAT tests were 70%. Moreover, Xpert MTB/RIF detected TB in 61% of smear-negative, culture-positive samples, whereas the other NAATs detected TB in only about a quarter of these samples. Finally, although some TB cases were identified as drug-resistant by one test but drug-sensitive by another, the small number of drug-resistant cases means no firm conclusions can be made about the accuracy of drug resistance determination by the various tests.
What Do These Findings Mean?
Although these findings are likely to be affected by the study's small size, they suggest that Xpert MTB/RIF may provide a more accurate rapid diagnosis of TB than smear microscopy and other currently available NAAT tests in regions where HIV and TB are endemic (i.e., always present). Indeed, the reported accuracy of Xpert MTB/RIF for TB diagnosis—85% sensitivity and 97% specificity—has the potential to save more than 400,000 lives per year. Taken together with the results of other recent studies (including an accompanying article by Lawn et al. that investigates the use of Xpert MTB/RIF for screening for HIV-associated TB and rifampicin resistance), these findings support the WHO recommendation that Xpert MTB/RIF, rather than smear microscopy, should be the initial test in HIV-infected individuals who might have TB.
Additional Information
Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001061.
This study is further discussed in a PLoS Medicine Perspective by Carlton Evans; a related PLoS Medicine Research Article by Lawn et al. is also available
WHO provides information (in several languages) on all aspects of tuberculosis, including general information on tuberculosis diagnostics and specific information on the Xpert MTB/RIF test; further information about WHO's endorsement of Xpert MTB/RIF is included in a recent Strategic and Technical Advisory Group for Tuberculosis report
WHO also provides information about tuberculosis and HIV
The US National Institute of Allergy and Infectious Diseases has detailed information on tuberculosis and HIV/AIDS
The US Centers for Disease Control and Prevention also has information about tuberculosis, including information on the diagnosis of and on tuberculosis and HIV co-infection
Information is available from Avert, an international AIDS charity on many aspects of HIV/AIDS, including information on HIV-related tuberculosis (in English and Spanish)
doi:10.1371/journal.pmed.1001061
PMCID: PMC3144192  PMID: 21814495
13.  What Patient Factors Predict Physicians’ Decision Not to Treat Latent Tuberculosis Infection in Tuberculosis Contacts? 
PLoS ONE  2013;8(9):e76552.
Objective
The study aimed to determine factors that are associated with physicians’ decision to offer treatment for latent tuberculosis infection (LTBI) in contacts of patients with tuberculosis.
Methods
We performed a nested case-control study in a cohort of contacts of patients with pulmonary tuberculosis who had a tuberculin skin test (TST) ≥ 10 mm. Cases were those who were offered treatment for LTBI. Controls were randomly selected from those who were not offered treatment for LTBI by the reviewing physician. Odds ratios were estimated by multivariate logistic regression.
Results
There were 195 cases and 279 controls. The following factors were significantly (positively or negatively) associated with being offered LTBI treatment in the multivariate analysis: female gender (OR 2.9; 95% CI 1.6–5.5), TST conversion (OR 3.9; 2.0–7.9), TST > 20 mm (OR 4.1; 1.8–9.1, for TST of 21–30 mm and OR 7.9; 2.6–23.8, for TST >30 mm), sputum smear positive index case (OR 12.7; 4.5–36.1), being overseas-born and immigration more than 2 years ago (OR 0.1; 0.06–0.3), being a health care worker (OR 0.2; 0.1–0.6), being a non-household contact of the TB index case (OR 0.3; 0.2–0.6) and age >35 years (OR 0.2; 0.1–0.5 for age 35 to 54.9 years and OR 0.04; 0.01–0.2 for age ≥55 years). Previous BCG vaccine and chest x-ray findings were not significantly associated with physicians’ decision to offer treatment for LTBI.
Conclusions
Most factors that influenced physicians’ decisions on treatment for LTBI were based on evidence of an association with risk of developing TB or risk of having an adverse reaction to treatment for LTBI. However, the decreased likelihood of offering treatment for LTBI to people born overseas, men and health care workers, was apparently not based on any evidence of risk. Efforts should be made to ensure that these groups are given access to treatment for LTBI.
doi:10.1371/journal.pone.0076552
PMCID: PMC3786986  PMID: 24098794
14.  Immunological and Viral Features in Patients with Overactive Bladder Associated with Human T-Cell Lymphotropic Virus Type 1 Infection 
Journal of medical virology  2012;84(11):1809-1817.
The majority of patients infected with human T-cell lymphotropic virus-type 1 (HTLV-1) are considered carriers, but a high frequency of urinary symptoms of overactive bladder, common in HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) have been documented in these patients. The aim of this study was to determine if immunological and viral factors that are seen in HAM/TSP are also observed in these patients. Participants were classified as HTLV-1 carriers (n=45), HTLV-1 patients suffering from overactive bladder (n=45) and HAM/TSP (n=45). Cells from HTLV-1 overactive bladder patients produced spontaneously more proinflammatory cytokines than carriers. TNF-α and IL-17 levels were similar in HAM/TSP and HTLV-1 overactive bladder patients. High proviral load was found in patients with overactive bladder and HAM/TSP and correlated with proinflammatory cytokines. In contrast with findings in patients with HAM/TSP, serum levels of Th1 chemokines were similar in HTLV-1 overactive bladder and carriers. Exogenous addition of regulatory cytokines decreased spontaneous IFN-γ production in cell cultures from HTLV-1 overactive bladder patients. The results show that HTLV-1 overactive bladder and HAM/TSP patients have in common some immunological features as well as similar proviral load profile. The data show that HTLV-1 overactive bladder patients are still able to down regulate their inflammatory immune response. In addition, these patients express levels of chemokines similar to carriers, which may explain why they have yet to develop the same degree of spinal cord damage as seen in patients with HAM/TSP. These patients present symptoms of overactive bladder, which may be an early sign of HAM/TSP.
doi:10.1002/jmv.23341
PMCID: PMC3457650  PMID: 22997085
HTLV-1; immune response; cytokines; chemokines; proviral load
15.  Systems Biology Approaches Reveal a Specific Interferon-Inducible Signature in HTLV-1 Associated Myelopathy 
PLoS Pathogens  2012;8(1):e1002480.
Human T-lymphotropic virus type 1 (HTLV-1) is a retrovirus that persists lifelong in the host. In ∼4% of infected people, HTLV-1 causes a chronic disabling neuroinflammatory disease known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The pathogenesis of HAM/TSP is unknown and treatment remains ineffective. We used gene expression microarrays followed by flow cytometric and functional assays to investigate global changes in blood transcriptional profiles of HTLV-1-infected and seronegative individuals. We found that perturbations of the p53 signaling pathway were a hallmark of HTLV-1 infection. In contrast, a subset of interferon (IFN)-stimulated genes was over-expressed in patients with HAM/TSP but not in asymptomatic HTLV-1 carriers or patients with the clinically similar disease multiple sclerosis. The IFN-inducible signature was present in all circulating leukocytes and its intensity correlated with the clinical severity of HAM/TSP. Leukocytes from patients with HAM/TSP were primed to respond strongly to stimulation with exogenous IFN. However, while type I IFN suppressed expression of the HTLV-1 structural protein Gag it failed to suppress the highly immunogenic viral transcriptional transactivator Tax. We conclude that over-expression of a subset of IFN-stimulated genes in chronic HTLV-1 infection does not constitute an efficient host response but instead contributes to the development of HAM/TSP.
Author Summary
Infection with the Human T Lymphotropic virus is widespread in the tropics and subtropics, where it causes a chronic disabling disease of the nervous system abbreviated as HAM/TSP. There is no effective treatment available for HAM/TSP, because it is not understood how the virus causes the neuronal damage that results in the clinical symptoms of weakness and paralysis of the legs. Here, we compared the frequencies of cell populations and gene expression profiles from diseased and asymptomatic HTLV-1 carriers to identify abnormalities in biological pathways that cause HAM/TSP. We discovered a distinct group of genes that is over-expressed in white blood cells in patients with HAM/TSP, but not asymptomatic HTLV-1 carriers or patients with the clinically similar disease multiple sclerosis. The expression of these genes is induced by interferons, a group of anti-viral proteins that are usually beneficial to the host but can also cause inflammation. We also found that interferons did not efficiently suppress HTLV-1 protein expression in vitro. We conclude that interferons do not control chronic HTLV-1 infection but instead contribute to the development of HAM/TSP. Our study provides new insights into the development of HTLV-1-associated diseases and opens new areas of therapeutic intervention.
doi:10.1371/journal.ppat.1002480
PMCID: PMC3266939  PMID: 22291590
16.  High Prevalence of Skin Disorders among HTLV-1 Infected Individuals Independent of Clinical Status 
Background
Human T-cell lymphotropic virus type 1 (HTLV-1) infection can increase the risk of developing skin disorders. This study evaluated the correlation between HTLV-1 proviral load and CD4+ and CD8+ T cells count among HTLV-1 infected individuals, with or without skin disorders (SD) associated with HTLV-1 infection [SD-HTLV-1: xerosis/ichthyosis, seborrheic dermatitis or infective dermatitis associated to HTLV-1 (IDH)].
Methods
A total of 193 HTLV-1-infected subjects underwent an interview, dermatological examination, initial HTLV-1 proviral load assay, CD4+ and CD8+ T cells count, and lymphproliferation assay (LPA).
Results
A total of 147 patients had an abnormal skin condition; 116 (79%) of them also had SD-HTLV-1 and 21% had other dermatological diagnoses. The most prevalent SD-HTLV-1 was xerosis/acquired ichthyosis (48%), followed by seborrheic dermatitis (28%). Patients with SD-HTLV-1 were older (51 vs. 47 years), had a higher prevalence of myelopathy/tropical spastic paraparesis (HAM/TSP) (75%), and had an increased first HTLV-1 proviral load and basal LPA compared with patients without SD-HTLV-1. When excluding HAM/TSP patients, the first HTLV-1 proviral load of SD-HTLV-1 individuals remains higher than no SD-HTLV-1 patients.
Conclusions
There was a high prevalence of skin disorders (76%) among HTLV-1-infected individuals, regardless of clinical status, and 60% of these diseases are considered skin disease associated with HTLV-1 infection.
Author Summary
HTLV-1 infection may increase the risk of developing skin disorders. A total of 193 HTLV-1 infected subjects were studied, including asymptomatic carriers and HAM/TSP patients. Of the subjects, 76% had an abnormal skin condition, with a high prevalence both among HTLV-1 asymptomatic carriers and HAM/TSP patients. The most prevalent SD-HTLV-1 was xerosis/acquired ichthyosis (48%), followed by seborrheic dermatitis (28%). Patients with SD-HTLV-1 were older (51 vs. 47 years), had a higher prevalence of myelopathy/tropical spastic paraparesis (HAM/TSP) (75%) and an increased first HTLV-1 proviral load compared with patients without SD-HTLV-1. When excluding HAM/TSP patients, the first HTLV-1 proviral load of SD-HTLV-1 individuals remains higher than no SD-HTLV-1 patients. Thus, skin diseases are highly prevalent among HTLV-1-infected individuals.
doi:10.1371/journal.pntd.0002546
PMCID: PMC3820737  PMID: 24244779
17.  Differential Live Mycobacterium tuberculosis-, M. bovis BCG-, Recombinant ESAT6-, and Culture Filtrate Protein 10-Induced Immunity in Tuberculosis▿  
The high prevalence of Mycobacterium tuberculosis makes it imperative that immune responses to evaluate could be predictive of infection. We investigated live Mycobacterium- and recombinant antigen-induced cytokine and chemokine responses in patients with active tuberculosis (TB) compared with those of healthy controls from an area where TB is endemic (ECs). M. tuberculosis-, M. bovis BCG-, ESAT6-, and culture filtrate protein 10 (CFP10)-induced responses were determined in peripheral blood mononuclear cells from patients with pulmonary TB (n = 38) and ECs (n = 39). The levels of the cytokines gamma interferon (IFN-γ) and interleukin-10 (IL-10) and the chemokines CCL2, CCL3, and CXCL9 were measured. The levels of M. tuberculosis- and BCG-induced IFN-γ secretion were significantly reduced (P = 0.002 and P < 0.01, respectively), while the amount of IL-10 induced by both virulent (P < 0.01) and avirulent (P = 0.002) mycobacteria was increased in patients with TB. The ESAT6-induced IFN-γ responses were increased in the patients with TB (P = 0.013) compared with those in the EC group. When tuberculin skin test (TST)-negative (TST−; induration, <10 mm) and TST-positive (TST+) donors were studied separately, both TST− and TST+ individuals showed increased IFN-γ responses to M. tuberculosis compared with the responses of the patients with TB (P = 0.037 and P = 0.006, respectively). However, only TST+ ECs showed reduced IFN-γ responses to ESAT6 (P = 0.008) compared with the responses of the patients with TB. The levels of M. tuberculosis-induced CCL2 (P = 0.006) and CXCL9 (P = 0.017) were greater in the patients with TB. The levels of CCL3 secretion in response to Mycobacterium and antigen stimulation were comparable between the two groups. While the levels of ESAT6-induced chemokines did not differ between the patients with TB and the ECs, the levels of CFP10-induced CCL2 (P = 0.01) and CXCL9 (P = 0.001) were increased in the patients. These data indicate differential host IFN-γ, CXCL9, and CCL2 responses to live mycobacteria and mycobacterial antigens and have implications for the identification of potential biomarkers of infection which could be used for the diagnosis of TB.
doi:10.1128/CVI.00091-09
PMCID: PMC2708395  PMID: 19439524
18.  Screening for HIV-Associated Tuberculosis and Rifampicin Resistance before Antiretroviral Therapy Using the Xpert MTB/RIF Assay: A Prospective Study 
PLoS Medicine  2011;8(7):e1001067.
In a prospective study, Stephen Lawn and colleagues find that pre-ART screening with Xpert MTB/RIF increased tuberculosis case detection by 45% compared to smear microscopy in HIV-positive patients at high risk of TB risk. AE competing interests must also pull through to the proof. “The Academic Editor, Madhukar Pai, declares that he consults for the Bill & Melinda Gates Foundation (BMGF). The BMGF supported FIND which was involved in the development of the Xpert MTB/RIF assay. He also co-chairs the Stop TB Partnership's New Diagnostics Working Group that was involved in the WHO endorsement of the Xpert assay.” Linked: Scott pmed.1001061; Evans pmed.1001064; Dowdy pmed.1001063
Background
The World Health Organization has endorsed the Xpert MTB/RIF assay for investigation of patients suspected of having tuberculosis (TB). However, its utility for routine TB screening and detection of rifampicin resistance among HIV-infected patients with advanced immunodeficiency enrolling in antiretroviral therapy (ART) services is unknown.
Methods and Findings
Consecutive adult HIV-infected patients with no current TB diagnosis enrolling in an ART clinic in a South African township were recruited regardless of symptoms. They were clinically characterised and invited to provide two sputum samples at a single visit. The accuracy of the Xpert MTB/RIF assay for diagnosing TB and drug resistance was assessed in comparison with other tests, including fluorescence smear microscopy and automated liquid culture (gold standard) and drug susceptibility testing. Of 515 patients enrolled, 468 patients (median CD4 cell count, 171 cells/µl; interquartile range, 102–236) produced at least one sputum sample, yielding complete sets of results from 839 samples. Mycobacterium tuberculosis was cultured from 81 patients (TB prevalence, 17.3%). The overall sensitivity of the Xpert MTB/RIF assay for culture-positive TB was 73.3% (specificity, 99.2%) compared to 28.0% (specificity, 100%) using smear microscopy. All smear-positive, culture-positive disease was detected by Xpert MTB/RIF from a single sample (sensitivity, 100%), whereas the sensitivity for smear-negative, culture-positive TB was 43.4% from one sputum sample and 62.3% from two samples. Xpert correctly identified rifampicin resistance in all four cases of multidrug-resistant TB but incorrectly identified resistance in three other patients whose disease was confirmed to be drug sensitive by gene sequencing (specificity, 94.1%; positive predictive value, 57%).
Conclusions
In this population of individuals at high risk of TB, intensive screening using the Xpert MTB/RIF assay increased case detection by 45% compared with smear microscopy, strongly supporting replacement of microscopy for this indication. However, despite the ability of the assay to rapidly detect rifampicin-resistant disease, the specificity for drug-resistant TB was sub-optimal.
Please see later in the article for the Editors' Summary
Editors' Summary
Background
Tuberculosis (TB)—a contagious bacterial infection that mainly affects the lungs—is a leading cause of illness and death among people who are infected with HIV, the virus that causes AIDS by destroying the immune system, which leaves infected individuals susceptible to other infections. TB is caused by Mycobacterium tuberculosis, which is spread in airborne droplets when people with the disease cough or sneeze. Its symptoms include a persistent cough, weight loss, and night sweats. Diagnostic tests for TB include chest X-rays, sputum smear analysis (microscopic examination of mucus coughed up from the lungs for M. tuberculosis bacilli), and mycobacterial liquid culture (the growth of M. tuberculosis from sputum and determination of its drug sensitivity). TB can be cured by taking several drugs daily for six months, although the recent emergence of multidrug-resistant TB (MDR-TB) is making the disease increasingly hard to treat.
Why Was This Study Done?
TB is a major problem in clinics that provide antiretroviral therapy (ART) for HIV-positive people in resource-limited settings. Not only is it a major cause of sickness and mortality in those affected by it, but TB (especially MDR-TB) can also spread to other patients attending the same clinic for health services. Rapid diagnosis and appropriate treatment are very important to reduce these risks. Unfortunately, sputum smear analysis—the mainstay of TB diagnosis in resource-limited settings—only detects about a fifth of TB cases when used as a screening tool before initiating ART. Chest X-rays are costly and don't always detect TB, and liquid culture—the gold standard method for TB diagnosis—is costly, technically difficult, and slow. Consequently, the World Health Organization (WHO) recently endorsed a new test for the investigation of patients suspected of having TB, especially in regions where HIV infection and MDR-TB are common. Xpert MTB/RIF is an automated DNA test that detects M. tuberculosis and DNA differences that make the bacteria resistant to the drug rifampicin (an indicator of MDR-TB) within 2 hours. In this study, the researchers investigate whether Xpert MTB/RIF could be used as a routine screening test to increase TB detection among HIV-positive people initiating ART.
What Did the Researchers Do and Find?
The researchers collected sputum from HIV-infected adults with no current TB diagnosis enrolling at an ART clinic in a South African township where HIV infection and TB are both common. They then compared the diagnostic accuracy of Xpert MTB/RIF (performed at a centralized laboratory) with that of several other tests, including liquid culture (the reference test). Nearly a fifth of the patients had culture-positive TB. Xpert MTB/RIF identified three-quarters of these patients (a sensitivity of 73.3%). By contrast, the sensitivity of smear microscopy was 28%. The new test's specificity (the proportion of patients with a negative Xpert MTB/RIF result among patients without TB) was 99.2%. That is, Xpert MTB/RIF had a low false-positive rate. Notably, Xpert MTB/RIF detected all cases of smear-positive, culture-positive TB but only 43.4% of smear-negative, culture-positive cases from a single sputum sample; it detected 62.3% of such cases when two sputum samples were analyzed. Finally, Xpert MTB/RIF correctly identified rifampicin resistance in all four patients who had MDR-TB but incorrectly identified resistance in three patients with drug-sensitive TB.
What Do These Findings Mean?
In this population of HIV-positive patients with a high TB risk, pre-ART screening with Xpert MTB/RIF increased case detection by 45% compared to smear microscopy, a finding that needs confirming in other settings. Importantly, Xpert MTB/RIF reduced the delay in diagnosis of TB from more than 20 days to two days. This delay would be reduced further by doing the assay at ART clinics rather than at a centralized testing facility, but the diagnostic accuracy of point-of-care testing needs evaluating. Overall, these findings (and those of an accompanying article by Scott et al. that examines the performance of Xpert MTB/RIF in an area where HIV infection is common) support the replacement of smear microscopy with Xpert MTB/RIF for pre-ART TB screening (provided misdiagnosis of rifampicin resistance can be reduced). These findings also suggest that routine screening with Xpert MTB/RIF could reduce the risk of MDR-TB outbreaks in HIV care and treatment settings and improve outcomes for HIV-positive patients with MDR-TB who currently often die before a diagnosis of TB can be made.
Additional Information
Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001056.
This study is further discussed in a PLoS Medicine Perspective by Carlton Evans; a related PLoS Medicine Research Article by Scott et al. is also available
WHO provides information (in several languages) on all aspects of tuberculosis, including general information on tuberculosis diagnostics and specific information on the Xpert MTB/RIF test; further information about WHO's endorsement of Xpert MTB/RIF is included in a recent Strategic and Technical Advisory Group for Tuberculosis report
WHO also provides information about tuberculosis and HIV
The US National Institute of Allergy and Infectious Diseases has detailed information on tuberculosis and HIV/AIDS
The US Centers for Disease Control and Prevention also has information about tuberculosis, including information on the diagnosis of and on tuberculosis and HIV co-infection
Information is available from Avert, an international AIDS charity, on many aspects of HIV/AIDS, including information on HIV-related tuberculosis (in English and Spanish)
doi:10.1371/journal.pmed.1001067
PMCID: PMC3144215  PMID: 21818180
19.  Prevalence of latent tuberculosis infection in Sudan: a case–control study comparing interferon-γ release assay and tuberculin skin test 
BMC Public Health  2013;13:1128.
Background
Most people exposed to M. tuberculosis show no evidence of clinical disease. Five to 10% of individuals with latent infection progress to develop overt disease during their life time. Identification of people with latent TB infection will increase case detection rates and may dictate new treatment policies to control tuberculosis. This study aimed to determine LTBI point prevalence in a population from Sudan using two different diagnostic methods: the tuberculin skin test (TST) and the IFN-γ release assay (IGRA).
Methods
This was a prospective, community-based and case-controlled study. Following informed consent, household contacts (HHCs; n = 98) of smear-positive index cases and Community controls (CCs; 186), were enrolled. Tuberculin skin test (TST), whole blood stimulation with ESAT-6/CFP-10 ± TB7.7 antigens or purified protein derivative (PPD) and IFN-γ levels determination with ELISA were performed. The levels of IFN-γ and TST induration between the CCs and the HHCs were compared using student t-test, Chi-square and Kappa coefficient. Pearson correlation test was used to compare TST and IFN-γ. P levels of <0.05 were considered significant.
Results
TST induration of ≥ 10 mm gave an LTBI point prevalence of 327 cases/1000 individuals among HHCs compared to 126 cases/1000 individuals among CCs (p = 0.000). PPD-induced IFN-γ release assay gave an LTBI point prevalence of 418 cases/1000 individuals among HHCs compared to 301 cases/1000 individuals among CCs (p =0.06). On the other hand ESAT-6/CFP-10 ± TB7.7-induced IFN-γ gave an LTBI point prevalence of 429 cases/1000 individuals among HHCs compared to 268 cases/1000 individuals among CCs (p = 0.01). IFN-γ productions levels induced by ESAT-6/CPF-10 ± TB7.7 antigens in HHCS and CCs were not significantly different from those induced by PPD (p = 0.7).
Conclusion
IFN-γ release assay (IGRA) gave higher LTBI point prevalence compared to TST in HHCs and CCs. PPD gave comparable results to ESAT-6/CFP-10 ± TB7.7 antigens in whole blood IFN-γ release, making it a cheap alternative to the recombinant antigens.
doi:10.1186/1471-2458-13-1128
PMCID: PMC4029475  PMID: 24313987
Latent Tb infection; IFN-γ release assay; TST; Sudan; Contact tracing
20.  Antiretroviral Therapy for Prevention of Tuberculosis in Adults with HIV: A Systematic Review and Meta-Analysis 
PLoS Medicine  2012;9(7):e1001270.
In a systematic review and meta-analysis, Amitabh Suthar and colleagues investigate the association between antiretroviral therapy and the reduction in the incidence of tuberculosis in adults with HIV infection.
Background
Human immunodeficiency virus (HIV) infection is the strongest risk factor for developing tuberculosis and has fuelled its resurgence, especially in sub-Saharan Africa. In 2010, there were an estimated 1.1 million incident cases of tuberculosis among the 34 million people living with HIV worldwide. Antiretroviral therapy has substantial potential to prevent HIV-associated tuberculosis. We conducted a systematic review of studies that analysed the impact of antiretroviral therapy on the incidence of tuberculosis in adults with HIV infection.
Methods and Findings
PubMed, Embase, African Index Medicus, LILACS, and clinical trial registries were systematically searched. Randomised controlled trials, prospective cohort studies, and retrospective cohort studies were included if they compared tuberculosis incidence by antiretroviral therapy status in HIV-infected adults for a median of over 6 mo in developing countries. For the meta-analyses there were four categories based on CD4 counts at antiretroviral therapy initiation: (1) less than 200 cells/µl, (2) 200 to 350 cells/µl, (3) greater than 350 cells/µl, and (4) any CD4 count.
Eleven studies met the inclusion criteria. Antiretroviral therapy is strongly associated with a reduction in the incidence of tuberculosis in all baseline CD4 count categories: (1) less than 200 cells/µl (hazard ratio [HR] 0.16, 95% confidence interval [CI] 0.07 to 0.36), (2) 200 to 350 cells/µl (HR 0.34, 95% CI 0.19 to 0.60), (3) greater than 350 cells/µl (HR 0.43, 95% CI 0.30 to 0.63), and (4) any CD4 count (HR 0.35, 95% CI 0.28 to 0.44). There was no evidence of hazard ratio modification with respect to baseline CD4 count category (p = 0.20).
Conclusions
Antiretroviral therapy is strongly associated with a reduction in the incidence of tuberculosis across all CD4 count strata. Earlier initiation of antiretroviral therapy may be a key component of global and national strategies to control the HIV-associated tuberculosis syndemic.
Review Registration
International Prospective Register of Systematic Reviews CRD42011001209
Please see later in the article for the Editors' Summary.
Editors' Summary
Background
Tuberculosis—a contagious bacterial infection— is a global public-health problem. In 2010, 8.8 million people developed active tuberculosis and 1.4 million people died from the disease. Tuberculosis can be cured by taking powerful antibiotics regularly for several months, and between 1995 and 2010, 46 million people with tuberculosis were successfully treated using DOTS—a directly observed antibiotic regimen designed by the World Health Organization (WHO). Now, though, the HIV epidemic is compromising global tuberculosis control efforts. HIV-positive people are very susceptible to tuberculosis because HIV, the virus that causes AIDS, destroys the immune system cells (including CD4 lymphocytes) that normally combat tuberculosis. In 2010, 1.1 million of the new (incident) cases of tuberculosis were among the 34 million people living with HIV, and 350,000 people died of HIV-associated tuberculosis, making tuberculosis the leading cause of death among HIV-positive people. To tackle HIV-associated tuberculosis, which occurs mainly in developing countries, WHO now recommends that HIV and tuberculosis programs use collaborative approaches such as the Three I's for HIV/TB strategy—intensified tuberculosis case-finding among HIV-positive people, isoniazid preventative therapy for HIV-positive people without active tuberculosis, and (tuberculosis) infection control in healthcare facilities, social settings, and households.
Why Was This Study Done?
Despite progress in scaling up the Three I's for HIV/TB strategy, complementary interventions are still needed to prevent tuberculosis in HIV-positive people. Antiretroviral therapy (ART) lowers the viral load of people infected with HIV and restores their immune system function and could, therefore, prevent HIVassociated tuberculosis, in addition to treating HIV infection. WHO recommends ART for all HIV-positive adults with a CD4 count of less than 350 cells/μl of blood and for all HIVpositive, tuberculosis-positive individuals irrespective of their CD4 count. However, the evidence for ART's preventative impact on tuberculosis has not been systematically examined. Here, the researchers undertake a systematic review (a search that uses predefined criteria to identify all the research on a given topic) and a meta-analysis (a statistical method for combining the results of studies) to investigate the impact of ART initiated at various CD4 counts on the development of tuberculosis in HIV-positive adults in developing countries.
What Did the Researchers Do and Find?
The researchers found 11 studies that compared tuberculosis incidence by ART status in HIV-infected adults over periods longer than six months on average in developing countries and undertook meta-analyses of these studies based on four categories of CD4 count at ART initiation (less than 200 cells/μl, 200–350 cells/μl, greater than 350 cells/μl, and any CD4 count). For all these categories, ART was strongly associated with a reduction in the incidence of tuberculosis. For example, the meta-analysis of the two studies that reported on participants in whom ART was initiated at a CD4 count less than 200 cells/μl yielded a hazard ratio (HR) of 0.16. That is, study participants starting ART when their CD4 count was below 200 cells/μl were about one-sixth as likely to develop tuberculosis as participants not receiving ART. In the metaanalysis of all 11 studies, study participants receiving ART were about one-third as likely to develop tuberculosis as study participants receiving no ART, irrespective of their CD4 count (HR 0.35). Importantly, the CD4 count at which ART was initiated did not significantly alter the magnitude of ART's preventive effect on tuberculosis development.
What Do These Findings Mean?
These findings suggest that ART is strongly associated with a reduction in the incidence of tuberculosis in HIV-positive adults in developing countries, whatever the CD4 count at ART initiation. Because most of the studies in this meta-analysis were observational, these results do not show that ART causes a reduction in tuberculosis incidence—other unknown factors shared by the study participants who received ART may be responsible for their lower tuberculosis incidence. Moreover, factors such as variations in diagnostic methods among the studies included in this meta-analysis may have affected the accuracy of these findings. Nevertheless, the key finding that ART is associated with a significant reduction in tuberculosis cases among adults with CD4 counts greater than 350 cells//μl should be considered by healthcare providers, policymakers, and people living with HIV when weighing the benefits and risks of early ART initiation. It also suggests that early ART initiation (in combination with expanded HIV testing) could be a key component of future global and national strategies to control HIV-associated tuberculosis.
Additional Information
Please access these websites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001270.
WHO provides information on all aspects of tuberculosis, including information on tuberculosis and HIV, on the Three I's for HIV/TB, and on ART for tuberculosis prevention (some information is in several languages)
The TB/HIV Working Group is part of the Stop TB Partnership, which is working toward tuberculosis elimination; patient stories about tuberculosis/HIV co-infection are also available on their site
The US Centers for Disease Control and Prevention has information about tuberculosis and about tuberculosis and HIV co-infection
The US National Institute of Allergy and Infectious Diseases also has detailed information on all aspects of tuberculosis including HIV-associated tuberculosis
Information is available from Avert, an international AIDS charity, on HIV-related tuberculosis (in English and Spanish), and from Aidsmap, a non-governmental organization, on HIV-associated tuberculosis
doi:10.1371/journal.pmed.1001270
PMCID: PMC3404110  PMID: 22911011
21.  Tuberculin Skin Test Reversion following Isoniazid Preventive Therapy Reflects Diversity of Immune Response to Primary Mycobacterium tuberculosis Infection 
PLoS ONE  2014;9(5):e96613.
Rationale
Healthy household contacts (HHC) of individuals with Tuberculosis (TB) with Tuberculin Skin Test (TST) conversions are considered to harbor latent Mycobacterium tuberculosis (Mtb), and at risk for TB. The immunologic, clinical, and public health implications of TST reversions that occur following Isoniazid preventive therapy (IPT) remain controversial.
Objectives
To measure frequency of TST reversion following IPT, and variation in interferon-gamma (IFN-γ) responses to Mtb, in healthy Ugandan TB HHC with primary Mtb infection evidenced by TST conversion.
Methods
Prospective cohort study of healthy, HIV-uninfected, TST-negative TB HHC with TST conversions. Repeat TST was performed 12 months following conversion (3 months following completion of 9 month IPT course) to assess for stable conversion vs. reversion. Whole blood IFN-γ responses to Mtb antigen 85B (MtbA85B) and whole Mtb bacilli (wMtb) were measured in a subset (n = 27 and n = 42, respectively) at enrollment and TST conversion, prior to initiation of IPT.
Results
Of 122 subjects, TST reversion was noted in 25 (20.5%). There were no significant differences in demographic, clinical, or exposure variables between reverters and stable converters. At conversion, reverters had significantly smaller TST compared to stable converters (13.7 mm vs 16.4 mm, respectively; p = 0.003). At enrollment, there were no significant differences in IFN-γ responses to MtbA85B or wMTB between groups. At conversion, stable converters demonstrated significant increases in IFN-γ responses to Ag85B and wMtb compared to enrollment (p = 0.001, p<0.001, respectively), while there were no significant changes among reverters.
Conclusions
TST reversion following IPT is common following primary Mtb infection and associated with unique patterns of Mtb-induced IFN-γ production. We have demonstrated that immune responses to primary Mtb infection are heterogeneous, and submit that prospective longitudinal studies of cell mediated immune responses to Mtb infection be prioritized to identify immune phenotypes protective against development of TB disease.
doi:10.1371/journal.pone.0096613
PMCID: PMC4010490  PMID: 24796677
22.  Tuberculin Skin Testing Compared with T-Cell Responses to Mycobacterium tuberculosis-Specific and Nonspecific Antigens for Detection of Latent Infection in Persons with Recent Tuberculosis Contact 
The tuberculin skin test (TST) is used for the identification of latent tuberculosis (TB) infection (LTBI) but lacks specificity in Mycobacterium bovis BCG-vaccinated individuals, who constitute an increasing proportion of TB patients and their contacts from regions where TB is endemic. In previous studies, T-cell responses to ESAT-6 and CFP-10, M. tuberculosis-specific antigens that are absent from BCG, were sensitive and specific for detection of active TB. We studied 44 close contacts of a patient with smear-positive pulmonary TB and compared the standard screening procedure for LTBI by TST or chest radiographs with T-cell responses to M. tuberculosis-specific and nonspecific antigens. Peripheral blood mononuclear cells were cocultured with ESAT-6, CFP-10, TB10.4 (each as recombinant antigen and as a mixture of overlapping synthetic peptides), M. tuberculosis sonicate, purified protein derivative (PPD), and short-term culture filtrate, using gamma interferon production as the response measure. LTBI screening was by TST in 36 participants and by chest radiographs in 8 persons. Nineteen contacts were categorized as TST negative, 12 were categorized as TST positive, and 5 had indeterminate TST results. Recombinant antigens and peptide mixtures gave similar results. Responses to TB10.4 were neither sensitive nor specific for LTBI. T-cell responses to ESAT-6 and CFP-10 were less sensitive for detection of LTBI than those to PPD (67 versus 100%) but considerably more specific (100 versus 72%). The specificity of the TST or in vitro responses to PPD will be even less when the proportion of BCG-vaccinated persons among TB contacts evaluated for LTBI increases.
doi:10.1128/CDLI.8.6.1089-1096.2001
PMCID: PMC96231  PMID: 11687445
23.  T-Cell Responses to the Mycobacterium tuberculosis-Specific Antigen ESAT-6 in Brazilian Tuberculosis Patients  
Infection and Immunity  2002;70(12):6707-6714.
The Mycobacterium tuberculosis-specific ESAT-6 antigen induces highly potent T-cell responses and production of gamma interferon (IFN-γ), which play a critical role in protective cell-mediated immunity against tuberculosis (TB). In the present study, IFN-γ secretion by peripheral blood mononuclear cells (PBMCs) in response to M. tuberculosis ESAT-6 in Brazilian TB patients was investigated in relation to clinical disease types, such as pleurisy and cavitary pulmonary TB. Leprosy patients, patients with pulmonary diseases other than TB, and healthy donors were assayed as control groups. Sixty percent of the TB patients indeed recognized M. tuberculosis ESAT-6, as did 50% of the leprosy patients and 60% of the non-TB controls. Nevertheless, the levels of IFN-γ in response to the antigen ESAT, but not to antigen 85B (Ag85B) and purified protein derivative (PPD), were significantly lower in controls than in patients with treated TB or pleural or cavitary TB. Moreover, according to Mycobacterium bovis BCG vaccination status, only 59% of the vaccinated TB patients responded to ESAT in vitro, whereas 100% of them responded to PPD. Both CD4 and CD8 T cells were able to release IFN-γ in response to ESAT. The present data demonstrate the specificity of ESAT-6 of M. tuberculosis and its ability to discriminate TB patients from controls, including leprosy patients. However, to obtain specificity, it is necessary to include quantitative IFN-γ production in response to the antigen as well, and this might limit the use of ESAT-6-based immunodiagnosis of M. tuberculosis infection in an area of TB endemicity.
doi:10.1128/IAI.70.12.6707-6714.2002
PMCID: PMC132944  PMID: 12438345
24.  Virologic and immunologic outcome of HAART in Human Immunodeficiency Virus (HIV)-1 infected patients with and without tuberculosis (TB) and latent TB infection (LTBI) in Addis Ababa, Ethiopia 
Background
HIV/TB coinfection remains a major challenge even after the initiation of HAART. Little is known about Mycobacterium tuberculosis (Mtb) specific immune restoration in relation to immunologic and virologic outcomes after long-term HAART during co-infections with latent and active TB.
Methods
A total of 232 adults, including 59 HIV patients with clinical TB (HIV + TB+), 125 HIV patients without clinical TB (HIV + TB-), 13 HIV negative active TB patients (HIV-TB+), and 10 HIV negative Tuberculin Skin TST positive (HIV-TST+), and 25 HIV-TST- individuals were recruited. HAART was initiated in 113 HIV + patients (28 TB + and 85 TB-), and anti-TB treatment for all TB cases. CD4+ T-cell count, HIV RNA load, and IFN-γ responses to ESAT-6/CFP-10 were measured at baseline, 6 months (M6), 18 months (M18) and 24 months (M24) after HAART initiation.
Results
The majority of HIV + TB- (70%, 81%, 84%) as well as HIV + TB + patients (60%, 77%, 80%) had virologic success (HIV RNA < 50 copies/ml) by M6, M18 and M24, respectively. HAART also significantly increased CD4+ T-cell counts at 2 years in HIV + TB + (from 110.3 to 289.9 cells/μl), HIV + TB- patients (197.8 to 332.3 cells/μl), HIV + TST- (199 to 347 cells/μl) and HIV + TST + individuals (195 to 319 cells/μl). Overall, there was no significant difference in the percentage of patients that achieved virologic success and in total CD4+ counts increased between HIV patients with and without TB or LTBI. The Mtb specific IFN-γ response at baseline was significantly lower in HIV + TB + (3.6 pg/ml) compared to HIV-TB + patients (34.4 pg/ml) and HIV + TST + (46.3 pg/ml) individuals; and in HIV-TB + patients compared to HIV-TST + individuals (491.2 pg/ml). By M18 on HAART, the IFN-γ response remained impaired in HIV + TB + patients (18.1 pg/ml) while it normalized in HIV + TST + individuals (from 46.3 to 414.2 pg/ml).
Conclusions
Our data show that clinical and latent TB infections do not influence virologic and immunologic outcomes of ART in HIV patients. Despite this, HAART was unable to restore optimal TB responsiveness as measured by Mtb specific IFN-γ response in HIV/TB patients. Improvement of Mtb-specific immune restoration should be the focus of future therapeutic strategies.
doi:10.1186/1742-6405-10-18
PMCID: PMC3718701  PMID: 23842109
HIV; Tuberculosis; HAART
25.  Prevalence of HTLV-1/2 in Pregnant Women Living in the Metropolitan Area of Rio de Janeiro 
Background
HTLV-1/2 infection can cause severe and disabling diseases in children and adults. The aim of the study was to estimate the prevalence of HTLV-1/2 infection in pregnant women living in the metropolitan area of Rio de Janeiro.
Methodology/Principal Findings
1,204 pregnant women were tested upon hospital admission for delivery in two public hospitals in the cities of Rio de Janeiro and Mesquita, between November, 2012 and April, 2013. The samples were screened by chemiluminescent microparticle immunoassay (CMIA) and reactive ones were confirmed by Western blot (WB). Epi-info software was used for building the database and performing the statistical analysis. Eight patients had confirmed HTLV-1/2 infection (7 HTLV-1, one HTLV-2), equivalent to a prevalence rate of 0.66%. Two further reactive screening tests had negative Western blot results and therefore were considered negative in the statistical analysis. All HTLV-1/2-positive patients were born in Rio de Janeiro, most were non-Caucasian (87.5%), in a stable relationship (62.5%), had at least ten years of formal education (62.5%) and a monthly family income of up to US$600.00 (87.5%). There was only one case of coinfection with syphilis and none with HIV. The mean age of the infected women was 28.4 (SD = 6.3) years and of the seronegative ones was 24.8 (SD = 6.5) (p = 0.10). The median number of pregnancies were 3.0 and 1.0 (p = 0.06) and the median number of sexual partners were 3.5 and 3.0 (p = 0.33) in the seropositive and negative groups, respectively. There were no statistically significant differences between the groups.
Conclusions/Significance
A significant prevalence of HTLV-1/2 was found in our population. The socio-epidemiological profile of carrier mothers was similar to the controls. Such findings expose the need for a public health policy of routine HTLV-1/2 screening in antenatal care, since counselling and preventive measures are the only strategies currently available to interrupt the chain of transmission and the future development of HTLV-1/2-related diseases.
Author Summary
HTLV-1/2 are retroviruses transmitted by blood products, sexual contact and from mother to child, mainly through breastfeeding. The infection has a characteristic geographical distribution with endemic areas often neighbouring very low prevalence areas. Infection is life long and although asymptomatic in most cases, it can cause severe and disabling diseases in children and adults. There is currently no cure, vaccine or effective treatment for HTLV-1/2 infections. Our research is the first to study the prevalence of HTLV-1/2 in pregnant women living in the metropolitan area of Rio de Janeiro, the second largest in Brazil. 1,204 pregnant women were tested upon hospital admission for delivery in two public hospitals in the cities of Rio de Janeiro and Mesquita, between November, 2012 and April, 2013 and a significant prevalence of HTLV-1/2 was found (0.66%). The socio-epidemiological profile of carrier mothers was similar to the controls'. Epidemiological knowledge is fundamental for the elaboration of public health policies such as routine HTLV-1/2 screening in antenatal care, since counselling and preventive measures, mainly avoidance of breastfeeding, are the only strategies currently available to interrupt the chain of transmission and the future development of HTLV-1/2-related diseases.
doi:10.1371/journal.pntd.0003146
PMCID: PMC4154655  PMID: 25188386

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