Suman Laal, Ph.D.

LipC (Rv0220) Is an Immunogenic Cell Surface Esterase of Mycobacterium tuberculosis
Shen, Guomiao; Singh, Krishna; Chandra, Dinesh; Serveau-Avesque, Carole; Maurin, Damien; Canaan, Stephane; Singla, Rupak; Behera, Digambar; Laal, Suman
2012 JAN ;80(1):243-253, Infection & immunity
We have reported previously the identification of novel proteins of Mycobacterium tuberculosis by the immunoscreening of an expression library of M. tuberculosis genomic DNA with sera obtained from M. tuberculosis-infected rabbits at 5 weeks postinfection. In this study, we report the further characterization of one of these antigens, LipC (Rv0220). LipC is annotated as a member of the Lip family based on the presence of the consensus motif 'GXSXG' characteristic of esterases. Although predicted to be a cytoplasmic enzyme, we provide evidence that LipC is a cell surface protein that is present in both the cell wall and the capsule of M. tuberculosis. Consistent with this localization, LipC elicits strong humoral immune responses in both HIV-negative (HIV(-)) and HIV-positive (HIV(+)) tuberculosis (TB) patients. The absence of anti-LipC antibodies in sera from purified protein derivative-positive (PPD(+)) healthy subjects confirms its expression only during active M. tuberculosis infection. Epitope mapping of LipC identified 6 immunodominant epitopes, 5 of which map to the exposed surface of the modeled LipC protein. The recombinant LipC (rLipC) protein also elicits proinflammatory cytokine and chemokine responses from macrophages and pulmonary epithelial cells. rLipC can hydrolyze short-chain esters with the carbon chain containing 2 to 10 carbon atoms. Together, these studies demonstrate that LipC is a novel cell surface-associated esterase of M. tuberculosis that is highly immunogenic and elicits both antibodies and cytokines/chemokines
— id: 150799, year: 2012, vol: 80, page: 243, stat: Journal Article,

Systematic review and meta-analysis of antigen detection tests for the diagnosis of tuberculosis
Flores L.L.; Steingart K.R.; Dendukuri N.; Schiller I.; Minion J.; Pai M.; Ramsay A.; Henry M.; Laal S.
2011 ;18(10):1616-1627, Clinical & vaccine immunology
Tests that detect Mycobacterium tuberculosis antigens in clinical specimens could provide rapid direct evidence of active disease. We performed a systematic review to assess the diagnostic accuracy of antigen detection tests for active tuberculosis (TB) according to standard methods and summarized test performance using bivariate random effects meta-analysis. Overall, study quality was a concern. For pulmonary TB (47 studies, 5,036 participants), sensitivity estimates ranged from 2% to 100% and specificity from 33% to 100%. Lipoarabinomannan (LAM) was the antigen most frequently targeted (23 studies, 49%). The pooled sensitivity of urine LAM was higher in HIV-infected than HIV-uninfected individuals (47%; 95% confidence interval [CI], 26 to 68% versus 14%; 95% CI, 4 to 38%); pooled specificity estimates were similar: 96%; 95% CI, 81 to 100% and 97%; 95% CI, 86 to 100%, respectively. For extrapulmonary TB (21 studies, 1,616 participants), sensitivity estimates ranged from 0% to 100% and specificity estimates from 62% to 100%. Five studies targeting LAM, ESAT-6, Ag85 complex, and the 65-kDa antigen in cerebrospinal fluid, when pooled, yielded the highest sensitivity (87%; 95% CI, 61 to 98%), but low specificity (84%; 95% CI, 60 to 95%). Because of the limited number of studies targeting any specific antigen other than LAM, we could not draw firm conclusions about the overall clinical usefulness of these tests. Further studies are warranted to determine the value of LAM detection for TB meningitis in high-HIV-prevalence settings. Considering that antigen detection tests could be translated into rapid point-of-care tests, research to improve their performance is urgently needed.
— id: 138727, year: 2011, vol: 18, page: 1616, stat: Journal Article,

Commercial serological tests for the diagnosis of active pulmonary and extrapulmonary tuberculosis: an updated systematic review and meta-analysis
Steingart, Karen R; Flores, Laura L; Dendukuri, Nandini; Schiller, Ian; Laal, Suman; Ramsay, Andrew; Hopewell, Philip C; Pai, Madhukar
2011 Aug;8(8):e1001062-e1001062, PLoS medicine
BACKGROUND: Serological (antibody detection) tests for tuberculosis (TB) are widely used in developing countries. As part of a World Health Organization policy process, we performed an updated systematic review to assess the diagnostic accuracy of commercial serological tests for pulmonary and extrapulmonary TB with a focus on the relevance of these tests in low- and middle-income countries. METHODS AND FINDINGS: We used methods recommended by the Cochrane Collaboration and GRADE approach for rating quality of evidence. In a previous review, we searched multiple databases for papers published from 1 January 1990 to 30 May 2006, and in this update, we add additional papers published from that period until 29 June 2010. We prespecified subgroups to address heterogeneity and summarized test performance using bivariate random effects meta-analysis. For pulmonary TB, we included 67 studies (48% from low- and middle-income countries) with 5,147 participants. For all tests, estimates were variable for sensitivity (0% to 100%) and specificity (31% to 100%). For anda-TB IgG, the only test with enough studies for meta-analysis, pooled sensitivity was 76% (95% CI 63%-87%) in smear-positive (seven studies) and 59% (95% CI 10%-96%) in smear-negative (four studies) patients; pooled specificities were 92% (95% CI 74%-98%) and 91% (95% CI 79%-96%), respectively. Compared with ELISA (pooled sensitivity 60% [95% CI 6%-65%]; pooled specificity 98% [95% CI 96%-99%]), immunochromatographic tests yielded lower pooled sensitivity (53%, 95% CI 42%-64%) and comparable pooled specificity (98%, 95% CI 94%-99%). For extrapulmonary TB, we included 25 studies (40% from low- and middle-income countries) with 1,809 participants. For all tests, estimates were variable for sensitivity (0% to 100%) and specificity (59% to 100%). Overall, quality of evidence was graded very low for studies of pulmonary and extrapulmonary TB. CONCLUSIONS: Despite expansion of the literature since 2006, commercial serological tests continue to produce inconsistent and imprecise estimates of sensitivity and specificity. Quality of evidence remains very low. These data informed a recently published World Health Organization policy statement against serological tests. Please see later in the article for the Editors' Summary
— id: 137459, year: 2011, vol: 8, page: e1001062, stat: Journal Article,

Antibodies against immunodominant antigens of Mycobacterium tuberculosis in subjects with suspected tuberculosis in the United States compared by HIV status
Achkar, Jacqueline M; Jenny-Avital, Elisabeth; Yu, Xian; Burger, Susanne; Leibert, Eric; Bilder, Patrick W; Almo, Steven C; Casadevall, Arturo; Laal, Suman
2010 Mar;17(3):384-392, Clinical & vaccine immunology
The immunodominance of Mycobacterium tuberculosis proteins malate synthase (MS) and MPT51 has been demonstrated in case-control studies with patients from countries in which tuberculosis (TB) is endemic. The value of these antigens for the serodiagnosis of TB now is evaluated in a cross-sectional study of pulmonary TB suspects in the United States diagnosed to have TB, HIV-associated TB, or other respiratory diseases (ORD). Serum antibody reactivity to recombinant purified MS and MPT51 was determined by enzyme-linked immunosorbent assays (ELISAs) of samples from TB suspects and well-characterized control groups. TB suspects were diagnosed with TB (n = 87; 49% sputum microscopy negative, 20% HIV(+)) or ORD (n = 63; 58% HIV(+)). Antibody reactivity to MS and MPT51 was significantly higher in U.S. HIV(+)/TB samples than in HIV(-)/TB samples (P < 0.001), and it was significantly higher in both TB groups than in control groups with latent TB infection (P < 0.001). Antibody reactivity to both antigens was higher in U.S. HIV(+)/TB samples than in HIV(+)/ORD samples (P = 0.052 for MS, P = 0.001 for MPT51) but not significantly different between HIV(-)/TB and HIV(-)/ORD. Among U.S. HIV(+) TB suspects, a positive anti-MPT51 antibody response was strongly and significantly associated with TB (odds ratio, 11.0; 95% confidence interval, 2.3 to 51.2; P = 0.002). These findings have implications for the adjunctive use of TB serodiagnosis with these antigens in HIV(+) subjects
— id: 133488, year: 2010, vol: 17, page: 384, stat: Journal Article,

Potential role for ESAT6 in dissemination of M. tuberculosis via human lung epithelial cells
Kinhikar, Arvind G; Verma, Indu; Chandra, Dinesh; Singh, Krishna K; Weldingh, Karin; Andersen, Peter; Hsu, Tsungda; Jacobs, William R Jr; Laal, Suman
2010 Jan;75(1):92-106, Molecular microbiology
ESAT6 has recently been demonstrated to cause haemolysis and macrophage lysis. Our studies demonstrate that ESAT6 causes cytolysis of type 1 and type 2 pneumocytes. Both types of pneumocytes express membrane laminin, and ESAT6 exhibits dose-dependent binding to both cell types and to purified human laminin. While minimal ESAT6 was detected on the surface of Mycobacterium tuberculosis grown in vitro, exogenously provided ESAT6 specifically associated with the bacterial cell surface, and the bacterium-associated ESAT6 retained its cytolytic ability. esat6 transcripts were upregulated approximately 4- to approximately 13-fold in bacteria replicating in type 1 cells, and approximately 3- to approximately 5 fold in type 2 cells. In vivo, laminin is primarily concentrated at the basolateral surface of pneumocytes where they rest on the basement membrane, which is composed primarily of laminin and collagen. The upregulation of esat6 transcripts in bacteria replicating in pneumocytes, the specific association of ESAT6 with the bacterial surface, the binding of ESAT6 to laminin and the lysis of pneumocytes by free and bacterium-associated ESAT6 together suggest a scenario wherein Mycobacterium tuberculosis replicating in pneumocytes may utilize surface ESAT6 to anchor onto the basolateral laminin-expressing surface of the pneumocytes, and damage the cells and the basement membrane to directly disseminate through the alveolar wall
— id: 106091, year: 2010, vol: 75, page: 92, stat: Journal Article,

Neutralization efficiency and presence of anti-V3 antibodies in plasma of HIV-1 infected Northern Indians
Choudhary, Alok K.; Dutta, Subhashree; Wig, Naveet; Biswas, A.; Andrabi, Raiees; Kalra, Rajesh; Bhasin, Rama; Pazner, Susan Zolla; Laal, Suman; Luthra, Kalpana
2009 JUN ;51(10):160-160, Journal of acquired immune deficiency syndromes. JAIDS
— id: 113759, year: 2009, vol: 51, page: 160, stat: Journal Article,

Identification of Mycobacterium avium KatG protein (MAV_2753) as a possible serodiagnostic marker for MAC disease
Gupta, Kapil; Khuller, Gopal Krishan; Wanchu, Ajay; Laal, Suman; Latawa, Romica; Verma, Indu
2009 Feb;58(2):170-172, Journal of infection
— id: 95778, year: 2009, vol: 58, page: 170, stat: Journal Article,

Mycobacterium avium KatG protein (MAV_2753): a putative candidate for the serodiagnosis of MAC disease
Gupta, Kapil; Wanchu, Ajay; Latawa, Romica; Laal, S.; Khuller, G. K.; Verma, Indu
2009 JUN ;51(10):168-168, Journal of acquired immune deficiency syndromes. JAIDS
— id: 113760, year: 2009, vol: 51, page: 168, stat: Journal Article,

Peptide-based antibody detection for tuberculosis diagnosis
Shen, Guomiao; Behera, Digambar; Bhalla, Manpreet; Nadas, Arthur; Laal, Suman
2009 Jan;16(1):49-54, Clinical & vaccine immunology
Tuberculosis (TB) is a major cause of morbidity and mortality, especially in developing countries. Despite significant limitations, microscopy remains the cornerstone of the global TB control strategy. As the TB epidemic escalates, new diagnostic methods that are accurate and also economical and simple to manufacture and deploy are urgently needed. Although several promising antigens have been identified and evaluated in recent years, the reproducible production of high-quality recombinant mycobacterial proteins with minimal batch-to-batch variation is difficult, laborious, and expensive. To determine the feasibility of devising a synthetic peptide-based diagnostic test for TB, we have delineated the immunodominant epitopes of three candidate antigens, Ag85B, BfrB, and TrxC, that were previously identified to be immunogenic in TB patients. The results demonstrate that combinations of carefully selected synthetic peptides derived from highly immunogenic proteins can be the basis for devising an immunodiagnostic test for TB
— id: 92172, year: 2009, vol: 16, page: 49, stat: Journal Article,

Peptides of a novel Mycobacterium tuberculosis-specific cell wall protein for immunodiagnosis of tuberculosis
Singh, Krishna K; Sharma, Naresh; Vargas, Diana; Liu, Zhentong; Belisle, John T; Potharaju, Visalakshi; Wanchu, Ajay; Behera, Digambar; Laal, Suman
2009 Aug 15;200(4):571-581, Journal of infectious diseases
The sequencing of the Mycobacterium tuberculosis genome revealed the existence of several genes encoding novel proteins with unknown functions, one of which is the proline-threonine repetitive protein (PTRP; Rv0538). Genomic studies of various mycobacterial species and M. tuberculosis clinical isolates demonstrate that ptrp is specific to the M. tuberculosis complex and ubiquitous in clinical isolates. Enzyme-linked immunosorbent assay, Western blot analysis, and electron microscopic evaluation of M. tuberculosis subcellular fractions and intact bacteria confirm that PTRP is a cell wall protein. Antibodies to PTRP are present in serum specimens from human immunodeficiency virus (HIV)-negative, tuberculosis (TB)-positive and HIV-positive, TB-positive patients but not purified protein derivative (PPD)-negative or PPD-positive healthy control subjects, demonstrating its diagnostic potential. Epitope mapping of PTRP delineated 4 peptides that can identify >80% of sputum smear-positive and >50% of smear-negative, HIV-negative, TB-positive patients and >80% of HIV-positive, TB-positive patients. These results demonstrate that immunodominant epitopes of carefully selected M. tuberculosis-specific proteins can be used to devise a simple peptide-based serodiagnostic test for TB
— id: 101637, year: 2009, vol: 200, page: 571, stat: Journal Article,

Performance of purified antigens for serodiagnosis of pulmonary tuberculosis: a meta-analysis
Steingart, Karen R; Dendukuri, Nandini; Henry, Megan; Schiller, Ian; Nahid, Payam; Hopewell, Philip C; Ramsay, Andrew; Pai, Madhukar; Laal, Suman
2009 Feb;16(2):260-276, Clinical & vaccine immunology
Serological antibody detection tests for tuberculosis may offer the potential to improve diagnosis. Recent meta-analyses have shown that commercially available tests have variable accuracies and a limited clinical role. We reviewed the immunodiagnostic potential of antigens evaluated in research laboratories (in-house) for the serodiagnosis of pulmonary tuberculosis and conducted a meta-analysis to evaluate the performance of comparable antigens. Selection criteria included the participation of at least 25 pulmonary tuberculosis patients and the use of purified antigens. Studies evaluating 38 kDa, MPT51, malate synthase, culture filtrate protein 10, TbF6, antigen 85B, alpha-crystallin, 2,3-diacyltrehalose, 2,3,6-triacyltrehalose, 2,3,6,6'-tetraacyltrehalose 2'-sulfate, cord factor, and TbF6 plus DPEP (multiple antigen) were included in the meta-analysis. The results demonstrated that (i) in sputum smear-positive patients, sensitivities significantly >or=50% were provided for recombinant malate synthase (73%; 95% confidence interval [CI], 58 to 85) and TbF6 plus DPEP (75%; 95% CI, 50 to 91); (ii) protein antigens achieved high specificities; (iii) among the lipid antigens, cord factor had the best overall performance (sensitivity, 69% [95% CI, 28 to 94]; specificity, 91% [95% CI, 78 to 97]); (iv) compared with the sensitivities achieved with single antigens (median sensitivity, 53%; range, 2% to 100%), multiple antigens yielded higher sensitivities (median sensitivity, 76%; range, 16% to 96%); (v) in human immunodeficiency virus (HIV)-infected patients who are sputum smear positive, antibodies to several single and multiple antigens were detected; and (vi) data on seroreactivity to antigens in sputum smear-negative or pediatric patients were insufficient. Potential candidate antigens for an antibody detection test for pulmonary tuberculosis in HIV-infected and -uninfected patients have been identified, although no antigen achieves sufficient sensitivity to replace sputum smear microscopy. Combinations of select antigens provide higher sensitivities than single antigens. The use of a case-control design with healthy controls for the majority of studies was a limitation of the review. Efforts are needed to improve the methodological quality of tuberculosis diagnostic studies
— id: 95779, year: 2009, vol: 16, page: 260, stat: Journal Article,

Biomarkers for clinical and incipient tuberculosis: performance in a TB-endemic country
Wanchu, Ajay; Dong, Yuxin; Sethi, Sunil; Myneedu, V P; Nadas, Arthur; Liu, Zhentong; Belisle, John; Laal, Suman
2008 ;3(4):e2071-e2071, PLoS ONE
BACKGROUND: Simple biomarkers are required to identify TB in both HIV(-)TB(+) and HIV(+)TB(+) patients. Earlier studies have identified the M. tuberculosis Malate Synthase (MS) and MPT51 as immunodominant antigens in TB patients. One goal of these investigations was to evaluate the sensitivity and specificity of anti-MS and -MPT51 antibodies as biomarkers for TB in HIV(-)TB(+) and HIV(+)TB(+) patients from a TB-endemic setting. Earlier studies also demonstrated the presence of these biomarkers during incipient subclinical TB. If these biomarkers correlate with incipient TB, their prevalence should be higher in asymptomatic HIV(+) subjects who are at a high-risk for TB. The second goal was to compare the prevalence of these biomarkers in asymptomatic, CD4(+) T cell-matched HIV(+)TB(-) subjects from India who are at high-risk for TB with similar subjects from US who are at low-risk for TB. METHODS AND RESULTS: Anti-MS and -MPT51 antibodies were assessed in sera from 480 subjects including PPD(+) or PPD(-) healthy subjects, healthy community members, and HIV(-)TB(+) and HIV(+)TB(+) patients from India. Results demonstrate high sensitivity (approximately 80%) of detection of smear-positive HIV(-)TB(+) and HIV(+)TB(+) patients, and high specificity (>97%) with PPD(+) subjects and endemic controls. While approximately 45% of the asymptomatic HIV(+)TB(-) patients at high-risk for TB tested biomarker-positive, >97% of the HIV(+)TB(-) subjects at low risk for TB tested negative. Although the current studies are hampered by lack of knowledge of the outcome, these results provide strong support for the potential of these biomarkers to detect incipient, subclinical TB in HIV(+) subjects. CONCLUSIONS: These biomarkers provide high sensitivity and specificity for TB diagnosis in a TB endemic setting. Their performance is not compromised by concurrent HIV infection, site of TB and absence of pulmonary manifestations in HIV(+)TB(+) patients. Results also demonstrate the potential of these biomarkers for identifying incipient subclinical TB in HIV(+)TB(-) subjects at high-risk for TB
— id: 78837, year: 2008, vol: 3, page: e2071, stat: Journal Article,

A systematic review of commercial serological antibody detection tests for the diagnosis of extrapulmonary tuberculosis
Steingart, Karen R; Henry, Megan; Laal, Suman; Hopewell, Philip C; Ramsay, Andrew; Menzies, Dick; Cunningham, Jane; Weldingh, Karin; Pai, Madhukar
2007 Nov;83(985):705-712, Postgraduate medical journal
Conventional diagnostic tests for tuberculosis have several limitations and are often unhelpful in establishing the diagnosis of extrapulmonary tuberculosis. Although commercial serological antibody based tests are available, their usefulness in the diagnosis of extrapulmonary tuberculosis is unknown. A systematic review was conducted to assess the accuracy of commercial serological antibody detection tests for the diagnosis of extrapulmonary tuberculosis. In a comprehensive search, 21 studies that reported data on sensitivity and specificity for extrapulmonary tuberculosis were identified. These studies evaluated seven different commercial tests, with Anda-TB IgG accounting for 48% of the studies. The results showed that (1) all commercial tests provided highly variable estimates of sensitivity (range 0.00-1.00) and specificity (range 0.59-1.00) for all extrapulmonary sites combined; (2) the Anda-TB IgG kit showed highly variable sensitivity (range 0.26-1.00) and specificity (range 0.59-1.00) for all extrapulmonary sites combined; (3) for all tests combined, sensitivity estimates for both lymph node tuberculosis (range 0.23-1.00) and pleural tuberculosis (range 0.26-0.59) were poor and inconsistent; and (4) there were no data to determine the accuracy of the tests in children or in patients with HIV infection, the two groups for which the test would be most useful. At present, commercial antibody detection tests for extrapulmonary tuberculosis have no role in clinical care or case detection
— id: 78838, year: 2007, vol: 83, page: 705, stat: Journal Article,

A systematic review of commercial serological antibody detection tests for the diagnosis of extrapulmonary tuberculosis
Steingart, Karen R; Henry, Megan; Laal, Suman; Hopewell, Philip C; Ramsay, Andrew; Menzies, Dick; Cunningham, Jane; Weldingh, Karin; Pai, Madhukar
2007 Oct;62(10):911-918, Thorax
Conventional diagnostic tests for tuberculosis have several limitations and are often unhelpful in establishing the diagnosis of extrapulmonary tuberculosis. Although commercial serological antibody based tests are available, their usefulness in the diagnosis of extrapulmonary tuberculosis is unknown. A systematic review was conducted to assess the accuracy of commercial serological antibody detection tests for the diagnosis of extrapulmonary tuberculosis. In a comprehensive search, 21 studies that reported data on sensitivity and specificity for extrapulmonary tuberculosis were identified. These studies evaluated seven different commercial tests, with Anda-TB IgG accounting for 48% of the studies. The results showed that (1) all commercial tests provided highly variable estimates of sensitivity (range 0.00-1.00) and specificity (range 0.59-1.00) for all extrapulmonary sites combined; (2) the Anda-TB IgG kit showed highly variable sensitivity (range 0.26-1.00) and specificity (range 0.59-1.00) for all extrapulmonary sites combined; (3) for all tests combined, sensitivity estimates for both lymph node tuberculosis (range 0.23-1.00) and pleural tuberculosis (range 0.26-0.59) were poor and inconsistent; and (4) there were no data to determine the accuracy of the tests in children or in patients with HIV infection, the two groups for which the test would be most useful. At present, commercial antibody detection tests for extrapulmonary tuberculosis have no role in clinical care or case detection
— id: 78839, year: 2007, vol: 62, page: 911, stat: Journal Article,

Commercial serological antibody detection tests for the diagnosis of pulmonary tuberculosis: a systematic review
Steingart, Karen R; Henry, Megan; Laal, Suman; Hopewell, Philip C; Ramsay, Andrew; Menzies, Dick; Cunningham, Jane; Weldingh, Karin; Pai, Madhukar
2007 Jun;4(6):e202-e202, PLoS medicine
BACKGROUND: The global tuberculosis epidemic results in nearly 2 million deaths and 9 million new cases of the disease a year. The vast majority of tuberculosis patients live in developing countries, where the diagnosis of tuberculosis relies on the identification of acid-fast bacilli on unprocessed sputum smears using conventional light microscopy. Microscopy has high specificity in tuberculosis-endemic countries, but modest sensitivity which varies among laboratories (range 20% to 80%). Moreover, the sensitivity is poor for paucibacillary disease (e.g., pediatric and HIV-associated tuberculosis). Thus, the development of rapid and accurate new diagnostic tools is imperative. Immune-based tests are potentially suitable for use in low-income countries as some test formats can be performed at the point of care without laboratory equipment. Currently, dozens of distinct commercial antibody detection tests are sold in developing countries. The question is 'do they work?' METHODS AND FINDINGS: We conducted a systematic review to assess the accuracy of commercial antibody detection tests for the diagnosis of pulmonary tuberculosis. Studies from all countries using culture and/or microscopy smear for confirmation of pulmonary tuberculosis were eligible. Studies with fewer than 50 participants (25 patients and 25 control participants) were excluded. In a comprehensive search, we identified 68 studies. The results demonstrate that (1) overall, commercial tests vary widely in performance; (2) sensitivity is higher in smear-positive than smear-negative samples; (3) in studies of smear-positive patients, Anda-TB IgG by enzyme-linked immunosorbent assay shows limited sensitivity (range 63% to 85%) and inconsistent specificity (range 73% to 100%); (4) specificity is higher in healthy volunteers than in patients in whom tuberculosis disease is initially suspected and subsequently ruled out; and (5) there are insufficient data to determine the accuracy of most commercial tests in smear microscopy-negative patients, as well as their performance in children or persons with HIV infection. CONCLUSIONS: None of the commercial tests evaluated perform well enough to replace sputum smear microscopy. Thus, these tests have little or no role in the diagnosis of pulmonary tuberculosis. Lack of methodological rigor in these studies was identified as a concern. It will be important to review the basic science literature evaluating serological tests for the diagnosis of pulmonary tuberculosis to determine whether useful antigens have been described but their potential has not been fully exploited. Activities leading to the discovery of new antigens with immunodiagnostic potential need to be intensified
— id: 78840, year: 2007, vol: 4, page: e202, stat: Journal Article,

Mycobacterium tuberculosis malate synthase- and MPT51-based serodiagnostic assay as an adjunct to rapid identification of pulmonary tuberculosis
Achkar, Jacqueline M; Dong, Yuxin; Holzman, Robert S; Belisle, John; Kourbeti, Irene S; Sherpa, Tsering; Condos, Rany; Rom, William N; Laal, Suman
2006 Nov;13(11):1291-1293, Clinical & vaccine immunology
The 81-kDa malate synthase (MS; Rv 1837c) and the 27-kDa MPT51 (Rv 3803c) of Mycobacterium tuberculosis are immunodominant antigens recognized by serum antibodies from approximately 80% of human immunodeficiency virus-negative smear-positive tuberculosis patients from India. We now provide evidence that the use of the MS/MPT51-based serodiagnostic assay can serve as an adjunct to sputum microscopy in the rapid diagnosis of pulmonary tuberculosis
— id: 70310, year: 2006, vol: 13, page: 1291, stat: Journal Article,

Novel approach for differential diagnosis of HIV infections in the face of vaccine-generated antibodies: utility for detection of diverse HIV-1 subtypes
Khurana, Surender; Needham, James; Park, Susan; Mathieson, Bonnie; Busch, Michael P; Nemo, George; Nyambi, Phillipe; Zolla-Pazner, Susan; Laal, Suman; Mulenga, Joseph; Chomba, Elwyn; Hunter, Eric; Allen, Susan; McIntyre, James; Hewlett, Indira; Lee, Sherwin; Tang, Shixing; Cowan, Elliot; Beyrer, Chris; Altfeld, Marcus; Yu, Xu G; Tounkara, Anatole; Koita, Ousmane; Kamali, Anatoli; Nguyen, Nga; Graham, Barney S; Todd, Deborah; Mugenyi, Peter; Anzala, Omu; Sanders, Eduard; Ketter, Nzeera; Fast, Patricia; Golding, Hana
2006 Nov 1;43(3):304-312, Journal of acquired immune deficiency syndromes. JAIDS
Because increasing numbers of HIV vaccine candidates are being tested globally, it is essential to differentiate vaccine- from virus-induced antibodies. Most of the currently tested vaccines contain multiple viral components. As a result, many vaccine recipients give positive results in FDA-licensed HIV serodetection tests. We have identified conserved sequences in Env-gp41 and Gag-p6, which are recognized soon after infection but are not included in most HIV vaccine candidates. A new HIV serodetection assay, the HIV-SELECTEST, was established that distinguishes between vaccine-induced antibodies and seroconversion due to true HIV infections. It is important to make this assay globally relevant, because many clinical trials are conducted around the world where most HIV infections are due to non-B subtype HIV-1. Therefore, the current study examined the reactivity of plasma samples from >3,000 infections with diverse HIV subtypes worldwide. The HIV-SELECTEST performed at >99% specificity and sensitivity. Both recent and established infections with clades A, B, C, D, E, F, G, J, and CRFs were detected. Antibodies elicited by other vaccinations or infections endemic to the clinical trial sites did not react in this assay. Therefore, HIV-SELECTEST could be an important differential diagnostic tool for HIV vaccine trials, blood banks, and population screening worldwide
— id: 78810, year: 2006, vol: 43, page: 304, stat: Journal Article,

Mycobacterium tuberculosis malate synthase is a laminin-binding adhesin
Kinhikar, Arvind G; Vargas, Diana; Li, Hualin; Mahaffey, Spencer B; Hinds, Laura; Belisle, John T; Laal, Suman
2006 May;60(4):999-1013, Molecular microbiology
Mycobacterium tuberculosis (M. tb) uses the glyoxalate bypass for intracellular survival in vivo. These studies provide evidence that the M. tb malate synthase (MS) has adapted to function as an adhesin which binds to laminin and fibronectin. This binding is achieved via the unique C-terminal region of the M. tb MS. The ability to function as an adhesin necessitates extracellular localization. We provide evidence that despite the absence of a Sec-translocation signal sequence the M. tb MS is secreted/excreted, and is anchored on the cell wall by an undefined mechanism. The MS of Mycobacterium smegmatis is cytoplasmic but the M. tb MS expressed in M. smegmatis localizes to the cell wall and enhances the adherence of the bacteria to lung epithelial A549 cells. Antibodies to the C-terminal laminin/fibronectin-binding domain interfere with the binding of the M. tb MS to laminin and fibronectin and reduce the adherence of M. tb to A549 cells. Coupled to the earlier evidence of in vivo expression of M. tb MS during active but not latent infection in humans, these studies show that a housekeeping enzyme of M. tb contributes to its armamentarium of virulence promoting factors
— id: 67530, year: 2006, vol: 60, page: 999, stat: Journal Article,

Disease state differentiation and identification of tuberculosis biomarkers via native antigen array profiling
Sartain, Mark J; Slayden, Richard A; Singh, Krishna K; Laal, Suman; Belisle, John T
2006 Nov;5(11):2102-2113, Molecular & cellular proteomics
A critical element of tuberculosis control is early and sensitive diagnosis of infection and disease. Our laboratories recently showed that different stages of disease were distinguishable via two-dimensional Western blot analyses of Mycobacterium tuberculosis culture filtrate proteins. However, this methodology is not suitable for high throughput testing. Advances in protein microarray technology provide a realistic mechanism to screen a large number of serum samples against thousands of proteins to identify biomarkers of disease states. Techniques were established for separation of native M. tuberculosis cytosol and culture filtrate proteins, resulting in 960 unique protein fractions that were used to generate protein microarrays. Evaluation of serological reactivity from 42 patients in three tuberculosis disease states and healthy purified protein derivative-positive individuals demonstrated that human immunodeficiency virus (HIV)-negative cavitary and noncavitary tuberculosis (TB) patients' sera recognized 126 and 59 fractions, respectively. Sera from HIV patients coinfected with TB recognized 20 fractions of which five overlapped with those recognized by non-HIV TB patients' sera and 15 were unique to the HIV+TB+ disease state. Identification of antigens within the reactive fractions yielded 11 products recognized by both cavitary and noncavitary TB patients' sera and four proteins (HspX, MPT64, PstS1, and TrxC) specific to cavitary TB patients. Moreover four novel B cell antigens (BfrB, LppZ, SodC, and TrxC) of human tuberculosis were identified
— id: 78841, year: 2006, vol: 5, page: 2102, stat: Journal Article,

Immunodiagnosis
Laal, Suman
Tuberculosis Philadelphia : Lippincott Williams & Wilkins, 2004,
— id: 3964, year: 2004, vol: , page: 186, stat: Chapter,

The Mycobacterium tuberculosis complex-restricted gene cfp32 encodes an expressed protein that is detectable in tuberculosis patients and is positively correlated with pulmonary interleukin-10
Huard, Richard C; Chitale, Sadhana; Leung, Mary; Lazzarini, Luiz Claudio Oliveira; Zhu, Hongxia; Shashkina, Elena; Laal, Suman; Conde, Marcus B; Kritski, Afranio L; Belisle, John T; Kreiswirth, Barry N; Lapa e Silva, Jose Roberto; Ho, John L
2003 Dec;71(12):6871-6883, Infection & immunity
Human tuberculosis (TB) is caused by the bacillus Mycobacterium tuberculosis, a subspecies of the M. tuberculosis complex (MTC) of mycobacteria. Postgenomic dissection of the M. tuberculosis proteome is ongoing and critical to furthering our understanding of factors mediating M. tuberculosis pathobiology. Towards this end, a 32-kDa putative glyoxalase in the culture filtrate (CF) of growing M. tuberculosis (originally annotated as Rv0577 and hereafter designated CFP32) was identified, cloned, and characterized. The cfp32 gene is MTC restricted, and the gene product is expressed ex vivo as determined by the respective Southern and Western blot testing of an assortment of mycobacteria. Moreover, the cfp32 gene sequence is conserved within the MTC, as no polymorphisms were found in the tested cfp32 PCR products upon sequence analysis. Western blotting of M. tuberculosis subcellular fractions localized CFP32 predominantly to the CF and cytosolic compartments. Data to support the in vivo expression of CFP32 were provided by the serum recognition of recombinant CFP32 in 32% of TB patients by enzyme-linked immunosorbent assay (ELISA) as well as the direct detection of CFP32 by ELISA in the induced sputum samples from 56% of pulmonary TB patients. Of greatest interest was the observation that, per sample, sputum CFP32 levels (a potential indicator of increasing bacterial burden) correlated with levels of expression in sputum of interleukin-10 (an immunosuppressive cytokine and a putative contributing factor to disease progression) but not levels of gamma interferon (a key cytokine in the protective immune response in TB), as measured by ELISA. Combined, these data suggest that CFP32 serves a necessary biological function(s) in tubercle bacilli and may contribute to the M. tuberculosis pathogenic mechanism. Overall, CFP32 is an attractive target for drug and vaccine design as well as new diagnostic strategies
— id: 78842, year: 2003, vol: 71, page: 6871, stat: Journal Article,

Specificity and diversity of antibodies to Mycobacterium tuberculosis arabinomannan
Navoa, Josephine Anne D; Laal, Suman; Pirofski, Liise-Anne; McLean, Gary R; Dai, Zhongdong; Robbins, John B; Schneerson, Rachel; Casadevall, Arturo; Glatman-Freedman, Aharona
2003 Jan;10(1):88-94, Clinical & diagnostic laboratory immunology
Arabinomannan (AM) is a polysaccharide antigen of the mycobacterial capsule. However, it is uncertain whether AM constitutes an immunologically distinct fraction of Mycobacterium tuberculosis. In this study, we analyzed the repertoire and specificity of antibodies to AM by using AM-binding murine monoclonal antibodies (MAbs) and human serum samples. Murine MAbs were found to be diverse in their specificity to AM and cross-reactivity with other arabinose-containing mycobacterial polysaccharides, with MAb 9d8 binding exclusively to AM. Human antibodies to AM were detected in serum samples from patients with pulmonary tuberculosis (TB), as well as in those from healthy, purified protein derivative-negative controls, with significantly higher titers among patients. The binding of human antibodies to AM was inhibited by MAb 9d8 in three patients with TB but not in controls. MAb 5c11, which recognizes other mycobacterial arabinose-containing carbohydrates in addition to AM, inhibited the binding of serum samples from 75% of patients and 76% of controls. Analysis of human antibodies with murine MAbs to human V(H) determinants demonstrated diversity among antibodies to AM with qualitative and quantitative differences compared with antibodies to lipoarabinomannan. In summary, our study suggests that antibodies to AM are diverse and heterogeneous with respect to antigen recognition and V(H) determinant expression, with human serum samples containing different subsets of antibodies to AM with the specificities of AM-binding murine MAbs. One MAb and a subset of human antibodies bind AM specifically, suggesting that this polysaccharide is antigenically distinct and is expressed in human infection
— id: 78843, year: 2003, vol: 10, page: 88, stat: Journal Article,

Combined use of serum and urinary antibody for diagnosis of tuberculosis
Singh, Krishna K; Dong, Yuxin; Hinds, Laura; Keen, Marc A; Belisle, John T; Zolla-Pazner, Susan; Achkar, Jacqueline M; Nadas, Arthur J; Arora, Vijay K; Laal, Suman
2003 Aug 1;188(3):371-377, Journal of infectious diseases
Efforts to devise immunoassays for tuberculosis (TB) that can be adapted to rapid formats are ongoing. The present study was aimed at determining whether urinary anti-Mycobacterium tuberculosis antibodies are present in patients with TB, to evaluate the feasibility of developing a urine antibody-based diagnostic test. Urinary antibodies directed against the culture filtrate proteins of M. tuberculosis, MPT 32, and the 81-kDa GlcB protein were detectable in patients with TB, although the sensitivity of antibody detection was lower (53%-64%), compared with serum antibodies (68%-77%). Surprisingly, with all 3 antigens, the use of paired serum and urine samples provided higher sensitivities of antibody detection than either single specimen, and anti-GlcB antibodies were present in the serum and/or urine of 39 (90%) of 43 smear-positive patients with TB. Although, with the current methods and antigens, the level of sensitivity is insufficient to design a urinary antibody diagnostic test, these studies provide the foundation for further studies on the development of a urine antibody-based immunoassay for TB
— id: 39136, year: 2003, vol: 188, page: 371, stat: Journal Article,

Homogeneity of antibody responses in tuberculosis patients
Samanich K; Belisle JT; Laal S
2001 Jul;69(7):4600-4609, Infection & immunity
The goals of the present study were twofold: (i) to compare the repertoires of antigens in culture filtrates of in vitro-grown Mycobacterium tuberculosis that are recognized by antibodies from noncavitary and cavitary tuberculosis (TB) patients and (ii) to determine the extent of variation that exists between the antigen profiles recognized by individual TB patients. Lipoarabinomannan-free culture filtrate proteins of M. tuberculosis were fractionated by one-dimensional (1-D) and 2-D polyacrylamide gel electrophoresis, and the Western blots were probed with sera from non-human immunodeficiency virus (non-HIV)-infected cavitary and noncavitary TB patients and from HIV-infected, noncavitary TB patients. In contrast to earlier studies based on recombinant antigens of M. tuberculosis which suggested that antibody responses in TB patients were heterogeneous (K. Lyashchenko et al., 1998, Infect. Immun. 66:3936-3940, 1998), our studies with native culture filtrate proteins show that the antibody responses in TB patients show significant homogeneity in being directed against a well-defined subset of antigens. Thus, there is a well-defined subset of culture filtrate antigens that elicits antibodies during noncavitary and cavitary disease. In addition, another set of antigens is recognized primarily by cavitary TB patients. The mapping with individual patient sera presented here suggests that serodiagnostic tests based on the subset of antigens recognized during both noncavitary and cavitary TB will enhance the sensitivity of antibody detection in TB patients, especially in difficult-to-diagnose, smear-negative, noncavitary TB patients
— id: 20613, year: 2001, vol: 69, page: 4600, stat: Journal Article,

Antigens of Mycobacterium tuberculosis Expressed during Preclinical Tuberculosis: Serological Immunodominance of Proteins with Repetitive Amino Acid Sequences
Singh KK; Zhang X; Patibandla AS; Chien P Jr; Laal S
2001 Jun;69(6):4185-4191, Infection & immunity
Four antigens of Mycobacterium tuberculosis that are expressed in vivo after aerosol infection but prior to the development of clinical tuberculosis (TB) in rabbits were identified by immunoscreening of an expression library of M. tuberculosis genomic DNA with sera obtained 5 weeks postinfection. Three of the proteins identified, PirG (Rv3810), polymorphic GC-repetitive sequence (PE-PGRS; Rv3367), and proline-threonine repetitive protein (PTRP) (Rv0538), have multiple tandem repeats of unique amino acid sequences and have characteristics of surface or secreted proteins. The fourth protein, MtrA (Rv3246c), is a response regulator of a putative two-component signal transduction system, mtrA-mtrB, of M. tuberculosis. All four antigens were recognized by pooled sera from TB patients and not from healthy controls, confirming their in vivo expression during active infection in humans. Three of the antigens (PE-PGRS, PTRP, and MtrA) were also recognized by retrospective preclinical TB sera obtained, prior to the clinical manifestation of TB, from human immunodeficiency virus-TB patients, suggesting that they are potential candidates for devising diagnostic tests for active, preclinical TB
— id: 20669, year: 2001, vol: 69, page: 4185, stat: Journal Article,

HIV phenotype correlates with the relative amounts of lymphocyte function-related molecule 1 (LFA-1) and major histocompatibility complex (MHC) class II in the virion envelope [In Process Citation]
Bastiani Lallos L; Cecilia D; Fenyo EM; Laal S; Zolla-Pazner S
2000 Jul 28;14(11):1523-1531, AIDS
OBJECTIVE: The biological phenotype of HIV-1 has been associated with various aspects of its infectivity, including syncytium formation and coreceptor usage. Adhesion molecules, present on both the target cell and the virus, have also been shown to play a role in the infectious process. A possible correlation between the presence of adhesion molecules in the envelope of HIV-1 with the biological phenotype of the virus is examined. DESIGN: The envelopes of 56 isolates of HIV-1 of known biological phenotype were analyzed for the presence of lymphocyte function-related molecule 1 (LFA-1) and major histocompatibility complex (MHC) class II molecules. METHODS: The coreceptor usage of each isolate was determined in a GHOST cell or a U87.CD4 infectivity assay. The presence of LFA-1 and MHC class II in each virus envelope was then determined using a virus-binding enzyme-linked immunosorbent assay (ELISA). RESULTS: Viruses using the chemokine receptor CCR5 have relatively higher levels of MHC class II than LFA-1 in their envelopes compared with those using CXCR4. CONCLUSIONS: The finding that there is a differential incorporation of MHC class II and LFA-1 molecules by CXCR4- and CCR5-using viruses augments the list of properties contributing to the biological phenotype of HIV-1. This may explain, in part, how CXCR4-using viruses are able to bind to and infect a broader range of cell types than CCR5-using viruses, and why CXCR4-using viruses are associated with a more aggressive disease course
— id: 9226, year: 2000, vol: 14, page: 1523, stat: Journal Article,

Serodiagnostic potential of culture filtrate antigens of Mycobacterium tuberculosis
Samanich KM; Keen MA; Vissa VD; Harder JD; Spencer JS; Belisle JT; Zolla-Pazner S; Laal S
2000 Jul;7(4):662-668, Clinical & diagnostic laboratory immunology
Our studies of the humoral responses of tuberculosis (TB) patients have defined the repertoire of culture filtrate antigens of Mycobacterium tuberculosis that are recognized by antibodies from cavitary and noncavitary TB patients and demonstrated that the profile of antigens recognized changes with disease progression (K. Samanich et al., J. Infect. Dis. 178:1534-1538, 1998). We have identified several antigens with strong serodiagnostic potential. In the present study we have evaluated the reactivity of cohorts of human immunodeficiency virus (HIV)-negative, smear-positive; HIV-negative, smear-negative; and HIV-infected TB patients, with three of the candidate antigens, an 88-kDa protein, antigen (Ag) 85C, and MPT32, and compared the reactivity of the same patient cohort with the 38-kDa antigen and Ag 85A. We have also compared the reactivity of native Ag 85C and MPT32 with their recombinant counterparts. The evaluation of the reactivity was done by a modified enzyme-linked immunosorbent assay described earlier (S. Laal et al., Clin. Diag. Lab. Immunol. 4:49-56, 1997), in which all sera are preadsorbed against Escherichia coli lysates to reduce the levels of cross-reactive antibodies. Our results demonstrate that (i) antigens identified on the basis of their reactivity with TB patients' sera provide high sensitivities for serodiagnosis, (ii) recombinant Ag 85C and MPT32, expressed in E. coli, show reduced reactivity with human TB sera, and (iii) of the panel of antigens tested, the 88-kDa protein is the most promising candidate for serodiagnosis of TB in HIV-infected individuals. Moreover, these results reaffirm that both the extent of the disease and the bacterial load may play a role in determining the antigen profile recognized by antibodies
— id: 11616, year: 2000, vol: 7, page: 662, stat: Journal Article,

Exclusion of HIV coreceptors CXCR4, CCR5, and CCR3 from the HIV envelope
Lallos LB; Laal S; Hoxie JA; Zolla-Pazner S; Bandres JC
1999 Jul 1;15(10):895-897, AIDS research & human retroviruses
Both HIV-1 primary isolates and laboratory strains incorporate cell-derived molecules into their envelopes depending on the host cell in which they are grown. This incorporation is not random and, specifically, HIV-1 has been shown to select against the incorporation into its surface of CD4, its main receptor. In this study, we have looked at the incorporation of HIV coreceptors CXCR4, CCR5, and CCR3 into the HIV envelope. For this purpose, we grew HIV-1 primary isolate BZ167 in several cell lines and PBMCs, and the envelope profiles of the resulting viruses were determined with a virus-binding ELISA. While the virus particle gained several molecules when passed through the different cell lines (e.g., ICAM-3, LFA-1, ICAM-1, or MHC class II), BZ167 never incorporated significant levels of CXCR4, CCR5, or CCR3 into its envelope even though some or all of the cell lines in which it was grown expressed them. These results show that HIV-1 selects against the incorporation of these chemokine receptors into its envelope molecule, as it does against the incorporation of CD4
— id: 6159, year: 1999, vol: 15, page: 895, stat: Journal Article,

Selective exclusion of HIV coreceptors from HIV virions
Bastiani L; Laal S; Hoxie JA; Zolla-Pazner S; Bandres JC
1998 Feb 1-5;5:85-85, Conference on Retroviruses & Opportunistic Infections
Both primary isolates and laboratory strains of HIV-1 incorporate different cell-derived molecules into their envelopes depending on the host cell in which they are grown. This incorporation in not random; for example, HIV-1 has been shown to exclude CD4 from its envelope. In this study we have examined the incorporation into the envelope of HIV-1 of three coreceptors -- CXCR4,CCR5, and CCR3-- as well as CD4. For this purpose, the HIV-1 SI primary isolate BZ167 was passaged into PHA-stimulated PBMC and CEM-SS cells and the incorporation of various cell-derived molecules into the virion envelope was determined with a virus binding ELISA. As previously shown, BZ167 grown in PHA-stimulated PBMC expressed most of the adhesion molecules tested (such as LFA-1, ICAM-1, and MHC class II), while CEM-SS-grown BZ167 did not express significant levels of any of these adhesion molecules. CEM-SS cells express both CD4 and CXCR4 whereas PHA-stimulated PBMCs express CD4, CXCR4, CCR5, and CCR3. In contrast, BZ167 grown in either cell type lacked CD4 as well as the coreceptors, CXCR4, CCR5, and CCR3
— id: 6008, year: 1998, vol: 5, page: 85, stat: Journal Article,

Delineation of human antibody responses to culture filtrate antigens of Mycobacterium tuberculosis
Samanich KM; Belisle JT; Sonnenberg MG; Keen MA; Zolla-Pazner S; Laal S
1998 Nov;178(5):1534-1538, Journal of infectious diseases
This study was undertaken to define the antigens in culture filtrates of actively replicating Mycobacterium tuberculosis that are recognized by antibodies from tuberculosis (TB) patients. Two-dimensional Western blots were probed with sera from healthy controls and TB patients that were preabsorbed with Escherichia coli lysates to deplete cross-reactive antibodies. Antibodies from TB patients recognized 26 of the >100 culture filtrate proteins, and the repertoire changed with disease progression. Only 12 of 26 antigens, including 3 proteins implicated in colonization and invasion by mycobacteria (MPT51, MPT32, and 85C), and 9 (as yet undefined proteins) were reactive with sera from TB patients with early noncavitary or cavitary disease. Eight additional antigens, including 4 undefined proteins, were recognized only by sera from a subset of patients with advanced cavitary disease. Studies suggest that 3 of the antigens recognized by sera from patients with early TB (85C, MPT32, and a 88-kDa protein) have strong serodiagnostic potential
— id: 7777, year: 1998, vol: 178, page: 1534, stat: Journal Article,

Host cell-dependent alterations in envelope components of human immunodeficiency virus type 1 virions
Bastiani L; Laal S; Kim M; Zolla-Pazner S
1997 May;71(5):3444-3450, Journal of virology
In addition to gp41 and gp120, an array of cell adhesion molecules is present on the envelope of human immunodeficiency virus type 1 (HIV-1). To examine the role of the host cell in the acquisition of these molecules by virions, both laboratory-adapted and primary isolates were sequentially passaged into different host cells. Viruses obtained from the various host cells were examined for the presence of 10 different cell-derived molecules by a virus binding enzyme-linked immunosorbent assay. Virus progeny raised in peripheral blood mononuclear cells expressed most of the adhesion molecules tested, with the level of LFA-1 being the highest. When viruses were passaged into CEM-SS or SupT1 cells, the expression of most of the adhesion molecules on the virus envelope was lost. In contrast, when viruses were passaged into MT2 cells, the virus progeny bore high levels of LFA-3, ICAM-1, and major histocompatibility complex classes I and II. These studies demonstrate for the first time the host cell dependence of the adhesion molecule profile present on the envelope of primary isolates of HIV-1. The presence of several adhesion molecules that have not previously been identified as components of the envelope of either laboratory or primary isolates is also described. In addition, we show that the adhesion molecule profile of the virions is acquired, or lost, within one passage and is maintained with subsequent passages in the same cell type
— id: 9243, year: 1997, vol: 71, page: 3444, stat: Journal Article,

Host cell-dependent alterations in HIV envelope components
Bastiani L; Laal S; Zolla-Pazner S
1997 Jan 22-26;4:148-148, Conference on Retroviruses & Opportunistic Infections
We have recently demonstrated that HIV(IIIB), when grown in different host cells, carried different adhesion molecules in its envelope. In the current study, we have examined the acquisition of adhesion molecules by primary isolates of HIV-1 under similar circumstances. Five PBMC-grown primary isolates of HIV-1 were passaged through CEM-SS, MT2, or SupT1 cells and back passaged in PBMC. The adhesion molecule profile on the progeny virions from each host cell was determined with a virus binding ELISA. The parental and back passaged primary isolates expressed high levels of LFA-1, ICAM-1, ICAM-3, and MHC class II. The profile on the envelope of the MT2-grown virions was altered in that they now bear high levels of LFA-3, ICAM-1, and MHC class I and II. In contrast, few of the adhesion molecules were detected on virions from CEM-SS or SupT1 cells. These alterations in the envelope profile of adhesion molecules were detected after a single passage and were maintained with additional passages in the same cell line. Since adhesion molecules play a critical role in virus-cell interactions, these studies suggest that the tropism of HIV-1 for various cells may be critically dependent on the host cell in which the virion was produced
— id: 6002, year: 1997, vol: 4, page: 148, stat: Journal Article,

Surrogate marker of preclinical tuberculosis in human immunodeficiency virus infection: antibodies to an 88-kDa secreted antigen of Mycobacterium tuberculosis
Laal S; Samanich KM; Sonnenberg MG; Belisle JT; O'Leary J; Simberkoff MS; Zolla-Pazner S
1997 Jul;176(1):133-143, Journal of infectious diseases
Antibodies to purified, size-fractionated secreted proteins of Mycobacterium tuberculosis in sera from patients with human immunodeficiency virus (HIV) infection and active tuberculosis (HIV/TB patients), and in stored sera obtained from the same patients prior to clinical manifestation of TB, were evaluated by ELISA, and the repertoire of antigens recognized was analyzed by immunoblotting. Compared with non-HIV/TB patients, HIV/TB patients had lower levels of anti-mycobacterial antibodies, and these were directed toward a restricted set of antigens. Antibodies to an 88-kDa secreted antigen were present in the sera of 74% of HIV/TB patients during the years (1.5-6) prior to manifestation of active, clinical tuberculosis, although only 66% were positive by the time tuberculosis was diagnosed. The presence of antibodies to the 88-kDa antigen can serve as a surrogate marker for identifying HIV-infected persons with active, subclinical disease who are at a high risk of developing clinical tuberculosis
— id: 7953, year: 1997, vol: 176, page: 133, stat: Journal Article,

Human humoral responses to antigens of Mycobacterium tuberculosis: immunodominance of high-molecular-mass antigens
Laal S; Samanich KM; Sonnenberg MG; Zolla-Pazner S; Phadtare JM; Belisle JT
1997 Jan;4(1):49-56, Clinical & diagnostic laboratory immunology
The selection of antigens of Mycobacterium tuberculosis for most studies of humoral responses in tuberculosis patients has been restricted to molecules that were either immunodominant in immunized animals or amenable to biochemical purification rather than those that were reactive with the human immune system. Delineation of antigens that elicit humoral responses during the natural course of disease progression in humans has been hindered by the presence of cross-reactive antibodies to conserved regions on ubiquitous prokaryotic antigens in sera from healthy individuals and tuberculosis patients. The levels of cross-reactive antibodies in the sera were reduced by preadsorption with Escherichia coli lysates, prior to studying their reactivity against a large panel of M. tuberculosis antigens to which the human immune system may be exposed during natural infection and disease. Thus, reactivity against pools of secreted, cellular, and cell wall-associated antigens of M. tuberculosis was assessed by an enzyme-linked immunosorbent assay (ELISA). Initial results suggested that the secreted protein preparation contained antigens most frequently recognized by the humoral responses of pulmonary tuberculosis patients. The culture filtrate proteins were subsequently size fractionated by preparative polyacrylamide gel electrophoresis, characterized by reaction with murine monoclonal antibodies to known antigens of M. tuberculosis by an ELISA, and assessed for reactivity with tuberculous and nontuberculous sera. Results show that a secreted antigen of 88 kDa elicits a strong antibody response in a high percentage of patients with pulmonary tuberculosis. This and other antigens identified on the basis of their reactivity with patient sera may prove useful for developing serodiagnosis for tuberculosis
— id: 9247, year: 1997, vol: 4, page: 49, stat: Journal Article,

Humoral responses in tuberculosis
Laal, Suman
Tuberculosis Boston : Little Brown, 1996,
— id: 4834, year: 1996, vol: , page: ?, stat: Chapter,

Synergistic neutralization of human immunodeficiency virus type 1 by combinations of human monoclonal antibodies
Laal S; Burda S; Gorny MK; Karwowska S; Buchbinder A; Zolla-Pazner S
1994 Jun;68(6):4001-4008, Journal of virology
The ability of antibodies to the V3 region and the CD4-binding domain (CD4bd) of human immunodeficiency virus type 1 (HIV-1) to act in synergy to neutralize HIV has been demonstrated previously. However, synergy between antibodies to other HIV-1 epitopes has not been studied. We have used 21 combinations of human monoclonal antibodies (MAbs) directed against different epitopes of the gp120 and gp41 proteins of HIV-1 to evaluate their ability to act in synergy to neutralize HIV-1. Combinations of anti-V3 and anti-CD4bd antibodies, anti-V3 and anti-gp120 C-terminus antibodies, anti-CD4bd and anti-C-terminus antibodies, anti-V3 and anti-gp41 antibodies, and anti-CD4bd and anti-gp41 antibodies were tested. Our results show that some, but not all anti-V3 antibodies can act in synergy with anti-CD4bd antibodies. In addition, for the first time, antibodies to the C-terminus region have been found to act in synergy with the anti-CD4bd antibodies. Various anti-CD4bd MAbs also act in synergy when used together. The use of such cocktails of human MAbs for passive immunization against HIV-1 may prove to be important for therapy in postexposure settings and for prevention of maternal-fetal transmission of the virus. The results also provide information on the types of antibodies that should be elicited by an effective vaccine
— id: 9263, year: 1994, vol: 68, page: 4001, stat: Journal Article,

Human monoclonal antibodies to HIV-1 define synergistic activities leading to enhanced neutralization
Zolla-Pazner S; Laal S; Burda S; Buchbinder A
1994 ;46:38-47, Antibiotics & chemotherapy
— id: 9266, year: 1994, vol: 46, page: 38, stat: Journal Article,

Synergistic interaction of various pairs of human monoclonal antibodies in the neutralization of HIV-1
Laal S; Burda S; Buchbinder A; Zolla-Pazner S
1993 Dec 12-16;1:67-67, National Conference on Human Retroviruses & Related Infections
Studies have shown that combinations of anti-V3 region and anti-CD4 binding domain (CD4bd) monoclonal antibodies are synergistic for neutralization of HIV-1. We have used a panel of human monoclonal antibodies (humAb) directed against various immunogenic regions on the HIV envelope, to perform a systematic analysis of the epitopes that must be targeted simultaneously to achieve enhanced neutralization. The panel included 2 humAbs to the gp120-V3 region, 3 to the gp120-CD4bd, 2 to the gp120-C-terminal region, 1 each to gp41 cluster I (aa571-641) and II (aa644-663), and 1 (2F5 from H. Kattinger) to a gp41 region adjacent to cluster II (aa662-667). The assay used was the modified syncytium formation microassay (Nara et al., Laal et al.) that measures cell free virus neutralization. The results show that a) not all anti-V3 and anti-CD4bd humAb pairs show synergy; b) anti-CD4bd humAbs can act in synergy both with anti-V3 and anti-C-terminus humAbs; c) anti-V3 humAb 447-D is able to act in synergy with 5 different anti-CD4bd humAbs; d)none of the anti-gp41 humAbs is able to act in synergy with either anti-V3 or anti-CD4bd humAbs. These results have important implications for rational design of regimens for active and passive immunization against HIV
— id: 6012, year: 1993, vol: 1, page: 67, stat: Journal Article,

A rapid, automated microtiter assay for measuring neutralization of HIV-1
Laal S; Burda S; Sharpe S; Zolla-Pazner S
1993 Aug;9(8):781-785, AIDS research & human retroviruses
A sensitive, rapid, and quantitative ELISA for p24 is described, which can be used as the read-out to test for HIV-1 neutralization in the syncytium-forming microassay and can replace the counting of syncytia under the microscope. This assay can be used reliably for divergent strains of HIV-1, including those that do not induce syncytia. The new read-out permits evaluation of neutralization at 3 days rather than 5 days. Using this assay, the neutralizing activity of several new and previously described human monoclonal antibodies against HIV-1 was characterized
— id: 9271, year: 1993, vol: 9, page: 781, stat: Journal Article,

Antibodies to M. tuberculosis antigens in HIV-1 infected individuals
Laal S; Pazner SZ
1993 Dec 12-16;1:189-189, National Conference on Human Retroviruses & Related Infections
Identification of molecules of M. tuberculosis which are expressed and immunogenic in the context of a dysfunctional immune system may provide insights into the pathogenesis of reactivation of latent tuberculosis. Use of such antigens (and antibodies to them) for the design of serodiagnostic tests may yield specific and sensitive assays that could be used for screening populations at high risk of reactivation. Antibodies to antigens of M. tuberculosis have been studied in the sera of HIV-seropositive individuals with confirmed clinical pulmonary TB. Sera were obtained from these individuals (a) several months prior to clinical manifestations of TB (b) at the time of clinical diagnosis of TB and (c) post-diagnosis. Unfractionated total cellular proteins from four strains of M. tuberculosis obtained from HIV-TB patients, and grown after minimal passaging in the laboratory, were used as antigen. The sera were depleted of antibodies to cross-reactive, ubiquitous bacterial antigens prior to being tested in an ELISA employing the pooled M. tuberculosis antigen preparation. Results show that antibodies to M. tuberculosis were detectable several months prior to clinical TB in 7/10 individuals. Only 4/10 patients had anti-M. tuberculosis antibodies at the time of clinical diagnosis of tuberculosis, and the proportion increased (9/15) after initiation of anti-tuberculosis therapy. The identity of the M. tuberculosis antigens identified by these patients is being analyzed by SDS-PAGE and immunoblotting
— id: 6013, year: 1993, vol: 1, page: 189, stat: Journal Article,

Epitopes of HIV-1 glycoproteins recognized by the human immune system
Laal S; Zolla-Pazner S
1993 ;56:91-111, Chemical immunology
— id: 9274, year: 1993, vol: 56, page: 91, stat: Journal Article,

Recombinant fusion protein identified by lepromatous sera mimics native Mycobacterium leprae in T-cell responses across the leprosy spectrum
Laal S; Sharma YD; Prasad HK; Murtaza A; Singh S; Tangri S; Misra RS; Nath I
1991 Feb 1;88(3):1054-1058, Proceedings of the National Academy of Sciences of the United States of America
Pooled polyvalent sera from lepromatous leprosy patients were used to screen a lambda gt11 recombinant DNA expression library of Mycobacterium leprae in order to identify the relevant antigens recognized by the human immune response. Of the 300,000 phages screened, 4 clones were identified that coded for fusion proteins of the same molecular mass. The fusion protein from clone LSR2 was tested for immunoreactivity in assays using peripheral blood cells and sera from 11 laboratory personnel and 105 patients across the leprosy spectrum. LSR2 protein appears to be predominantly a T-cell antigen. It evokes similar lymphoproliferative responses as the native bacillus both at the individual level and in the leprosy spectrum as a whole. Though only 50% of patient sera with anti-M. leprae antibodies reacted with the fusion protein, the pattern of reactivity in the antibody responses was also similar for the various clinical types. The coding regions of clones LSR1 and LSR2 are identical. They show no homology with sequences stored in data banks and encode a protein of 89 amino acids with a calculated molecular mass of approximately 10 kDa
— id: 8355, year: 1991, vol: 88, page: 1054, stat: Journal Article,

Isolation and serological characterization of a Plasmodium vivax recombinant antigen
Sharma YD; Sharma VP; Ray P; Laal S; Sawant SD; Verma S
1991 Jun;59(6):1922-1926, Infection & immunity
A genomic library for Plasmodium vivax was constructed in lambda gt11 and immunologically screened with pooled serum samples from vivax patients. Six seroreactive clones were isolated, and one clone, denoted PV9, was studied further. This clone has an unusual base composition (65% G + C), does not share any homology with P. falciparum, and codes for an entirely new antigenic determinant. Antibodies (immunoglobulin G type) against the PV9-encoded polypeptide were produced in all vivax patients older than 15 years. This seroreactivity was lower among patients younger than 15 years (53%). The antigenic epitope(s) of the PV9-encoded polypeptide was recognized at a similar rate by serum samples from P. vivax patients who were living 350 to 973 km apart. Fifty percent of uninfected Indian adults were also seropositive, whereas all European and American (United States) sera tested were negative, suggesting that anti-PV9 antibodies persist after infection. The seroreactivity pattern of this antigen is similar to that of the immunity developed in malaria after repeated infections
— id: 15085, year: 1991, vol: 59, page: 1922, stat: Journal Article,

Nucleotide sequence and deduced amino acid sequence of Mycobacterium leprae gene showing homology to bacterial atp operon
Nath I; Laal S
1990 Aug 25;18(16):4935-4935, Nucleic acids research
— id: 15087, year: 1990, vol: 18, page: 4935, stat: Journal Article,

Human immune response to recombinant interferon gamma and protein antigen LSR2
Nath I; Laal S; Sivasai KS; Tangri S; Murtaza A; Singh S; Wilfred D; Misra RS
1990 Sep;41(3):324-325, Tropical medicine & parasitology
— id: 15086, year: 1990, vol: 41, page: 324, stat: Journal Article,

The nature and kinetics of a delayed immune response to purified protein derivative of tuberculin in the skin of lepromatous leprosy patients
Kaplan G; Laal S; Sheftel G; Nusrat A; Nath I; Mathur NK; Mishra RS; Cohn ZA
1988 Nov 1;168(5):1811-1824, Journal of experimental medicine
We have analyzed the nature and kinetics of a delayed, cell-mediated immune response to a purified protein derivative of tuberculin (PPD) in the skin of 154 naturally sensitized patients with lepromatous leprosy. After the intradermal injection of 5 U of PPD, biopsies were taken at 1-21 d and studied for the composition, extent, persistence, and organization of the emigratory cell response by light and electron microscopy. Induration of positive sites occurred promptly, reached a maximum diameter at 4 d, displayed a major extravasatory element, and was evident for as long as 21 d. The cellularity of the site exhibited a biphasic course, reached a maximum at 7 d, involved as much as 70% of the dermis and millions of new cells, and was elevated threefold above preinjection levels at 21 d. The emigratory cells were limited to T cells and circulating monocytes. T cells were more evident as they entered a preexisting lepromatous lesion containing parasitized macrophages and only occasional T cells many of the CD8+ phenotype. The predominant emigratory T cell was CD4+ although CD8+ cells were in evidence. The CD4/CD8 ratio of the lesions started at less than unity and in two distinct steps reached levels as high as 5:1. In most sites CD4+ cells were in the majority at 21 d. A well-defined granulomatous response with epithelioid and giant cells was apparent at 4 d, reached a maximum at 7 d, and involved all PPD sites at this time point. The generation of these differentiated mononuclear phagocytes from newly emigrated monocytes was never observed in the underlying lepromatous lesion but is a constant feature of the tuberculoid leprosy response. Epidermal thickening and keratinocyte proliferation, sequellae of the dermal reaction, reached a maximum at 7 d and gradually resolved by 3 wk. A constant feature of the PPD response was the extensive destruction of preexisting macrophages containing Mycobacterium leprae bacilli or their products. This was associated with the presence of and intimate contact with highly polarized lymphoid cells of unknown phenotype. Cell destruction did not involve other elements of the dermis and spared parasitized Schwann cells. Newly emigrated T cells and monocytes were never seen within the perineural sheath in contact with neural elements. It appears that a single antigenic stimulus leads to a very long-term, defined series of events with distinct temporal patterns. It includes waves of emigratory T cells, the maturation and organization of monocytes, the generation of killer cells, and the extensive destruction of parasitized macrophages.(ABSTRACT TRUNCATED AT 400 WORDS)
— id: 15088, year: 1988, vol: 168, page: 1811, stat: Journal Article,

Epidermal changes in reactional leprosy: keratinocyte Ia expression as an indicator of cell-mediated immune responses
Thangaraj H; Laal S; Thangaraj I; Nath I
1988 Sep;56(3):401-407, International journal of leprosy & other mycobacterial diseases
Significant epidermal changes were observed in lesions of leprosy patients undergoing type 1 (reversal) and type 2 (erythema nodosum leprosum, ENL) reactions. Using indirect immunofluorescence and frozen sections stained with the appropriate monoclonal antibodies, an increase in epidermal cell layers, the presence of Ia on keratinocytes, an increase in Langerhans' cell numbers, and scattered T cells within the epidermis were seen in both types of reactions. Although borderline tuberculoid patients with type 1 reactions showed the consistent presence of Ia on all keratinocytes, lepromatous patients undergoing ENL reactions showed only a patchy distribution. Taken together, these studies indicate that local T-cell activation leading to the production of terminal lymphokine, such as interferon-gamma, with subsequent induction of Ia on epidermal cells may be an important event in reactional leprosy states. It is of interest that the hitherto considered 'anergic' lepromatous patients should recover temporary T-cell reactivity during the natural course of the disease
— id: 15089, year: 1988, vol: 56, page: 401, stat: Journal Article,

Type 1 reactions in leprosy--heterogeneity in T-cell functions related to the background leprosy type
Laal S; Mishra RS; Nath I
1987 Sep;55(3):481-493, International journal of leprosy & other mycobacterial diseases
Nineteen each of paucibacillary borderline tuberculoid (BT) and multibacillary borderline borderline (BB)/borderline lepromatous (BL) leprosy patients undergoing type 1 reactions were compared with nonreactional stable patients of the appropriate leprosy type. In the BT reactional group, both phytohemagglutinin-induced and, more importantly, antigen-induced lymphoproliferation was reduced in 80%-90% of the patients. On the other hand, leukocyte migration inhibition was reduced in 40% and remained unchanged in the others. Suppressor-cell activity as evaluated by a costimulant assay was also reduced in a majority of the reactional BT individuals. In contrast, the bacilliferous BB and BL patients in reaction showed significant general improvement in leukocyte migration inhibition (p less than 0.001) and antigen-induced lymphoproliferation (p less than 0.05) as compared to the expected hyporesponsive/anergic uncomplicated BB-BL patients. Suppressor-cell activity also recovered during the reactional phase. However, no significant differences were observed in either of the reactional or stable leprosy types in the numbers of total T cells (OKT3+) and their subsets as defined by OKT4+ (helper/inducer) and OKT8+ (suppressor/cytotoxic) functional phenotypes. Moreover, during type 1 reactions the 48-hr delayed-type hypersensitivity (DTH) responses after intradermal injection of Mycobacterium leprae antigens continued to reflect the background leprosy type rather than the functional perturbations in the circulating T cells. Only a marginal increase in DTH was observed in some BT reactional individuals. No consistent pattern in the above in vitro T-cell-related responses was discernable in the same individuals 4-6 months after subsidence of reactions. The clinical entity of type 1 reactions encompassing paucibacillary and multibacillary leprosy shows a heterogeneity/dichotomy in T-cell responses which may reflect different immunological mechanisms underlying the reactional state
— id: 15090, year: 1987, vol: 55, page: 481, stat: Journal Article,

Influence of delayed immune reactions on human epidermal keratinocytes
Kaplan G; Witmer MD; Nath I; Steinman RM; Laal S; Prasad HK; Sarno EN; Elvers U; Cohn ZA
1986 May;83(10):3469-3473, Proceedings of the National Academy of Sciences of the United States of America
The epidermal changes that occur in human cutaneous immune responses have been investigated in the tuberculin reaction and in the lesions of tuberculoid and lepromatous leprosy and cutaneous leishmaniasis. In each situation, there was a dermal accumulation of monocytes and T cells, and the epidermis exhibited thickening. In the tuberculin response, the thickness of the epidermis sometimes doubled in 48-72 hr, and this was attributed to increases in both size and number of keratinocytes. In addition, the phenotype of the keratinocytes changed from Ia- to Ia+. Similar changes in keratinocyte Ia-antigen expression occurred in the epidermis overlying untreated tuberculoid leprosy and cutaneous leishmaniasis lesions, but not in lepromatous leprosy. We suggest that one or more epidermal growth factors may be generated in the course of a delayed immune reaction in the dermis
— id: 15091, year: 1986, vol: 83, page: 3469, stat: Journal Article,

Natural emergence of antigen-reactive T cells in lepromatous leprosy patients during erythema nodosum leprosum
Laal S; Bhutani LK; Nath I
1985 Dec;50(3):887-892, Infection & immunity
Fifteen lepromatous leprosy (LL) patients undergoing erythema nodosum leprosum (ENL) reactions were compared with 13 stable, uncomplicated, anergic individuals of the same leprosy background. ENL patients showed significant antigen-induced leukocyte migration inhibition (migration index = 0.058 +/- 0.01), paralleling the values obtained with a responder tuberculoid leprosy population (migration index = 0.04 +/- 0.004). Both phytohemagglutinin-induced general T-cell proliferation and, more significantly, antigen-induced lymphoproliferation were enhanced during the acute phase of the reaction. Suppressor cell activity, monitored by a costimulant assay, showed enhanced antigen-stimulated suppression of mitogen responses. Interestingly, the improvement in in vitro T-cell responses was not reflected in dermal reactivity, since 48-h delayed-type hypersensitivity responses after intradermal injection of soluble Mycobacterium leprae antigens continued to be poor. After subsidence of reactional lesions, leukocyte migration inhibition, lymphoproliferation, and suppressor cell activity were reduced to the unresponsive state seen in stable LL patients. Significantly, perturbations of T-cell reactivity are detectable in ENL reactions, indicating the natural but transient emergence of antigen-induced T cells in LL
— id: 15092, year: 1985, vol: 50, page: 887, stat: Journal Article,

Differences in predominant T cell phenotypes and distribution pattern in reactional lesions of tuberculoid and lepromatous leprosy
Narayanan RB; Laal S; Sharma AK; Bhutani LK; Nath I
1984 Mar;55(3):623-628, Clinical & experimental immunology
The nature and histological pattern of the cutaneous infiltrates of 17 leprosy patients in reversal reactions (Type I) and erythema nodosum leprosum (Type II, ENL) were compared with tissues from 18 non-reactional borderline leprosy (BT, BL) and lepromatous leprosy (LL) patients using monoclonal antibodies and immunofluorescence. Reactional BT lesions showed a mild increase in OKT11+ pan T cells as compared to non-reactional tissues and a significant influx of OKT8+ (suppressor/cytotoxic) cells which were peripherally localized in the lymphocyte mantle surrounding the epithelioid cells. The Leu 3a+ (helper/inducer) cells were scattered amongst the lymphocytes and macrophages. The mean ratio (+/- s.d.) of Leu 3a+/OKT8+ cells was 1.88 +/- 0.64 in Type I BT reactions as compared to 2.95 +/- 0.95 in BT lesions. In contrast, lesions of BL reversal reactions and ENL showed a more marked increase in pan T cells with a preponderance of the helper/inducer subset, Leu 3a+/OKT8+ ratio being 2.26 +/- 0.61 and 0.93 +/- 0.57 in BL reactional and non-reactional lesions, respectively. Interestingly, this increase in the numbers of the T cells reached levels observed in BT lesions. The distribution pattern of OKT8+ cells was similar to Leu 3a+, both being diffusely scattered amongst the bacilli laden macrophages. Ia like antigens were present in all granulomas and were abundant on lymphocytes and macrophages and less conspicuous on epithelioid cells. T6+ Langerhans cells were uniformly increased in all reactional lesions. It would appear that the changes observed in both Type I and Type II reactions are similar in the lepromatous group of patients. They differ significantly from the BT reversal reaction in terms of the dominant T cell subset and the microanatomical distribution of the OKT8+ cells in the lesions
— id: 15093, year: 1984, vol: 55, page: 623, stat: Journal Article,