Challenges and advances in serological and molecular tests to aid leprosy diagnosis.

Leprosy is a neglected chronic infectious disease caused by obligate intracellular bacilli, Mycobacterium leprae and Mycobacterium lepromatosis. Despite multidrug therapy (MDT) success, leprosy accounts for more than 200,000 new cases yearly. Leprosy diagnosis remains based on the dermato-neurologic examination, but histopathology of skin biopsy and bacilloscopy of intradermal scraping are subsidiary diagnostic tests that require expertise and laboratory infrastructure. This minireview summarizes the state of the art of serologic tests to aid leprosy diagnosis, highlighting enzyme-linked immunosorbent assay (ELISA) and point-of-care tests (POCT) biotechnologies. Also, the impact of the postgenomic era on the description of new recombinantly expressed M. leprae-specific protein antigens, such as leprosy Infectious Disease Research Institute (IDRI) diagnostic (LID)-1 is summarized. Highly specific and sensitive molecular techniques to detect M. leprae DNA as the quantitative polymerase chain reaction (qPCR) and the loop-mediated isothermal amplification (LAMP) are briefly reviewed. Serology studies using phenolic glycolipid-I (PGL-I) semi-synthetic antigens, LID-1 fusion antigen, and the single fusion complex natural disaccharide-octyl (NDO)-LID show high sensitivity in multibacillary (MB) patients. However, serology is not applicable to paucibacillary patients, as they have weak humoral response and robust cell-mediated response, requiring tests for cellular biomarkers. Unlike ELISA-based tests, leprosy-specific POCT based on semi-synthetic PGL-I antigens and NDO-LID 1 antigen is easy to perform, cheaper, equipment-free, and can contribute to early diagnosis avoiding permanent incapacities and helping to interrupt M. leprae transmission. Besides its use to help diagnosis of household contacts or at-risk populations in endemic areas, potential applications of leprosy serology include monitoring MDT efficacy, identification of recent infection, especially in young children, as surrogate markers of disease progression to orient adult chemoprophylaxis and as a predictor of type 2 leprosy reactions. Advances in molecular biology techniques have reduced the complexity and execution time of qPCR confirming its utility to help diagnosis while leprosy-specific LAMP holds promise as an adjunct test to detect M. leprae DNA.

Prediction of the occurrence of leprosy reactions based on Bayesian networks.

Introduction: Leprosy reactions (LR) are severe episodes of intense activation of the host inflammatory response of uncertain etiology, today the leading cause of permanent nerve damage in leprosy patients. Several genetic and non-genetic risk factors for LR have been described; however, there are limited attempts to combine this information to estimate the risk of a leprosy patient developing LR. Here we present an artificial intelligence (AI)-based system that can assess LR risk using clinical, demographic, and genetic data. Methods: The study includes four datasets from different regions of Brazil, totalizing 1,450 leprosy patients followed prospectively for at least 2 years to assess the occurrence of LR. Data mining using WEKA software was performed following a two-step protocol to select the variables included in the AI system, based on Bayesian Networks, and developed using the NETICA software. Results: Analysis of the complete database resulted in a system able to estimate LR risk with 82.7% accuracy, 79.3% sensitivity, and 86.2% specificity. When using only databases for which host genetic information associated with LR was included, the performance increased to 87.7% accuracy, 85.7% sensitivity, and 89.4% specificity. Conclusion: We produced an easy-to-use, online, free-access system that identifies leprosy patients at risk of developing LR. Risk assessment of LR for individual patients may detect candidates for close monitoring, with a potentially positive impact on the prevention of permanent disabilities, the quality of life of the patients, and upon leprosy control programs.

A social network approach for the study of leprosy transmission beyond the household.

BACKGROUND: Mycobacterium leprae was the first microorganism directly associated with a disease, however, there are still important gaps in our understanding of transmission. Although household contacts are prioritized, there is evidence of the importance of extrahousehold contacts. The goal of this article is to contribute to our understanding of the transmission of leprosy ex-household. METHODS: We compare co-location data of 397 leprosy cases and 211 controls drawn from the Centro de Dermatologia Sanitária D. Libânia in Fortaleza, Brazil. We collected lifetime geolocation data related to residence, school attendance and workplace and developed novel methods to establish a critical distance (Rc) for exposure and evaluated the potential for transmission for residence, school and workplace. RESULTS: Our methods provide different threshold values of distance for residence, school and workplace. Residence networks demonstrate an Rc of about 500 m. Cases cluster in workplaces as well. Schools do not cluster cases. CONCLUSIONS: Our novel network approach offers a promising opportunity to explore leprosy transmission. Our networks confirm the importance of coresidence, provide a boundary and suggest a role for transmission in workplaces. Schools, on the other hand, do not demonstrate a clustering of cases. Our findings may have programmatic relevance.

Human Genetic Susceptibility of Leprosy Recurrence.

Host genetic susceptibility to leprosy has been intensively investigated over the last decades; however, there are no studies on the role of genetic variants in disease recurrence. A previous initiative identified three recurrent cases of leprosy for which none of the M. leprae strains, as obtained in the first and the second diagnosis, had any known genomic variants associated to resistance to Multidrug therapy; in addition, whole genome sequencing indicated that the same M. leprae was causing two out of the three recurrences. Thus, these individuals were suspected of being particularly susceptible to M. leprae infection, either as relapse or reinfection. To verify this hypothesis, 19 genetic markers distributed across 11 loci (14 genes) classically associated with leprosy were genotyped in the recurrent and in three matching non-recurrent leprosy cases. An enrichment of risk alleles was observed in the recurrent cases, suggesting the existence of a particularly high susceptibility genetic profile among leprosy patients predisposing to disease recurrence.

Mycobacterium leprae-Specific Antibodies in Multibacillary Leprosy Patients Decrease During and After Treatment With Either the Regular 12 Doses Multidrug Therapy (MDT) or the Uniform 6 Doses MDT.

Leprosy serology reflects the bacillary load of patients and multidrug therapy (MDT) reduces Mycobacterium leprae-specific antibody titers of multibacillary (MB) patients. The Clinical Trial for Uniform Multidrug Therapy Regimen for Leprosy Patients in Brazil (U-MDT/CT-BR) compared outcomes of regular 12 doses MDT/R-MDT and the uniform 6 doses MDT/U-MDT for MB leprosy, both of regimens including rifampicin, clofazimine, and dapsone. This study investigated the impact of R-MDT and U-MDT and the kinetic of antibody responses to M. leprae-specific antigens in MB patients from the U-MDT/CT-BR. We tested 3,400 serum samples from 263 MB patients (R-MDT:121; U-MDT:142) recruited at two Brazilian reference centers (Dona Libânia, Fortaleza, Ceará; Alfredo da Matta Foundation, Manaus, Amazonas). Enzyme-linked immunosorbent assays with three M. leprae antigens [NT-P-BSA: trisaccharide-phenyl of phenollic glycolipid-I antigen (PGL-I); LID-1: Leprosy Infectious Disease Research Institute Diagnostic 1 di-fusion recombinant protein; and ND-O-LID: fusion complex of disaccharide-octyl of PGL-I and LID-1] were performed using around 13 samples per patient. Samples were collected at baseline/M0, during MDT (R-MDT:M1-M12 months, U-MDT:M1-M6 months) and after MDT discontinuation (first, second year). Statistical significance was assessed by the Mann-Whitney U test for comparison between groups (p values < 0.05). Mixed effect multilevel regression analyses were used to investigate intraindividual serological changes overtime. In R-MDT and U-MDT groups, males predominated, median age was 41 and 40.5 years, most patients were borderline lepromatous and lepromatous leprosy (R-MDT:88%, U-MDT: 90%). The bacilloscopic index at diagnosis was similar (medians: 3.6 in the R-MDT and 3.8 in the U-MDT group). In R-MDT and U-MDT groups, a significant decline in anti-PGL-I positivity was observed from M0 to M5 (p = 0.035, p = 0.04, respectively), from M6 to M12 and at the first and second year posttreatment (p < 0.05). Anti-LID-1 antibodies declined from M0 to M6 (p = 0.024), M7 to M12 in the R-MDT; from M0 to M4 (p = 0.003), M5 to M12 in the U-MDT and posttreatment in both groups (p > 0.0001). Anti-ND-O-LID antibodies decreased during and after treatment in both groups, similarly to anti-PGL-I antibodies. Intraindividual serology results in R-MDT and U-MDT patients showed that the difference in serology decay to all three antigens was dependent upon time only. Our serology findings in MB leprosy show that regardless of the duration of the U-MDT and R-MDT, both of them reduce M. leprae-specific antibodies during and after treatment. In leprosy, antibody levels are considered a surrogate marker of the bacillary load; therefore, our serological results suggest that shorter U-MDT is also effective in reducing the patients' bacillary burden similarly to R-MDT. Clinical Trial Registration: ClinicalTrials.gov, NCT00669643.

Clinical trial for uniform multidrug therapy for leprosy patients in Brazil (U-MDT/CT-BR): adverse effects approach.

BACKGROUND: The Clinical Trial for Uniform Multidrug Therapy for Leprosy Patients in Brazil (U-MDT/CT-BR), designed to evaluate the effectiveness of a six-months regimen, assessed the adverse effects caused by the drugs. OBJECTIVE: Describe adverse effects due to MDT in U-MDT/CT-BR, comparing the uniform regimen (U-MDT) to the current WHO regimen (R-MDT). PATIENTS AND METHODS: After operational classification, patients were randomly allocated to the study groups. U-MDT PB and U-MDT MB groups, received the U-MDT regimen, six doses of MB-MDT (rifampicin, dapsone and clofazimine). R-MDT PB and R-MDT MB groups, received the WHO regimens: six doses (rifampicin and dapsone) for PB and 12 doses (rifampicin, dapsone and clofazimine) for MB. During treatment, patients returned monthly for clinical and laboratorial evaluation. Patients with single lesion were not included in this trial. RESULTS: Skin pigmentation (21.7%) and xerosis (16.9%) were the most frequent complaints among 753 patients. Laboratory exams showed hemoglobin concentration lower than 10g/dL in 23.3% of the patients, glutamic oxaloacetic transaminase (GOT) above 40U/L in 29.5% and glutamic pyruvic transaminase (GPT) above 40U/L in 28.5%. Twenty-four patients (3.2%) stopped dapsone intake due to adverse effects, of whom 16.6% due to severe anemia. One case of sulfone syndrome was reported. STUDY LIMITATIONS: Loss of some monthly laboratory sample collection. CONCLUSIONS: There was no statistical difference regarding adverse effects in the R-MDT and U-MDT groups but anemia was greater in patients from R-MDT/MB group, therefore adverse effects do not represent a constraint to recommend the six-month uniform regimen of treatment for all leprosy patients.

Clinical trial for uniform multidrug therapy for leprosy patients in Brazil (U-MDT/CT-BR): adverse effects approach

Abstract: BACKGROUND: The Clinical Trial for Uniform Multidrug Therapy for Leprosy Patients in Brazil (U-MDT/CT-BR), designed to evaluate the effectiveness of a six-months regimen, assessed the adverse effects caused by the drugs. OBJECTIVE: Describe adverse effects due to MDT in U-MDT/CT-BR, comparing the uniform regimen (U-MDT) to the current WHO regimen (R-MDT). Patients and methods: After operational classification, patients were randomly allocated to the study groups. U-MDT PB and U-MDT MB groups, received the U-MDT regimen, six doses of MB-MDT (rifampicin, dapsone and clofazimine). R-MDT PB and R-MDT MB groups, received the WHO regimens: six doses (rifampicin and dapsone) for PB and 12 doses (rifampicin, dapsone and clofazimine) for MB. During treatment, patients returned monthly for clinical and laboratorial evaluation. Patients with single lesion were not included in this trial. RESULTS: Skin pigmentation (21.7%) and xerosis (16.9%) were the most frequent complaints among 753 patients. Laboratory exams showed hemoglobin concentration lower than 10g/dL in 23.3% of the patients, glutamic oxaloacetic transaminase (GOT) above 40U/L in 29.5% and glutamic pyruvic transaminase (GPT) above 40U/L in 28.5%. Twenty-four patients (3.2%) stopped dapsone intake due to adverse effects, of whom 16.6% due to severe anemia. One case of sulfone syndrome was reported. STUDY LIMITATIONS: Loss of some monthly laboratory sample collection. CONCLUSIONS: There was no statistical difference regarding adverse effects in the R-MDT and U-MDT groups but anemia was greater in patients from R-MDT/MB group, therefore adverse effects do not represent a constraint to recommend the six-month uniform regimen of treatment for all leprosy patients.

Phylogenomics and antimicrobial resistance of the leprosy bacillus Mycobacterium leprae.

Leprosy is a chronic human disease caused by the yet-uncultured pathogen Mycobacterium leprae. Although readily curable with multidrug therapy (MDT), over 200,000 new cases are still reported annually. Here, we obtain M. leprae genome sequences from DNA extracted directly from patients' skin biopsies using a customized protocol. Comparative and phylogenetic analysis of 154 genomes from 25 countries provides insight into evolution and antimicrobial resistance, uncovering lineages and phylogeographic trends, with the most ancestral strains linked to the Far East. In addition to known MDT-resistance mutations, we detect other mutations associated with antibiotic resistance, and retrace a potential stepwise emergence of extensive drug resistance in the pre-MDT era. Some of the previously undescribed mutations occur in genes that are apparently subject to positive selection, and two of these (ribD, fadD9) are restricted to drug-resistant strains. Finally, nonsense mutations in the nth excision repair gene are associated with greater sequence diversity and drug resistance.

Phylogenomics and antimicrobial resistance of the leprosy bacillus Mycobacterium leprae

Leprosy is a chronic human disease caused by the yet-uncultured pathogen Mycobacterium leprae. Although readily curable with multidrug therapy (MDT), over 200,000 new cases are still reported annually. Here, we obtain M. leprae genome sequences from DNA extracted directly from patients' skin biopsies using a customized protocol. Comparative and phylogenetic analysis of 154 genomes from 25 countries provides insight into evolution and antimicrobial resistance, uncovering lineages and phylogeographic trends, with the most ancestral strains linked to the Far East. In addition to known MDT-resistance mutations, we detect other mutations associated with antibiotic resistance, and retrace a potential stepwise emergence of extensive drug resistance in the pre-MDT era. Some of the previously undescribed mutations occur in genes that are apparently subject to positive selection, and two of these (ribD, fadD9) are restricted to drug-resistant strains. Finally, nonsense mutations in the nth excision repair gene are associated with greater sequence diversity and drug resistance.

Leprosy: current situation, clinical and laboratory aspects, treatment history and perspective of the uniform multidrug therapy for all patients

Abstract: In this review, the most relevant and current epidemiological data, the main clinical, laboratory and therapeutical aspects of leprosy are presented. Detailed discussion of the main drugs used for leprosy treatment, their most relevant adverse effects, evolution of the therapeutic regimen, from dapsone as a monotherapy to the proposed polychemotherapy by World Health Organization (WHO) can be found in this CME. We specifically highlight the drug acceptability, reduction in treatment duration and the most recent proposal of a single therapeutic regimen, with a fixed six months duration, for all clinical presentations, regardless of their classification.

Uniform multidrug therapy for leprosy patients in Brazil (U-MDT/CT-BR): Results of an open label, randomized and controlled clinical trial, among multibacillary patients.

BACKGROUND: Leprosy control is based on early diagnosis and multidrug therapy. For treatment purposes, leprosy patients can be classified as paucibacillary (PB) or multibacillary (MB), according to the number of skin lesions. Studies regarding a uniform treatment regimen (U-MDT) for all leprosy patients have been encouraged by the WHO, rendering disease classification unnecessary. METHODOLOGY AND FINDINGS: An independent, randomized, controlled clinical trial conducted from 2007 to 2015 in Brazil, compared main outcomes (frequency of reactions, bacilloscopic index trend, disability progression and relapse rates) among MB patients treated with a uniform regimen/U-MDT (dapsone+rifampicin+clofazimine for six months) versus WHO regular-MDT/R-MDT (dapsone+rifampicin+clofazimine for 12 months). A total of 613 newly diagnosed, untreated MB patients with high bacterial load were included. There was no statistically significant difference in Kaplan-Meyer survival function regarding reaction or disability progression among patients in the U-MDT and R-MDT groups, with more than 25% disability progression in both groups. The full mixed effects model adjusted for the bacilloscopic index average trend in time showed no statistically significant difference for the regression coefficient in both groups and for interaction variables that included treatment group. During active follow up, four patients in U-MDT group relapsed representing a relapse rate of 2.6 per 1000 patients per year of active follow up (95% CI [0·81, 6·2] per 1000). During passive follow up three patients relapsed in U-MDT and one in R-MTD. As this period corresponds to passive follow up, sensitivity analysis estimated the relapse rate for the entire follow up period between 2·9- and 4·5 per 1000 people per year. CONCLUSION: Our results on the first randomized and controlled study on U-MDT together with the results from three previous studies performed in China, India and Bangladesh, support the hypothesis that UMDT is an acceptable option to be adopted in endemic countries to treat leprosy patients in the field worldwide. TRIAL REGISTRATION: ClinicalTrials.gov: NCT00669643.

Whole genome sequencing distinguishes between relapse and reinfection in recurrent leprosy cases.

BACKGROUND: Since leprosy is both treated and controlled by multidrug therapy (MDT) it is important to monitor recurrent cases for drug resistance and to distinguish between relapse and reinfection as a means of assessing therapeutic efficacy. All three objectives can be reached with single nucleotide resolution using next generation sequencing and bioinformatics analysis of Mycobacterium leprae DNA present in human skin. METHODOLOGY: DNA was isolated by means of optimized extraction and enrichment methods from samples from three recurrent cases in leprosy patients participating in an open-label, randomized, controlled clinical trial of uniform MDT in Brazil (U-MDT/CT-BR). Genome-wide sequencing of M. leprae was performed and the resultant sequence assemblies analyzed in silico. PRINCIPAL FINDINGS: In all three cases, no mutations responsible for resistance to rifampicin, dapsone and ofloxacin were found, thus eliminating drug resistance as a possible cause of disease recurrence. However, sequence differences were detected between the strains from the first and second disease episodes in all three patients. In one case, clear evidence was obtained for reinfection with an unrelated strain whereas in the other two cases, relapse appeared more probable. CONCLUSIONS/SIGNIFICANCE: This is the first report of using M. leprae whole genome sequencing to reveal that treated and cured leprosy patients who remain in endemic areas can be reinfected by another strain. Next generation sequencing can be applied reliably to M. leprae DNA extracted from biopsies to discriminate between cases of relapse and reinfection, thereby providing a powerful tool for evaluating different outcomes of therapeutic regimens and for following disease transmission.

Leprosy reactions: The predictive value of Mycobacterium leprae-specific serology evaluated in a Brazilian cohort of leprosy patients (U-MDT/CT-BR).

BACKGROUND: Leprosy reactions, reversal reactions/RR and erythema nodosum leprosum/ENL, can cause irreversible nerve damage, handicaps and deformities. The study of Mycobacterium leprae-specific serologic responses at diagnosis in the cohort of patients enrolled at the Clinical Trial for Uniform Multidrug Therapy Regimen for Leprosy Patients in Brazil/U-MDT/CT-BR is suitable to evaluate its prognostic value for the development of reactions. METHODOLOGY: IgM and IgG antibody responses to PGL-I, LID-1, ND-O-LID were evaluated by ELISA in 452 reaction-free leprosy patients at diagnosis, enrolled and monitored for the development of leprosy reactions during a total person-time of 780,930 person-days, i.e. 2139.5 person-years, with a maximum of 6.66 years follow-up time. PRINCIPAL FINDINGS: Among these patients, 36% (160/452) developed reactions during follow-up: 26% (119/452) RR and 10% (41/452) had ENL. At baseline higher anti-PGL-I, anti-LID-1 and anti-ND-O-LID seropositivity rates were seen in patients who developed ENL and RR compared to reaction-free patients (p<0.0001). Seroreactivity in reactional and reaction-free patients was stratified by bacilloscopic index/BI categories. Among BI negative patients, higher anti-PGL-I levels were seen in RR compared to reaction-free patients (p = 0.014). In patients with 0

Leprosy: current situation, clinical and laboratory aspects, treatment history and perspective of the uniform multidrug therapy for all patients.

In this review, the most relevant and current epidemiological data, the main clinical, laboratory and therapeutical aspects of leprosy are presented. Detailed discussion of the main drugs used for leprosy treatment, their most relevant adverse effects, evolution of the therapeutic regimen, from dapsone as a monotherapy to the proposed polychemotherapy by World Health Organization (WHO) can be found in this CME. We specifically highlight the drug acceptability, reduction in treatment duration and the most recent proposal of a single therapeutic regimen, with a fixed six months duration, for all clinical presentations, regardless of their classification.

Uniform multidrug therapy for leprosy patients in Brazil (U-MDT/CT-BR): results of an open label, randomized and controlled clinical trial, among multibacillary patients

BACKGROUND: Leprosy control is based on early diagnosis and multidrug therapy. For treatment purposes, leprosy patients can be classified as paucibacillary (PB) or multibacillary (MB), according to the number of skin lesions. Studies regarding a uniform treatment regimen (U-MDT) for all leprosy patients have been encouraged by the WHO, rendering disease classification unnecessary. METHODOLOGY AND FINDINGS: An independent, randomized, controlled clinical trial conducted from 2007 to 2015 in Brazil, compared main outcomes (frequency of reactions, bacilloscopic index trend, disability progression and relapse rates) among MB patients treated with a uniform regimen/U-MDT (dapsone+rifampicin+clofazimine for six months) versus WHO regular-MDT/R-MDT (dapsone+rifampicin+clofazimine for 12 months). A total of 613 newly diagnosed, untreated MB patients with high bacterial load were included. There was no statistically significant difference in Kaplan-Meyer survival function regarding reaction or disability progression among patients in the U-MDT and R-MDT groups, with more than 25% disability progression in both groups. The full mixed effects model adjusted for the bacilloscopic index average trend in time showed no statistically significant difference for the regression coefficient in both groups and for interaction variables that included treatment group. During active follow up, four patients in U-MDT group relapsed representing a relapse rate of 2.6 per 1000 patients per year of active follow up (95% CI [0·81, 6·2] per 1000). During passive follow up three patients relapsed in U-MDT and one in R-MTD. As this period corresponds to passive follow up, sensitivity analysis estimated the relapse rate for the entire follow up period between 2·9- and 4·5 per 1000 people per year. CONCLUSION: Our results on the first randomized and controlled study on U-MDT together with the results from three previous studies performed in China, India and Bangladesh, support the hypothesis that UMDT is an acceptable option to be adopted in endemic countries to treat leprosy patients in the field worldwide.

Can baseline ML Flow test results predict leprosy reactions? An investigation in a cohort of patients enrolled in the uniform multidrug therapy clinical trial for leprosy patients in Brazil.

BACKGROUND: The predictive value of the serology to detection of IgM against the Mycobacterium leprae-derived phenolic glycolipid-I/PGL-I to identify leprosy patients who are at higher risk of developing reactions remains controversial. Whether baseline results of the ML Flow test can predict leprosy reactions was investigated among a cohort of patients enrolled in The Clinical Trial for Uniform Multidrug Therapy for Leprosy Patients in Brazil (U-MDT/CT-BR). METHODS: This was a descriptive study focusing on the main clinical manifestations of leprosy patients enrolled in the U-MDT/CT-BR from March 2007 to February 2012 at two Brazilian leprosy reference centers. For research purposes, 753 leprosy patients were categorized according to a modified Ridley-Jopling (R&J) classification and according to the development of leprosy reactions (reversal reaction/RR and erythema nodosum leprosum/ENL), and whether they had a positive or negative bacillary index/BI. RESULTS: More than half of the patients (55.5 %) reported leprosy reaction: 18.3 % (138/753) had a RR and 5.4 % (41/753) had ENL. Leprosy reactions were more frequent in the first year following diagnosis, as seen in 27 % (205/753) of patients, while 19 % (142/753) developed reactions during subsequent follow-up. Similar frequencies of leprosy reactions and other clinical manifestations were observed in paucibacillary (PB) and multibacillary (MB) leprosy patients treated with U-MDT and regular MDT (R-MDT) (P = 0.43 and P = 0.61, respectively). Compared with PB patients, leprosy reactions were significantly more frequent in MB patients with a high BI, and more patients developed RR than ENL. However, RR and neuritis were also reported in patients with a negative BI. At baseline, the highest rate of ML Flow positivity was observed in patients with a positive BI, especially those who developed ENL, followed by patients who had neuritis and RR. Among reaction-free patients, 81.9 % were ML Flow positive, however, the differences were not statistically significant compared to reactional patients (P = 0.45). CONCLUSIONS: MB and PB patients treated with R-MDT and U-MDT showed similar frequencies of RR and other clinical manifestations. Positive ML Flow tests were associated with MB leprosy and BI positivity. However, ML Flow test results at baseline showed limited sensitivity and specificity for predicting the development of leprosy reactions.

Could Mycobacterium leprae Infection Be an Occupational Disease? A Survey in Healthcare Workers From an Endemic Area in the Amazonian Region.

A serologic survey was conducted to evaluate the prevalence of Mycobacterium leprae infection among healthcare workers and associated factors. Of 280 workers, 26 (9.3%) were positive using immunoglobulin M serology for PGL-I M. leprae antigen. Exposure to leprosy patients in the workplace was significantly associated with seropositivity (P=.044).

Description of leprosy classification at baseline among patients enrolled at the uniform multidrug therapy clinical trial for leprosy patients in Brazil.

The uniform multidrug therapy clinical trial, Brazil (U-MDT/CT-BR), database was used to describe and report the performance of available tools to classify 830 leprosy patients as paucibacillary (PB) and multibacillary (MB) at baseline. In a modified Ridley and Jopling (R&J) classification, considering clinical features, histopathological results of skin biopsies and the slit-skin smear bacterial load results were used as the gold standard method for classification. Anti-phenolic glycolipid-I (PGL-I) serology by ML Flow test, the slit skin smear bacterial load, and the number of skin lesions were evaluated. Considering the R&J classification system as gold standard, ML Flow tests correctly allocated 70% patients in the PB group and 87% in the MB group. The classification based on counting the number of skin lesions correctly allocated 46% PB patients and 99% MB leprosy cases. Slit skin smears properly classified 91% and 97% of PB and MB patients, respectively. Based on U-MDT/CT-BR results, classification of leprosy patients for treatment purposes is unnecessary because it does not impact clinical and laboratories outcomes. In this context, the identification of new biomarkers to detect patients at a higher risk to develop leprosy reactions or relapse remains an important research challenge.

Phage display and synthetic peptides as promising biotechnological tools for the serological diagnosis of leprosy.

BACKGROUND: The diagnosis of leprosy is primarily based on clinical manifestations, and there is no widely available laboratory test for the early detection of this disease, which is caused by Mycobacterium leprae. In fact, early detection and treatment are the key elements to the successful control of leprosy. METHODOLOGY/PRINCIPAL FINDINGS: Peptide ligands for antibodies from leprosy patients were selected from phage-displayed peptide libraries. Three peptide sequences expressed by reactive phage clones were chemically synthesized. Serological assays that used synthetic peptides were evaluated using serum samples from leprosy patients, household contacts (HC) of leprosy patients, tuberculosis patients and endemic controls (EC). A pool of three peptides identified 73.9% (17/23) of multibacillary (MB) leprosy patients using an enzyme-linked immunosorbent assay (ELISA). These peptides also showed some seroreactivities to the HC and EC individuals. The peptides were not reactive to rabbit polyclonal antisera against the different environmental mycobacteria. The same peptides that were conjugated to the carrier protein bovine serum albumin (BSA) induced the production of antibodies in the mice. The anti-peptide antibodies that were used in the Western blotting analysis of M. leprae crude extracts revealed a single band of approximately 30 kDa in one-dimensional electrophoresis and four 30 kDa isoforms in the two-dimensional gel. The Western blotting data indicated that the three peptides are derived from the same bacterial protein. CONCLUSIONS/SIGNIFICANCE: These new antigens may be useful in the diagnosis of MB leprosy patients. Their potentials as diagnostic reagents must be more extensively evaluated in future studies using a large panel of positive and negative sera. Furthermore, other test approaches using peptides should be assessed to increase their sensitivity and specificity in detecting leprosy patients. We have revealed evidence in support of phage-displayed peptides as promising biotechnological tools for the design of leprosy diagnostic serological assays.