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.

Comparison of antiphenolic glycolipid-1 antibody levels in seropositive contacts of leprosy after 2 years of single-dose rifampicin as postexposure prophylaxis.

Background: Leprosy is still a global problem, especially in developing countries, including Indonesia. Ineffective prevention of leprosy leads to active transmission of the disease. World Health Organization (WHO) recommend post-exposure prophylaxis (PEP) with single dose of rifampicin (SDR) for leprosy patients. Previous study showed protective effect of SDR against leprosy, especially for the first 2 years. Hence, the use of PEP and IgM anti PGL-1 examination are required to suspend the chain of leprosy transmission. This study evaluated the effectiveness of SDR administration by comparing IgM anti-PGL-1 antibody levels in seropositive household contacts before and after 2 years of SDR administration. Methods: Analytical observational laboratory study comparing IgM anti PGL-1 antibody levels before and after 2 years of SDR administration in leprosy contacts, with a prospective follow-up study design. We conducted this study from December 2022 to January 2023 at Dr. Mohammad Hoesin General Hospital Palembang. All seropositive household contacts of leprosy who had been administrated SDR 2 years ago were included, then PGL-1 antibody levels were examined. Results: The use of SDR showed significant improvement in leprosy contacts after 2 years (P=0.000). The median antibody level before SDR administration was 1,209.20 (615.81 - 4,353.60), which decrease to 146.03 (0 - 2,487.80) U/mL after 2 years. There was statistically significant relationship between history of BCG vaccination (P=0.003) and IgM PGL-1 antibody levels after 2 years of SDR administration. Conclusion: There is a significant decrease in IgM anti PGL-1 antibody levels among leprosy contacts after 2 years of SDR chemoprophylaxis administration.

High seropositivity against NDO-LID in a group of household contacts of leprosy patients. Are we close to leprosy elimination in Colombia?

INTRODUCTION: Leprosy is a chronic infectious disease caused by two mycobacteria (Mycobacterium leprae and Mycobacterium lepromatosis). The household contacts (HHC) of leprosy index cases are at higher risk of being infected with these mycobacteria. Therefore, serological testing in HHC would be an effective strategy to eliminate leprosy in Colombia. OBJECTIVE: To determine the seroprevalence and factors associated with the infection by M. leprae in HHC. METHODS: An observational study was conducted in 428 HHC located in the Colombian Caribbean, Andean, Pacific, and Amazonian regions. We evaluated the seropositivity and titrations of IgM, IgG, and protein A against NDO-LID. RESULTS: The evaluated HHC showed high seropositivity, precisely 36.9% anti-NDO-LID IgM, 28.3% anti-NDO-LID IgG, and 47.7% protein A. Furthermore, Protein A showed a greater capacity to detect infected individuals than other anti-NDO-LID conjugates (p < 0.0001). This study did not show differences in the seropositivity according to sex or age of the HHC (p > 0.05). Higher seropositivity for IgM was evidenced mainly in HHC located in the Colombian Pacific region (p 0.001). This research did not show differences in the seropositivity for these serological tests between HHC of PB or MB leprosy patients (p > 0.05). CONCLUSION: Leprosy transmission is still active between Colombian HHC. Consequently, controlling leprosy transmission in this population is fundamental to eradicating this disease.

Factors associated with the development of leprosy in Brazilian contacts: a systematic review.

People who interact with leprosy patients in their environment, neighborhood, family, or social relationships are at risk to develop the disease. This systematic review investigated the risk and protective factors associated with the development of leprosy in Brazilian contacts. The studies were found in Cochrane Library, PubMed (MEDLINE), Embase, Virtual Health Library, grey literature and hand search until July 2021. The study selection, data extraction and quality assessment were independently performed by two investigators. The quality assessment was performed using the Newcastle-Ottawa Scale (NOS). This review was registered in PROSPERO (CRD42020160680). Seventeen articles fulfilled the inclusion criteria (n=544). The immunological and molecular factors, such as Anti-phenolic Glycolipid Antibodies (Anti-PGL-1) seropositivity, negative Mitsuda test, absence of Bacillus Calmette-Guérin (BCG) scar, positive Polymerase Chain Reaction (PCR) in blood; age and race; conviviality, education, contact time and type of contact, as well as elements related to the index case (bacilloscopic index; genetic conditions, family relationships), and some combined factors were shown to be relevant risk factors associated with the development of the disease in Brazilian leprosy contacts. The protective factors reported were the presence of one or more BCG scars, positive Mitsuda test, and education level. All selected studies were considered of high quality according to NOS. The knowledge of disease-related risk and protective factors provides the scientific basis for decision-making in the management of the disease in leprosy contacts.

Design of a specific peptide against phenolic glycolipid-1 from Mycobacterium leprae and its implications in leprosy bacilli entry.

BACKGROUND: Mycobacterium leprae, the causative agent of Hansen's disease, causes neural damage through the specific interaction between the external phenolic glycolipid-1 (PGL-1) and laminin subunit alpha-2 (LAMA2) from Schwann cells. OBJECTIVE: To design a LAMA2-based peptide that targets PGL-1 from M. leprae. METHODS: We retrieved the protein sequence of human LAMA2 and designed a specific peptide using the Antimicrobial Peptide Database and physicochemical parameters for antimycobacterial peptide-lipid interactions. We used the AlphaFold2 server to predict its three-dimensional structure, AUTODOCK-VINA for docking, and GROMACS programs for molecular dynamics simulations. FINDINGS: We analysed 52 candidate peptides from LAMA2, and subsequent screening resulted in a single 60-mer peptide. The mapped peptide comprises four ß-sheets and a random coiled region. This peptide exhibits a 45% hydrophobic ratio, in which one-third covers the same surface. Molecular dynamics simulations show that our predicted peptide is stable in aqueous solution and remains stable upon interaction with PGL-1 binding. In addition, we found that PGL-1 has a preference for one of the two faces of the predicted peptide, which could act as the preferential binding site of PGL-1. MAIN CONCLUSIONS: Our LAMA2-based peptide targeting PGL-1 might have the potential to specifically block this key molecule, suggesting that the preferential region of the peptide is involved in the initial contact during the attachment of leprosy bacilli to Schwann cells.

Arginase 1 is a marker of protection against illness in contacts of leprosy patients.

Leprosy household contacts are generally more prone to develop the disease compared to the general population. Previous studies have demonstrated that genes related to the alternative activation (M2) profile in macrophages are associated with the increased bacillary load in multibacillary leprosy patients (MB), and that contacts of MB patients have a higher risk of contracting the disease. In addition, positive serological responses to PGL-1 or LID-1 are associated with a higher risk of disease. We performed a 5-year follow-up of contacts of leprosy patients and evaluated the pattern of gene and protein expression in cells from contacts that developed leprosy during this period. Leprosy household contacts had decreased soluble CD163 and heme oxygenase 1 (HO-1) serum levels when compared with healthy donors and leprosy patients. In contrast, arginase 1 activities were higher in contacts when compared with both healthy donors and leprosy patients. Of the contacts, 33 developed leprosy during the follow-up. Gene expression analysis revealed reduced ARG1 expression in these contacts when compared with contacts that did not develop disease. Arginase activity was a good predictive marker of protection in contacts (sensitivity: 90.0%, specificity: 96.77%) and the association with serology for anti-PGL-1 and anti-LID-1 increased the sensitivity to 100%. Altogether, the data presented here demonstrate a positive role of arginase against leprosy and suggest that the evaluation of arginase activity should be incorporated into leprosy control programs in order to aid in the decision of which contacts should receive chemoprophylaxis.

Spatial distribution pattern of new leprosy cases under 15 years of age and their contacts in Sobral, Ceará, Brazil.

This study's objective was to analyze the spatial distribution pattern of new leprosy cases under 15 years and their contacts. A cross-sectional descriptive study covering sociodemographic characteristics and spatial analysis was carried out. The participants were from the city of Sobral, Ceará and the study was conducted between August 2014 and September 2015. Contacts were identified by the persons responsible for the children. Seropositivity was determined with the NDO-LID antigen, and positive cases were plotted on Voronoi polygons. Nine new cases of leprosy under 15 years of age have been found. The average number of people living with the cases was higher than the number of people living with non-household contacts. All household contacts were aware of other leprosy cases and had a higher rate of seropositive tests than non-household contacts. The index cases lived in the poorest regions of the municipality and hyper-endemic areas. Spatial analysis revealed a cluster of subclinical infection within a radius of 102 meters, suggesting that non-household transmission is related to proximity with seropositive individuals. In conclusion, the search for new leprosy cases cannot be restricted to household contacts.

Can anti-PGL-I antibody isotypes differentiate leprosy contacts and leprosy patients?

BACKGROUND: Serological tests for antibody measurement in leprosy have a series of limitations in discriminating contacts and patients. The present paper intends to evaluate if association of more than one antibody isotype in serum samples may be a useful tool in leprosy diagnosis. METHODS: This study evaluated 395 leprosy contacts and 71 leprosy index cases living in endemic municipalities in Northeastern Brazil. The participants were evaluated according to their anti-phenolic glycolipid antigen-I isotype (PGL-I) profile. Serum anti-PGL-I IgM, IgG, and IgA were measured by indirect ELISA. RESULTS: A strong association was found for antibody positivity in MB leprosy index cases. The odds ratios were 6.11 (95% CI 3.08 - 12.16) for IgM, 3.31 (1.66 - 6.61) for IgG, and 16.97 (8.39 - 34.2) for IgA. For IgM associated with one or more isotypes, the OR was 21.0 (95% CI 10.11 - 43.64), and for IgG + IgA, the OR was 17.58 (6.23 - 49.54). The highest diagnostic sensitivity of 76.0% (95% CI 61.8 - 86.9) was observed for IgM, and the lowest value was 24.1% (13.0 - 38.2), which was observed for IgG + IgA isotypes. Regarding presumptive positive predictive values, the lowest value was obtained for IgM at 24.7% (95% CI 18.1 - 32.3), and the highest values were observed for IgM+ one or more isotypes and for IgG + IgA isotype at 60.0% (44.3 - 74.3) and 66.7% (41.0 - 86.7), respectively. CONCLUSIONS: The present work demonstrated that by associating two or more positive antibody isotypes, the risk of facing a real case of leprosy may increase.

Detection of Mycobacterium leprae infection in family clusters from six leprosy-endemic regions in Colombia.

BACKGROUND: Leprosy is a chronic infectious disease caused by Mycobacterium leprae, which continues to be endemic in tropical countries, making it necessary to implement strategies for its elimination. The objective of the current article was to detect M. leprae infection and associated factors through serological and epidemiological evaluation in family clusters of leprosy patients. METHODS: Mycobacterium leprae infection was determined in 50 family clusters of leprosy patients from the departments of Bolívar, Atlántico, Santander, Boyacá, Chocó and Antioquia through the detection of antibodies (protein A, IgM, IgG) against anti-natural octyl disacharide-leprosy IDRI diagnostic (NDO-LID). RESULTS: Higher seroconversion and elevated titers of these antibodies against NDO-LID were observed in the population of Chocó and Atlántico (p<0.05). Additionally, a higher frequency of infection was observed in large family groups that consumed armadillo meat and belonged to a low socioeconomic stratum (p<0.05). Multivariate analysis established that the main associated factors for a family cluster experiencing this infection were belonging to a vulnerable economic stratum and a large family group. CONCLUSIONS: This study found that the set of social and demographic variables (i.e. armadillo consumption, geographic area, low socioeconomic status and belonging to a large family cluster) are related to the promotion of seropositivity in family clusters.

Laboratory diagnosis of leprosy: Two staining methods from bacilloscopy and rapid ml flow test.

Background: The diagnosis of leprosy is based on the characteristic signs and symptoms of the disease, subsidized by laboratory tests. When positive, the bacilloscopy closes the diagnosis for leprosy. Phenolic glycolipid-I, or PGL-I, is a molecule in the bacillus cell wall that confers a greater immune response. The ML Flow test is an immunochromatographic test for the detection of anti-PGL-I IgM in human blood or serum. Methods: A prospective study with data collection and biological materials in patients with suspected leprosy from August 2020 to May 2021. For microscopy, intradermal smears were stained with Auramine O, and after reading under a fluorescence microscope, reviewed by Ziehl-Neelsen. The ML flow test was performed according to the Bührer-Sékula protocol. To assess the agreement between the methods, the Kappa index was estimated. Results: Of the 94 suspected leprosy patients, 31 (32.9%) were diagnosed with leprosy. There was moderate agreement between the results of the ML Flow and Auramine O tests (Kappa = 0.58) and substantial agreement between the ML Flow and Ziehl-Neelsen microscopy (Kappa = 0.72). In paucibacillary cases, serology was positive in 100% of patients. Conclusions: This study concluded that the Ziehl-Neelsen technique remains the best option for standard leprosy staining, and the ML flow test is more positive among the three techniques evaluated and can be an effective tool in the early diagnosis of leprosy cases.

Detection of anti-M. leprae antibodies in children in leprosy-endemic areas: A systematic review.

BACKGROUND: Leprosy elimination primarily targets transmission of Mycobacterium leprae which is not restricted to patients' households. As interruption of transmission is imminent in many countries, a test to detect infected asymptomatic individuals who can perpetuate transmission is required. Antibodies directed against M. leprae antigens are indicative of M. leprae infection but cannot discriminate between active and past infection. Seroprevalence in young children, however, reflects recent M. leprae infection and may thus be used to monitor transmission in an area. Therefore, this literature review aimed to evaluate what has been reported on serological tests measuring anti-M. leprae antibodies in children without leprosy below the age of 15 in leprosy-endemic areas. METHODS AND FINDINGS: A literature search was performed in the databases Pubmed, Infolep, Web of Science and The Virtual Health Library. From the 724 articles identified through the search criteria, 28 full-text articles fulfilled all inclusion criteria. Two additional papers were identified through snowballing, resulting in a total of 30 articles reporting data from ten countries. All serological tests measured antibodies against phenolic glycolipid-I or synthetic derivatives thereof, either quantitatively (ELISA or UCP-LFA) or qualitatively (ML-flow or NDO-LID rapid test). The median seroprevalence in children in endemic areas was 14.9% and was stable over time if disease incidence remained unchanged. Importantly, seroprevalence decreased with age, indicating that children are a suitable group for sensitive assessment of recent M. leprae infection. However, direct comparison between areas, solely based on the data reported in these studies, was impeded by the use of different tests and variable cut-off levels. CONCLUSIONS: Quantitative anti-PGL-I serology in young children holds promise as a screening test to assess M. leprae infection and may be applied as a proxy for transmission and thereby as a means to monitor the effect of (prophylactic) interventions on the route to leprosy elimination.

Epitope mapping from Mycobacterium leprae proteins: Convergent data from in silico and in vitro approaches for serodiagnosis of leprosy.

Knowledge of immunodominant B-cell epitopes is essential to design powerful diagnostic strategies aiming for antibody detection. Outstanding progress in computational prediction has achieved a significant contribution to the biomedical fields, including immunodiagnosis. In silico analysis may have an even more important role when information concerning antigens from etiologic agents of neglected diseases, such as leprosy, is scarce. The aim of this study was to provide mapping of B-cell epitopes from two Mycobacterium leprae-derived antigens (Ag85B and ML2055), confirm their antigenicity, and to assess the ability of in silico immunoinformatics tools to accurately predict them. Linear B-cell epitopes predicted by ABCpred and SVMTrip servers were compared to antigenic regions of synthetic overlapping peptides that exhibited reactivity to antibodies from patients with leprosy. Our in vitro results identified several immunodominant regions that had also been indicated by in silico prediction, providing agreement between experimental and simulated data. After chemical synthesis, we used enzyme-linked immunosorbent assays to determine the effectiveness of the first identified sequence (GTNVPAEFLENFVHG) which had 72 % sensitivity and 78 % specificity (AUC = 0.79) while the second one (PVSSEAQPGDPNAPS) had 72 % sensitivity and 93.8 % specificity (AUC = 0.85). Using dot blotting, an easy-to-read visual test, both peptides could distinguish sera from patients with leprosy from those with tuberculosis and from sera of healthy volunteers. Our findings suggest that these synthetic peptides, with some refinement, may be useful as serological diagnostic antigens for leprosy. In addition, it was displayed that immunoinformatics provides reliable information for mapping potential B-cell epitopes for development of peptide-based diagnostic assays for neglected diseases.

Mycobacterium indicus pranii vaccine immunoprophylaxis in anti-phenolic glycolipid-1-positive leprosy contacts - A pilot study from a tertiary care center in North India.

BACKGROUND: Contacts of leprosy patients have an increased risk of infection with Mycobacterium leprae. Contact tracing and chemo- or immunoprophylaxis are important means of preventing leprosy transmission. AIMS: We aimed to evaluate the efficacy of immunoprophylaxis with Mycobacterium indicus pranii vaccine in reducing anti-phenolic glycolipid-1 titers in household contacts of leprosy patients. METHODS: This prospective single-center study was conducted in a tertiary care center in North India from January 2015 to December 2016. Contacts of leprosy patients (both paucibacillary and multibacillary) were screened for anti-phenolic glycolipid-1 antibodies with enzyme-linked immunosorbent assay. Those found positive were given immunoprophylaxis with a single dose of Mycobacterium indicus pranii vaccine, and anti-phenolic glycolipid-1 titers were evaluated at six and 12 months. All contacts were clinically followed for three years. RESULTS: Of the 135 contacts of 98 leprosy patients that were screened, 128 were recruited. Seventeen of these contacts were positive for anti-phenolic glycolipid-1 antibodies and were given Mycobacterium indicus pranii vaccine. Two contacts were lost to follow-up. After immunoprophylaxis, anti-phenolic glycolipid-1 titers were negative in all patients at all intervals, and no contact developed any clinical signs or symptoms of leprosy during the three-year follow-up. LIMITATIONS: The small number of contacts studied, the short follow-up period and the absence of a control group were limitations of this study. Dicussion: We could not find any papers on natural decline of PGL 1 titres in contacts, although in leprosy patients, these titres may even increase after completion of treatment. However the titres do correlate with bacterial load (reference: Int J Lepr Other Mycobact Dis. 1998 Sep;66(3):356-64) so if the tires decrease or become negative it may be considered as an indirect evidence of bacillary clearance. Hence we may suggest the protective efficacy. Furthermore, as the editor mentioned, considering the small number of positive patients, a control group was not possible in the present pilot study, but such studies may be carried out in the future. CONCLUSION: Immunoprophylaxis with Mycobacterium indicus pranii vaccine is effective and safe in preventing disease in contacts of leprosy patients. However, these findings need to be replicated in larger studies.

Latent leprosy infection identified by dual RLEP and anti-PGL-I positivity: Implications for new control strategies.

The number of new cases of leprosy reported worldwide has remained essentially unchanged for the last decade despite continued global use of free multidrug therapy (MDT) provided to any diagnosed leprosy patient. In order to more effectively interrupt the chain of transmission, new strategies will be required to detect those with latent disease who contribute to furthering transmission. To improve the ability to diagnose leprosy earlier in asymptomatic infected individuals, we examined the combined use of two well-known biomarkers of M. leprae infection, namely the presence of M. leprae DNA by PCR from earlobe slit skin smears (SSS) and positive antibody titers to the M. leprae-specific antigen, Phenolic Glycolipid I (anti-PGL-I) from leprosy patients and household contacts living in seven hyperendemic cities in the northern state of Pará, Brazilian Amazon. Combining both tests increased sensitivity, specificity and accuracy over either test alone. A total of 466 individuals were evaluated, including 87 newly diagnosed leprosy patients, 52 post-treated patients, 296 household contacts and 31 healthy endemic controls. The highest frequency of double positives (PGL-I+/RLEP+) were detected in the new case group (40/87, 46%) with lower numbers for treated (12/52, 23.1%), household contacts (46/296, 15.5%) and healthy endemic controls (0/31, 0%). The frequencies in these groups were reversed for double negatives (PGL-I-/RLEP-) for new cases (6/87, 6.9%), treated leprosy cases (15/52, 28.8%) and the highest in household contacts (108/296, 36.5%) and healthy endemic controls (24/31, 77.4%). The data strongly suggest that household contacts that are double positive have latent disease, are likely contributing to shedding and transmission of disease to their close contacts and are at the highest risk of progressing to clinical disease. Proposed strategies to reduce leprosy transmission in highly endemic areas may include chemoprophylactic treatment of this group of individuals to stop the spread of bacilli to eventually lower new case detection rates in these areas.

Oct-6 transcriptional factor a possible biomarker for leprosy diagnosis.

Leprosy is an infectious disease caused by Mycobacterium leprae that affects the skin and nerves. The nerve damage in leprosy may be related to alterations in transcriptional factors, such as Krox-20, Oct-6, Sox-10. Thirty skin biopsies in leprosy patients and 15 non-leprosy skin biopsies were evaluated using RT-qPCR to assess Krox-20, Oct-6, and Sox-10 and these data was related with S-100 immunohistochemistry. Changes in gene expression were observed in the skin and dermal nerves of leprosy patients in Oct-6 and Sox-10. When comparing Oct-6 with S-100 IHC as diagnostic tests for leprosy, Oct-6 showed a sensitivity of 73.3%, and specificity of 100%, while S-100 IHC showed a sensitivity of 96.6% and specificity of 100%. Our data suggest Oct-6 could be an auxiliary biomarker specific to detecting changes in dermal nerves in leprosy and thus useful to health workers and pathologists with no expertise to observe nerve injuries in leprosy.

Chemical Synthesis of the Trisaccharide Epitope of Phenolic Glycolipid-1 Surface Antigen from Mycobacterium leprae.

PGL-1 epitope 1 bearing a p-aminoethylphenol group was efficiently synthesized by using linear synthetic routes. A method for efficient synthesis of oligosaccharides containing rhamnose rings was developed. The chemistry is flexible and could be used for the synthesis of other PGLs antigens. A biotinylated PGL-1 antigen 23 was synthesized and could be used as a probe for early detection of leprosy.

Leprosy surveillance study in a highly endemic Brazilian area using leprosy specific serologic tests and IFNγ whole blood assay.

This surveillance study evaluated leprosy-serologic tests and the IFNγ whole-blood-assay/WBA as adjunct diagnostic tools. Previously diagnosed leprosy index cases, intradomiciliary, peridomiciliary contacts from a Brazilian endemic area were enrolled during domiciliary visits. Physical evaluation was performed by trained nurses and leprosy diagnosis confirmed by expert dermatologist. ELISA detected IgM anti-PGL-I, IgG anti-LID-1, and IgM/IgG anti-ND-O-LID antibodies. Heparinized WBA plasma stimulated with LID-1, 46f + LID-1, ML0276 + LID-1 (24 h, 37 °C, 5% CO2) was tested for human IFNγ (QuantiFERON®-TB Gold/QFT-G; Qiagen). The survey included 1731 participants: 44 leprosy index cases, 64 intradomiciliary, 1623 peridomiciliary contacts. Women represented 57.7%, median age was 32 years, 72.2% had BCG scar. Leprosy prevalence was higher in intradomiciliary (8.57%) versus peridomiciliary contacts (0.67%), p < 0.001. Among 23 suspects, five leprosy cases were confirmed: 4 multibacillary/MB and 1 paucibacillary/PB. Leprosy incidence was 0.30%: 1.56% in intradomiciliary versus 0.25% in peridomiciliary (p = 0.028). Seropositivity rates were 1.9% to PGL-I, 4.9% to LID-1, and 1.0% to ND-O-LID. LID-1 positivity was higher in all groups; incident cases were LID-1 seropositive. ND-O-LID positivity was higher in intra- versus peridomiciliary contacts (p = 0.022). IFNγ WBA (40 index cases, 19 suspects, 35 intradomiciliary, 74 peridomiciliary contacts) showed higher LID-1/WBA positivity in peridomiciliary contacts (p > 0.05); significant differences among groups were seen with 46f + LID-1 but 0276 + LID-1 induced higher IFNγ levels. Incident cases were LID-1 seropositive, while IFNγ-WBA had marginal diagnostic application. As seropositivity indicates exposed individuals at higher risk of disease development, the utility of serologic screening for surveillance and prophylactic measures remains to be demonstrated.

Diagnostic accuracy of tests using recombinant protein antigens of Mycobacterium leprae for leprosy: A systematic review.

The aim of this systematic review was to investigate the studies that evaluated the sensitivity and specificity of serologic tests using recombinant protein antigens from Mycobacterium leprae for leprosy diagnosis. We included 13 studies that were available in PubMed, Brazilian Virtual Library of Health, Web of Science, ScienceDirect and Scopus. From these studies, we found that the recombinant serine-rich 45-kDa protein of M. leprae (ML0411) demonstrated high performance for multibacillary (MB) also to paucibacillary (PB) patients, although this study was tested only for Indian population. Despite that, studies using the ND-O-LID antigen have been able to more accurately identify new cases of leprosy among people living in endemic or non-endemic areas and household contacts in Brazil, Colombia, and the Philippines, especially when combined with other biomarkers. Finally, low sensitivity values for PB patients' antibodies response remain challenging for tests intended to diagnose clinical forms that comprise this classification in leprosy.

[The use of synthetic glycoconjugates as components of the immunochromatographic test for rapid serological diagnosis of leprosy.]

The glycoconjugates with BSA (bovine serum albumin) were synthesized using a next saccharide: disaccharide derivative M.leprae PGL-1 (phenolic glycolipid-1); a complex of the disaccharide fragment and the branched hexasaccharide fragment LAM (lipoarabinomannan); diarabinofuranose fragment LAM. These glycoconjugates were used as antigenic components for leprosy rapid serotest construction in immunochromatographic format (leprosy LF serotest). The data obtained with sera of leprosy patients, patients who have been in contact with leprosy, and healthy donors indicate that the most promising antigenic component is a BSA conjugate with two synthetic epitopes - a disaccharide derivative of PGL-1 and a branched hexasaccharide fragment of LAM. The leprosy LF serotest with such glycoconjugate demonstrated the greatest diagnostic sensitivity for main forms of leprosy - paucibacillary (PB) and multibacillary (MB).

Serological evidence of Toxoplasma gondii infection as potential risk for the development of lepromatous leprosy in an endemic area for both neglected tropical diseases in Brazil.

BACKGROUND: Mycobacterium leprae and Toxoplasma gondii infections are both neglected tropical diseases highly prevalent in Brazil. Infection with certain parasite species can significantly alter susceptibility to other important pathogens, and/or influence the development of pathology. Here we investigated the possible influence of M. leprae/T. gondii co-parasitism on the manifestation of leprosy and its clinical forms. METHODS: Participants (n = 291) were recruited in Campos dos Goytacazes city, Rio de Janeiro state, southeast Brazil, from August 2015 to December 2019 and clinically diagnosed for leprosy. Participants were selected based on the presence (patients) or absence (healthy controls) of the leprosy disease. Contacts of patients were also recruited for this study. Serum samples from patients (n = 199) with leprosy, contacts (n = 40) and healthy controls (n = 52) were investigated for levels of IgM and IgG anti-phenolic glycolipid-1 (PGL-1) by ELISA. Additionally, IgG antibody against soluble Toxoplasma antigen (STAg) was measured in sera samples from leprosy patients, contacts and healthy controls for Toxoplasma gondii serology by ELISA. Anti-PGL-1 IgG and IgM levels were compared using one-way ANOVA Kruskal-Wallis or Mann-Whitney, while Spearman test was used to correlate levels of IgG anti-STAg and IgM/IgG anti-PGL-1 from seropositive and seronegative individuals for T. gondii infection. The risk of T. gondii infection for leprosy disease was assessed using Fisher's test. RESULTS: Levels of IgM anti-PGL-1 antibodies were significantly higher in multibacillary (MB) patients compared to paucibacillary (PB) patients (P = 0.0068). Higher IgM and IgG levels anti-PGL-1 were detected in patients with the lepromatous forms. The serologic prevalence for T. gondii infection was 74.9%. We detected increased anti-STAg antibody levels in leprosy patients (79.4%), reaching 88.8% within those with lepromatous form of this disease. The leprosy risk increase in T. gondii seropositive individuals was two-fold (odds ratio [OR] = 2.055; 95% confidence intervals [95% CI]: 1.18-3.51) higher than those seronegative, and considering the lepromatous leprosy risk this increase was even dramatic (OR = 4.33; 95% CI: 1.76-9.69) in T. gondii seropositive individuals. Moreover the leprosy risk in T. gondii seropositive individuals was weakly correlated to the levels of IgG anti-STAg and IgM/IgG anti-PGL-1. CONCLUSIONS: Altogether, our results suggest that T. gondii infection may exert immunomodulatory properties that influence to the susceptibility of leprosy, mainly on its more severe clinical form. A better understanding of parasite immunomodulation can ultimately contribute to the development of medical applications.