Leprosy Household Contacts Research Articles

Levels of TGF- Serum Positively Correlated with Levels of IgM Anti PGL-1 In Household Contacts of Multibacillary Leprosy Patients

Background Leprosy is a chronic progressive infectious disease. Phenolic glycolipid-1 (PGL-1) is an antigen of Mycobacterium leprae, which can trigger the host’s antibody response. Transforming growth factorβ (TGF-β) plays an immunosuppressive role when the host is exposed to PGL-1 antigen or other M. leprae antigens. This study aims to determine the correlation of TGF-β levels with IgM anti-PGL-1 levels in patients with multibacillary leprosy household contacts. Methods Observational analytical study with a cross-sectional approach. The study subjects consisted of 48 household contact subjects and 24 non-household contact subjects aged 15-65 years old who were selected by consecutive sampling based on inclusion and exclusion criteria. 3 mL of venous blood samples were taken and then examined for IgM anti-PGL-1 and TGF-β with the ELISA kit. Data analysis was carried out using SPSS version 23, and a p-value <0.05 was significant. Results The mean level of IgM anti-PGL-1 in the household contact group was 685.46 ± 290.79 u/mL, while in the non-household contact group was 345.50 ± 206.58 u/mL. The mean TGF-β level in household contact groups was 256.69 ± 127.41 pg/mL, while in the non-household contact group was 144.85 ± 36.73 pg/mL (p<0.001). This study found a moderate positive relationship (r=0.450, p<0.001) between levels of TGF-β and IgM anti-PGL-1 household contacts and non-household contacts group. Conclusions The mean level of IgM anti-PGL-1 and TGF-β in household contacts is higher than in non-household contacts, with a significant difference. There is a moderate positive significant relationship between levels of TGF-β and IgM anti-PGL-1 household contacts and non-household contacts group.

High-resolution ultrasonography for early diagnosis of neural impairment in seropositive leprosy household contacts.

Leprosy household contacts (HC) represent a high-risk group for the development of the disease. Anti-PGL-I IgM seropositivity also increases the risk of illness. Despite significant advances in leprosy control, it remains a public health problem; and early diagnosis of this peripheral neuropathy represents one of the main goals of leprosy programs. The present study was performed to identify neural impairment in leprosy HC by analyzing differences in high-resolution ultrasonographic (US) measurements of peripheral nerves between leprosy HC and healthy volunteers (HV). Seventy-nine seropositive household contacts (SPHC) and 30 seronegative household contacts (SNHC) underwent dermato-neurological examination and molecular analysis, followed by high-resolution US evaluation of cross-sectional areas (CSAs) of the median, ulnar, common fibular and tibial nerves. In addition, 53 HV underwent similar US measurements. The US evaluation detected neural thickening in 26.5% (13/49) of the SPHC and only in 3.3% (1/30) among the SNHC (p = 0.0038). The CSA values of the common fibular and tibial nerves were significantly higher in SPHC. This group also had significantly greater asymmetry in the common fibular and tibial nerves (proximal to the tunnel). SPHC presented a 10.5-fold higher chance of neural impairment (p = 0.0311). On the contrary, the presence of at least one scar from the BCG vaccine conferred 5.2-fold greater protection against neural involvement detected by US (p = 0.0184). Our findings demonstrated a higher prevalence of neural thickening in SPHC and support the role of high-resolution US in the early diagnosis of leprosy neuropathy. The combination of positive anti-PGL-I serology and absence of a BCG scar can identify individuals with greater chances of developing leprosy neuropathy, who should be referred for US examination, reinforcing the importance of including serological and imaging methods in the epidemiological surveillance of leprosy HC.

Early diagnosis of neural impairment in seropositive leprosy household contacts: The experience of a reference center in Brazil.

Leprosy is an infectious disease that remains with a high number of new cases in developing countries. Household contacts have a higher risk for the development of the disease, but the neural impairment in this group is not well elucidated yet. Here, we measured the chance of occurrence of peripheral neural impairment in asymptomatic leprosy household. Contacts who present anti-PGL-I IgM seropositivity, through electroneuromyography (ENMG) evaluation. We recruited 361 seropositive contacts (SPC) from 2017 to 2021, who were subjected to an extensive protocol that included clinical, molecular, and electroneuromyographic evaluations. Our data revealed a positivity of slit skin smear and skin biopsy qPCR of 35.5% (128/361) and 25.8% (93/361) respectively. The electroneuromyographic evaluation of the SPC showed neural impairment in 23.5% (85/361), with the predominance of a mononeuropathy pattern in 62.3% (53/85). Clinical neural thickening was observed in 17.5% (63/361) of seropositive contacts, but among the individuals with abnormal ENMG, only 25.9% (22/85) presented neural thickening in the clinical exam. Ours results corroborates the need to make the approach to asymptomatic contacts in endemic countries more timely. Since leprosy in its early stages can present an indolent and subclinical evolution, serological, molecular, and neurophysiological tools are essential to break the disease transmission chain.

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.

RLEP LAMP for the laboratory confirmation of leprosy: towards a point-of-care test

BackgroundNucleic acid-based amplification tests (NAAT), above all (q)PCR, have been applied for the detection of Mycobacterium leprae in leprosy cases and household contacts with subclinical infection. However, their application in the field poses a range of technical challenges. Loop-mediated isothermal amplification (LAMP), as a promising point-of-care NAAT does not require sophisticated laboratory equipment, is easy to perform, and is applicable for decentralized diagnosis at the primary health care level. Among a range of gene targets, the M. leprae specific repetitive element RLEP is regarded as highly sensitive and specific for diagnostic applications. MethodsOur group developed and validated a dry-reagent-based (DRB) RLEP LAMP, provided product specifications for customization of a ready-to-use kit (intended for commercial production) and compared it against the in-house prototype. The assays were optimized for application on a Genie® III portable fluorometer. For technical validation, 40 “must not detect RLEP” samples derived from RLEP qPCR negative exposed and non-exposed individuals, as well as from patients with other conditions and a set of closely related mycobacterial cultures, were tested together with 25 “must detect RLEP” samples derived from qPCR confirmed leprosy patients. For clinical validation, 150 RLEP qPCR tested samples were analyzed, consisting of the following categories: high-positive samples of multibacillary (MB) leprosy patients (> 10.000 bacilli/extract), medium-positive samples of MB leprosy patients (1.001–10.000 bacilli/extract), low-positive samples of MB leprosy patients (1–1.000 bacilli/extract), endemic controls and healthy non-exposed controls; each n = 30. ResultsTechnical validation: both LAMP formats had a limit of detection of 1.000 RLEP copies, i.e. 43–27 bacilli, a sensitivity of 92% (in-house protocol)/100% (ready-to-use protocol) and a specificity of 100%. Reagents were stable for at least 1 year at 22 °C. Clinical validation: Both formats showed a negativity rate of 100% and a positivity rate of 100% for high-positive samples and 93–100% for medium positive samples, together with a positive predictive value of 100% and semi-quantitative results. The positivity rate for low-positive samples was 77% (in-house protocol)/43% (ready-to-use protocol) and differed significantly between both formats. ConclusionsThe ready-to-use RLEP DRB LAMP assay constitutes an ASSURED test ready for field-based evaluation trials aiming for routine diagnosis of leprosy at the primary health care level.

Predictive nomogram for leprosy using genetic and epidemiological risk factors in Southwestern China: Case-control and prospective analyses.

BackgroundThere is a high incidence of leprosy among house-contacts compared with the general population. We aimed to establish a predictive model using these genetic factors along with epidemiological factors to predict leprosy risk of leprosy household contacts (HHCs).MethodsWeighted genetic risk score (wGRS) encompassing genome wide association studies (GWAS) variants and five non-genetic factors were examined in a case–control design associated with leprosy risk including 589 cases and 647 controls from leprosy HHCs. We constructed a risk prediction nomogram and evaluated its performance by concordance index (C-index) and calibration curve. The results were validated using bootstrap resampling with 1000 resamples and a prospective design including 1100 HHCs of leprosy patients.FindingThe C-index for the risk model was 0·792 (95% confidence interval [CI] 0·768-0·817), and was confirmed to be 0·780 through bootstrapping validation. The calibration curve for the probability of leprosy showed good agreement between the prediction of the nomogram and actual observation. HHCs were then divided into the low-risk group (nomogram score ≤ 81) and the high-risk group (nomogram score > 81). In prospective analysis, 12 of 1100 participants had leprosy during 63 months’ follow-up. We generated the nomogram for leprosy in the validation cohort (C-index 0·773 [95%CI 0·658-0·888], sensitivity75·0%, specificity 66·8%). Interpretation The nomogram achieved an effective prediction of leprosy in HHCs. Using the model, the risk of an individual contact developing leprosy can be determined, which can lead to a rational preventive choice for tracing higher-risk leprosy contacts.FundingThe ministry of health of China, ministry of science and technology of China, Chinese academy of medical sciences, Jiangsu provincial department of science and technology, Nanjing municipal science and technology bureau.

Single-Nucleotide Polymorphisms in Genes Predisposing to Leprosy in Leprosy Household Contacts in Zhejiang Province, China.

PurposeGenome-wide association studies (GWAS) have identified multiple genetic variants associated with leprosy. To investigate the single and combined associations between single-nucleotide polymorphisms (SNPs) and the development of leprosy, we therefore performed generalized multi-analytical (GMDR) analysis in Chinese leprosy household contacts and constructed a risk prediction model.Patients and MethodsThis case–control study included 229 leprosy cases and 233 healthy household contacts in Zhejiang province, China. Participants were genotyped for 17 polymorphisms selected from GWAS. The Pearson χ2 test, logistic regression and GMDR analysis were performed to investigate gene–gene interactions and construct a risk prediction model for leprosy.ResultsThe genotype and the allele distributions of rs142179458, rs2275606, rs663743 and rs73058713 were significantly different between patients and controls. rs2275606, rs6478108, rs663743 and rs73058713 showed an association after adjusting for sex and age in the logistic regression. A five-way interaction model consisting of rs2058660, rs2275606, rs4720118, rs6478108 and rs780668 was chosen as the optimal model for determining leprosy susceptibility. The model classified 237 (51.3%) into the low-risk group and 225 (48.7%) individuals into the high-risk group. The area under the curve (AUC) of this model was 0.757 (95% CI: 0.712–0.803), and the odds ratio for leprosy between the high- and low-risk groups was 9.733 (95% CI: 6.384–14.960; P<0.001). The sensitivity and specificity of the model were observed to be 74.7% and 76.8%, respectively.ConclusionOur results suggest that rs2058660, rs2275606, rs4720118, rs6478108 and rs780668, five SNPs with a significant sole effect on leprosy, interact to confer a higher risk for the disease in leprosy household contacts (HHCs).

BCG vaccine and leprosy household contacts: Protective effect and probability to becoming sick during follow-up

BackgroundImmunoprophylaxis with Bacillus Calmette–Guérin (BCG) vaccine is still the most effective intervention in the prevention of leprosy among household contacts (HHCs) of leprosy patients. MethodsA retrospective cohort study using data of 5.061 HHCs for a period of 16 years (follow-up of 7 years per leprosy HHCs), evaluating the occurrence of disease as the main outcome and the presence or absence of BCG scars verified at the first evaluation. Statistical analyzes were performed using the relative risk, hazard ratio and survival curves by Kaplan-Meier test. ResultsA total of 92 contacts sickened, of which 41.3% (38/92) in the first year and 58.7% (54/92) in the course of the other years of follow-up. Of those who became sick, 62% (57/92) developed borderline tuberculoid (BT). The additional protective effect occurred for those who had 2 BCG scars at the first follow-up assessment (Relative Risk: 0.41; p = 0.007) when compared to those not previously exposed to the vaccine. The number of BCG scars examined at the first assessment (t0 = time zero) affected the occurrence of the outcome evidenced by the difference in survival curves throughout the follow-up (Log Rank, p = 0.041; Breslow, p = 0.012; Tarone-Ware, p = 0.020). Leprosy HHCs with 0 BCG scar at time zero (t0) have a shorter survival time (average time of 22 months between t0 and outcome) when compared to those with 2 BCG scars (average time of 36 months between t0 and outcome). ConclusionsVaccination of healthy individuals without signs and symptoms of leprosy is extremely important because BCG vaccine has an additional protective effect in those cases with 2 BCG scars throughout follow-up. Reducing the risk of leprosy HHCs becoming sick depends on preventive actions such as immunoprophylaxis and index cases treatment.

Molecular, immunological and neurophysiological evaluations for early diagnosis of neural impairment in seropositive leprosy household contacts.

BackgroundHousehold contacts constitute the highest risk group for leprosy development, and despite significant progress in the disease control, early diagnosis remains the primary goals for leprosy management programs.MethodsWe have recruited 175 seropositive and 35 seronegative household contacts from 2014 to 2016, who were subjected to an extensive protocol that included clinical, molecular (peripheral blood qPCR, slit-skin smear qPCR, skin biopsy qPCR) and electroneuromyographic evaluations.Results/Principal findingsThe positivity of peripheral blood qPCR of seropositive contacts was 40.6% (71/175) whereas only 8.6% (3/35) were qPCR positive in seronegative contacts (p = 0.0003). For the slit-skin smear, only 4% (7/175) of seropositive contacts presented positive bacilloscopy, whereas the qPCR detected 47.4% (83/175) positivity in this group compared with only 17.1% (6/35) in seronegative contacts (p = 0.0009). In the ENMG evaluation of contacts, 31.4% (55/175) of seropositives presented some neural impairment, and 13.3% (4/35) in seronegatives (p = 0.0163). The presence of neural thickening conferred a 2.94-fold higher chance of ENMG abnormality (p = 0.0031). Seropositive contacts presented a 4.04-fold higher chance of neural impairment (p = 0.0206). The peripheral blood qPCR positivity presented odds 2.08-fold higher towards neural impairment (OR, 2.08; p = 0.028). Contrarily, the presence of at least one BCG vaccine scar demonstrated 2.44-fold greater protection against neural impairment (OR = 0.41; p = 0.044).Conclusions/SignificanceELISA anti-PGL-I is the most important test in determining the increased chance of neural impairment in asymptomatic leprosy household contacts. The combination of the two assays (ELISA anti-PGL-I and peripheral blood qPCR) and the presence of BCG scar may identify individuals with higher chances of developing leprosy neuropathy, corroborating with the early diagnosis and treatment.

Anti-PGL1 salivary IgA/IgM, serum IgG/IgM, and nasal Mycobacterium leprae DNA in individuals with household contact with leprosy

Leprosy household contacts represent a group at high risk of developing the disease. The aim of this study was to detect Mycobacterium leprae subclinical infection in this group through serological and molecular parameters. Serum anti-PGL1 IgG/IgM and salivary anti-PGL1 IgA/IgM was investigated using an ELISA, and nasal carriage of M. leprae DNA was detected by PCR, in leprosy household contacts of paucibacillary (PB) and multibacillary (MB) household leprosy patients (n=135), their index cases (n=30), and in persons living in a low endemic city (n=17). Salivary anti-PGL1 IgA and IgM and serum anti-PGL1 IgG showed good correlation comparing contacts and index cases (p<0.01, p<0.005, and p<0.0001, respectively). This was not observed for serum anti-PGL1 IgM (p>0.05). A high frequency of anti-PGL1 IgM positivity was found in IgG-negative samples (p<0.0001). For IgG-positive samples, IgM antibodies were also positive in most of the samples. None of the 17 volunteers living in a low endemic city presented seropositivity for IgG; however, two of them showed positivity for anti-PGL1 IgM. M. leprae DNA was found in the nasal swabs of nine out of the 85 MB household leprosy contacts (10.6%) and in three out of the 50 PB household leprosy contacts (6.0%). We strongly suggest that serum IgG/IgM and salivary anti-PGL1 IgA/IgM measurements are used to follow leprosy household contacts.

Development of a new point-of-care test for leprosy

Development of a new point-of-care test for leprosy

Clinical and immunological evaluation after BCG-id vaccine in leprosy patients in a 5-year follow-up study.

IntroductionThe use of bacillus Calmette–Guérin (BCG) has long been considered a stimulus for immune reactivity in leprosy household contacts. Probably, the combination of multidrug therapy with BCG could facilitate the clearance of leprosy bacilli in the host, reduce relapse rates, and shorten the duration of skin-smear positivity.MethodsTo investigate the mechanism of action of BCG, a study involving 19 leprosy patients, eleven multibacillary (MB) and eight paucibacillary, was performed to assess the in vitro production of interleukin (IL)-10, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-6, and IL-17 in the supernatant of peripheral blood mononuclear cells, before and 30 days after inoculation with BCG intradermally (BCG-id). Peripheral blood mononuclear cells isolated by Ficoll–Hypaque gradient were cultivated with Concanavalin-A (Con-A), lipopolysccharides (LPS), or BCG. The supernatant was collected for ELISA quantification of cytokines. The immunohistochemistry of IFN-γ, IL-1, IL-10, IL-12, transforming growth factor (TGF)-β, and TNF-α was carried out in biopsies of skin lesions of leprosy patients before and 30 days after inoculation of BCG-id. These patients were followed up for 5 years to assess the therapeutic response to multidrug therapy, the occurrence of leprosy reactions, and the results of bacterial index and anti-PGL-1 serology after the end of treatment.ResultsThe results showed increased production of cytokines after BCG-id administration in MB and paucibacillary leprosy patients. There was statistically higher levels of TNF-α (P = 0.017) in MB patients and of IL-17 (P = 0.008) and IFN-γ (P = 0.037) in paucibacillary patients. Immunohistochemical staining, especially for TNF-α, was more intense in biopsies of MB leprosy patients taken after BCG-id administration, probably for induction of innate human immunity. The clinical evaluation suggests that BCG-id is able to induce a more effective therapeutic response, with reduction of the number and the intensity of leprosy reactions.ConclusionThese results suggest that BCG-id induces activation of the initial phase of immunocellular activity: innate human immunity (increase in TNF-α, IL-12 and macrophage activation). Therefore, we conclude that the use of BCG-id could be indicated as an adjuvant to multidrug therapy in treatment of leprosy patients.

Cytokine responses to Mycobacterium leprae unique proteins differentiate between Mycobacterium leprae infected and naïve armadillos

New diagnostic tools for early detection of leprosy are necessary to help reduce its transmission and severity. M. leprae unique proteins have been used to assess differences in human T-cell responses in leprosy patients, household contacts and endemic controls. In this study, we examined the response of M. leprae-infected armadillos to a variety of M. leprae recombinant antigen candidates currently being examined for diagnostic efficacy in humans. Among recently M. leprae infected armadillos, IFN-gamma expression was enhanced after stimulation of PBMC with all M. leprae recombinant proteins except for ML2283 (mean: 2.65 Relative Quantification (RQ)). The group mean stimulation index for M. leprae proteins ML0009, ML1601, ML2478 and ML2531 averaged 35.2 RQ and was significantly higher (P < 0.05) than that measured among the non-infected, naive group (mean 6.2 RQ). Although ML0840 tended to enhance IFN-gamma levels, the mean IFN-gamma transcript levels of the currently experimentally inoculated group (20.1 RQ) was not significantly different statistically (P = 0.10) from the mean of the naive group (7.5 RQ). Also no statistically significant differences were observed in IFN-gamma transcript levels between the resistant and currently experimentally inoculated group (P > 0.05) or between the resistant and the naive group (P > 0.05) after stimulation of PBMCs with all M. leprae recombinant proteins. Only low levels of TNF-alpha were observed across all groups after in vitro stimulation with all the antigens examined. These data suggest that armadillos can be used effectively to help identify M. leprae specific proteins that may be applied for monitoring T-cell responses in M. leprae infected hosts as their disease progresses as well as for the early diagnosis of leprosy.

Relative lack of clinical disease among household contacts of tuberculosis patients compared to leprosy households

The incidence of clinical tuberculosis and clinical leprosy among household members of tuberculosis and leprosy patients in Sri Lanka was studied. The study period was approximately 20 years (January 1981 to December 2001) and the total number of patients and contacts were 325 and 968 for tuberculosis and 726 and 3066 for leprosy, respectively. While none of the tuberculosis patient households had more than 1 patient nor any contacts who developed clinical disease during the observation period, 20% (148/726) of the leprosy patients had more than 1 patient in the family and 0.9% (13/1403) of their contacts who were followed-up developed clinical leprosy during the observation period. Although the tuberculosis patient household contacts did not develop clinical disease, in 79% (88/112) of contacts who were tested by Western blot analysis, there was serologic evidence of Mycobacterium tuberculosis infection. These data show that in populations of comparable socio-economic, environmental and geographic locations, tuberculosis and leprosy show very different transmission patterns. In general, in tuberculosis household contacts, in spite of exposure, infection did not proceed to clinical disease. In contrast, a significant number of leprosy household contacts developed clinical leprosy. These findings have implications in the design and implementation of control programmes for these two diseases.