ML0405 and ML2331 Are Antigens ofMycobacterium lepraewith Potential for Diagnosis of Leprosy

Abstract

Despite the success of multidrug therapy in reducing the number of registered leprosy cases worldwide, evidence suggests that Mycobacterium leprae continues to be transmitted. A serological diagnostic test capable of identifying and allowing treatment of early-stage disease could reduce transmission and prevent the onset of the disability, a common complication of the disease in later stages. Serological diagnosis based on antibody recognition of phenolic glycolipid I (PGL-I) cannot reliably identify individuals with lower bacterial indices (BI). One strategy that might improve this situation is the provision of highly specific serological antigens that may be combined with PGL-I to improve the sensitivity of diagnosis. Using serological expression cloning with a serum pool of untreated lepromatous leprosy (LL) patients, we identified 14 strongly reactive M. leprae proteins, 5 of which were previously unstudied. We present results suggesting that two of these proteins, ML0405 and ML2331, demonstrate the ability to specifically identify LL/borderline lepromatous (BL) patients on the basis of immunoglobulin G (IgG) reactivity. In a household contact study, LL index cases were identified on the basis of this reactivity, while household contacts of these patients demonstrated undetectable reactivity. At a serum dilution of 1:800, suitable to reduce background PGL-I IgM reactivity, two BL patients with a BI of <4 showed anti-human polyvalent immunoglobulin G, A, and M reactivity measured with a combination of ML0405, ML2331, and natural disaccharide O-linked human serum albumin (NDOHSA) (synthetic PGL-I) that was markedly higher than IgM reactivity to NDOHSA alone. We suggest that ML0405 and ML2331 may have utility in serological leprosy diagnosis.Leprosy is a devastating human disease caused by infection with Mycobacterium leprae bacilli. The disease predominantly affects the skin, although during infection, significant nerve destruction leads to deformities of the hand, foot, face, and, in some cases, eye (1). The disease is represented by a clinical spectrum. Lepromatous leprosy/borderline lepromatous (LL/ BL) patients represent one pole of the spectrum, demonstrating a high bacterial index (BI) and, as such, are classified as multibacillary (MB). LL/BL patients demonstrate high titers of M. leprae-specific antibodies and an absence of M. leprae-specific cell-mediated immunity. At the opposite pole, tuberculoid tuberculoid/borderline tuberculoid (TT/BT) patients demonstrate very low or absent BI and are designated paucibacillary. These individuals demonstrate significant M. leprae-specific cellmediated immunity and very low or absent titers of M. lepraespecific antibodies.Despite the success of multidrug therapy in reducing the number of registered leprosy cases worldwide, the annual rate of new case detection remains unchanged, at approximately 700,000 cases per year (33), with children representing 15% of new cases (18). This suggests that active transmission of M. leprae is still occurring, but the route and me...

Identification of serological biomarkers of infection, disease progression and treatment efficacy for leprosy

“…The titres of IgG antibodies to protein antigens generally correlate with the bacterial load and strong responses in MB patients can rival anti-PGL-I responses. The best protein candidates to assess the IgG response include ML0405, ML2331, ML2055 (Reece et al 2006, Duthie et al 2007), ML2028 (Launois et al 1994) and ML2038 , Kai et al 2008. A bioinformatics analysis predicted ML0405 to be one of the most promiscuous M. leprae proteins studied to date and to contain T cell epitopes predicted to be recognised by 50 out of 51 human leukocyte antigen-DR alleles along with seven potential T cell epitopes and 39 potential B cell epitopes, whereas ML2331 contained 24 potential B cell epitopes (Sampaio et al 2011).…”

“…Cloning and purification of recombinant M. leprae proteins -DNA sequences encoding 12 full-length M. leprae proteins were cloned from genomic DNA as previously described (Spencer et al 2005, Reece et al 2006. Purified recombinant proteins were produced by the Infectious Disease Research Institute (IDRI), Seattle, WA or Colorado State University (CSU), Fort Collins, CO, USA.…”

Bioinformatic Identification of Tandem Repeat Antigens of the Leishmania donovani Complex

“…The expressed TR-containing proteins were analyzed for seroreactivity using panels of patient and control sera. L. infantum soluble lysate antigen (SLA) was used as a positive control and a Mycobacterium leprae antigen ML2331 was used as an irrelevant antigen (26). Proteins were diluted in an enzyme-linked immunosorbent assay (ELISA) coating buffer, and 96-well plates were coated with 1 g of L. infantum SLA or 200 ng of individual recombinant antigens, followed by blocking with phosphate-buffered saline containing 0.05% Tween 20 and 1% bovine serum albumin.…”

“…rLinJ11.0070r2 and rLinJ25.1100TR showed intermediate reactivity to the VL patient sera; none of the four antigens were recognized by sera from healthy donors. VL patient sera showed only a weak antibody response to an irrelevant Mycobacterium leprae antigen ML2331 (26). Compared to the reactivity of the irrelevant antigen, rLinJ11.0070r2, rLinJ25.1100TR, rLinJ27.0400r2, and rLinJ29.29.0110TR, as well as L. infantum SLA, showed significantly stronger reactivity to the VL patient sera, whereas rLinJ21.2010TR or LinJ32.3710TR did not detect VL-specific antibodies (P Ͻ 0.05 on rLinJ11.0070r2, P Ͻ 0.01 on rLinJ25.1100TR, and P Ͻ 0.001 on rLinJ27.0400r2, rLinJ29.29.0110TR, and L. infantum SLA by unpaired t test).…”

Use of Protein Microarrays To Define the Humoral Immune Response in Leprosy Patients and Identification of Disease-State-Specific Antigenic Profiles

“…Recombinant forms of some of the more significant native proteins were subsequently created and tested (22,27,37). Recently, several groups have also used a postgenomic approach to discover new antigens for leprosy diagnosis (1,2,28,36,37). These studies all exploited genomic sequence for the identification of M. leprae-specific proteins or peptides that may be suitable for serodiagnosis of different disease states of leprosy.…”