Synergistic antigen combinations for the development of interferon gamma release assays for paucibacillary leprosy

Abstract

The development of immunodiagnostic tests for paucibacillary leprosy (PB) is based on Mycobacterium leprae specific-cell mediated immunity (CMI)/IFN-γ production. Recently, novel M. leprae protein antigens that stimulate CMI have been described. This study evaluated different M. leprae antigen combinations in whole blood assay (WBA). Five study groups were tested (20 per group): newly diagnosed, untreated PB patients and multibacillary leprosy patients (MB); household contacts of MB patients (HHC); healthy endemic controls (EC); pulmonary tuberculosis patients (TB). WBA (heparinized, 24 h 37 °C 5% CO₂) were stimulated with: 10 μg/ml of each individual M. leprae recombinant protein (rML) and five combinations of rML (46f + LID-1, ML0276 + LID-1, ML2055 + ML1632 + ML2044, ML0276 + 46f, ML2055 + LID-1)-M. leprae cell sonicate (MLCS, 10 μg/ml), PHA (1 μg/ml), and PBS alone. Human IFN-γ ELISA (QuantiFERON-TB Gold/QFT-G, Cellestis) was performed using stimulated plasma (arbitrary cut-off = 50 pg/ml). Three out of five antigen combinations (46f + LID-1, ML0276 + LID-1, ML2055 + ML1632 + ML2044) were able to increase the levels of IFN-γ production in WBA in a larger number of responders among both PB leprosy and contacts. However, the magnitude of IFN-γ responses was higher among contacts. The antigen combination (46f + ML0276) stimulated IFN-γ only in symptomatic PB leprosy patients and not in asymptomatic contacts. Few controls (EC, TB) responded to combinations (0-15%), indicating the specificity of the response in an endemic area with high BCG coverage. The synergistic effect of new combinations of M. leprae proteins upon IFN-γ production in WBA indicates their potential use for the development of an interferon gamma release assay/IGRA for the diagnosis of PB leprosy.

Evaluation of Novel Tools to Facilitate the Detection and Characterization of Leprosy Patients in China

“…Our own work has identified multiple antigens that elicit IFN γ upon incubation with blood from PB patients in Brazil [22–25]. The data presented here extend these activities to the Chinese leprosy population but, consistent with recent data, indicate that additional antigens or markers are needed to enhance the sensitivity of the response [26]. Our data indicate that TB patients and EC did not respond to M. leprae antigen stimulation whereas a large proportion of PB patients, and even some MB patients, did.…”

Diagnosis of mycobacterial skin infections

“…The resulting infiltration is evaluated after 48–72 hours. Due to low sensitivity (33–96 %) and specificity (62 %) and false‐positive results in patients vaccinated with BCG, TST is obsolete for the diagnosis of tuberculosis in adults in developed countries . However, in children as well as in developing countries, the TST is still a cornerstone to test for infection with M. tuberculosis .…”

Diagnose mykobakterieller Hautinfektionen

“…Aktuell werden neue Proteinantigene von M. leprae untersucht, um einen zuverlässigen IGRA für Lepra zu entwickeln , der insbesondere für die Diagnose einer paucibazillären Lepra wertvoll werden könnte.…”

“…Interferon-γ-Release-Assays (IGRA) werden zunehmend zum Screening auf Infektionen mit M. tuberculosis eingesetzt und empfohlen [18,19] Aktuell werden neue Proteinantigene von M. leprae untersucht, um einen zuverlässigen IGRA für Lepra zu entwickeln [20][21][22], der insbesondere für die Diagnose einer paucibazillären Lepra wertvoll werden könnte.…”