The leishmanin skin test (LST), used for almost a century to detect infection and immunity to Leishmania,   is no longer available due to a lack of antigen for  injection. Leishmaniasis is a major neglected tropical disease infecting millions each year throughout Low and Middle Income Countries (LMICs) and more recently in developed countries. Depending on the species of Leishmania parasite,  the severity of the disease can range from a self-healing cutaneous lesions to deadly visceral infection. In some regions including the Indian subcontinent,  improved control programs are emerging along  with better treatments and new vaccines are  entering clinical trials.  In other parts of the world however, leishmaniasis, and in particular visceral leishmaniasis cases are increasing such as  in East Africa and South America. To support recent advances against leishmaniasis, it has become necessary to re-introduce the LST for surveillance in regions with outbreaks and to use as a biomarker for vaccine efficacy. The research described in this paper provides the foundation and rational for the return of  the LST (1).

 The LST, also known as the Montenegro test after its inventor in 1926 (2), is analogous to the tuberculin skin test (TST) that has been widely used as a diagnostic test for tuberculosis. Both the LST and TST are based on the same principle that intradermal injection of antigens into the forearm of an immune individual causes a memory T-cell mediated delayed-type hypersensitivity (DTH) reaction that is visible as swelling at the site of injection after 48hrs. This is particularly relevant for leishmaniasis because the T-cell mediated cellular immunity last for a lifetime following cure compared to the antibody response which only last for months. Moreover, antibodies are not protective against Leishmania infection whereas memory Th1 cellular immunity is protective against infection. This is evident as visceral leishmaniasis cases that are cured by drug treatment and cutaneous Leishmania cases that are self-cured are typically immune to re-infection for life resulting from a strong T-cell immunity.  In fact, this is the basis for the development of a live attenuated Leishmania vaccine that is poised to enter human clinical trials (3, 4).

 One of the difficulties associated with surveillance of visceral leishmaniasis is that only about 10% of infected individuals develop disease and the development of symptoms  takes up to 6 months or more providing plenty of time for transmission to become well established. Consequently, monitoring symtomatic cases only is not effective for surveillance. In comparison, the LST is positive in asymptomatic cases and together with monitoring symptomatic cases can provide a more accurate readout of transmission and  improve surveillance. For example, the LST can be used to investigate the extent of transmission around an index  case.  Future studies can reveal whether the LST antigen can be used in an in vitro blood test  such as the interferon-gamma release assay (IGRA), also.  The IGRA does not require a 48 hr follow-up of patients to examine the skin reaction and can be performed on a small blood sample.

Another important consideration will be how to test the new  attenuated leishmania vaccine (3, 4) in a LMIC that could require tens of thousands of subjects and years of follow-up which is not feasible or affordable. This is where the LST described in the paper (1) will be essential. Using the LST as a primary endpoint, instead of determining case numbers, will dramatically reduce the numbers and the time to establish vaccine performance.  Using the LST as a measure for immunity is supported by this study (1) and  epidemiological studies in Bangladesh and Sudan showing that LST positive individuals are protected from disease.

 What is old is new in leishmaniasis and the return of the LST teaches us from the past how to move forward.

 Pictured:   Visceral leishmaniasis surveillance in an endemic village;  Bihar, India. Dr RN Pandey and Professor  Greg Matlashewski are observing a rapid diagnostic test strip for anti-leishmania antibodies in an endemic village. The leishmanin skin test  proposed in this study (1) will improve the detection of asymptomatic infections for surveillance   and will support the development of new vaccines for visceral leishmaniasis. 

References:

1. Dey R, Alshaweesh J, Singh KP, Lypaczewski P, Karmakar S, Klenow L, Paulini K, Kaviraj S, K1amhawi S, Valenzuela JG, Singh S, Hamano S, Satoskar AR, Gannavaram S, Nakhasi HL, Matlashewski G. Production of Leishmanin Skin Test antigen from Leishmania donovani for future reintroduction in the field.  Nature Commun. 2023 Nov 2;14(1):7028. doi: 10.1038/s41467-023-42732-2.

2.  Montenegro J. Cutaneous reaction in leishmaniasis. Arch Derm Syphilol 13:187-194, 1926

3. Zhang WW, Karmakar S, Gannavaram S, Dey R, Lypaczewski P, Ismail N, Siddiqui A, Simonyan V, Oliveira F, Coutinho-Abreu IV, DeSouza-Vieira T, Meneses C, Oristian J, Serafim TD, Musa A, Nakamura R, Saljoughian N, Volpedo G, Satoskar M, Satoskar S, Dagur PK, McCoy JP, Kamhawi S, Valenzuela JG, Hamano S, Satoskar AR, Matlashewski G, Nakhasi HL. A second generation leishmanization vaccine with a markerless attenuated Leishmania major strain using CRISPR gene editing. Nature Commun. 2020 Jul 10;11(1):3461. doi: 10.1038/s41467-020-17154-z. 

4. Karmakar S, Ismail N, Oliveira F, Oristian J, Zhang WW, Kaviraj S, Singh KP, Mondal A, Das S, Pandey K, Bhattacharya P, Volpedo G, Gannavaram S, Satoskar M, Satoskar S, Sastry RM, Oljuskin T, Sepahpour T, Meneses C, Hamano S, Das P, Matlashewski G, Singh S, Kamhawi S, Dey R, Valenzuela JG, Satoskar A, Nakhasi HL.Preclinical validation of a live attenuated dermotropic Leishmania vaccine against vector transmitted fatal visceral leishmaniasis. Commun Biol. 2021 Jul 30;4(1):929. doi: 10.1038/s42003-021-02446-x.