High performance of an enzyme linked immunosorbent assay for American tegumentary leishmaniasis diagnosis with Leishmania (Viannia) braziliensis amastigotes membrane crude antigens

Abstract

The diagnosis of American tegumentary leishmaniasis (ATL) still requires the design of more effective tools. Leishmania (Viannia) braziliensis is the causal agent of the 90% of Argentinean ATL cases. Considering the current knowledge, an ELISA based crude antigen (CA) for the diagnosis was designed. Ninety-nine subjects diagnosed as ATL, 27 as no-ATL, and 84 donors from non-ATL-endemic areas were included in this study. The current ATL diagnosis was based four techniques, dermal smear microscopic examination (parasitological test), PCR, Leishmanin skin test, and clinical records. We obtained CA extracts from promastigotes and amastigotes from macrophage cultures of different zymodemes of endemic Leishmania species circulating in the study area. Crude antigens from the 'local' main zymodeme of L. (V.) braziliensis showed the highest reactivity against anti-Leishmania antibodies compared to the other included species. The CA of amastigotes of this zymodeme was 3.4 fold more reactive than promastigotes one. Moreover, amastigote-membrane CA (MCA) were 3.6 fold more reactive than the soluble antigens. The MCA-ELISA reached a sensitivity and specificity of 98% (CI = 94.7%-100%) and 63.6% (53.9-73.1), respectively. When anti-Trypanosoma cruzi reactive sera were excluded, the specificity reached 98.4% (94.4-100), while the sensitivity was similar, with a positive predictive value (PV) of 98.6% (94.6-100) and negative PV of 96.3% (91.6-100). The performance of the MCA-ELISA results strongly contribute to the final diagnostic decision, since a non-reactive serological result almost discards the suspected ATL, because of its high negative PV. The developed PLOS ONE

The Trypanosoma cruzi Antigen and Epitope Atlas: antibody specificities in Chagas disease patients across the Americas

“…Serum samples were collected from clotted blood obtained by venipuncture and analysed for T. cruzi -specific antibodies with the following commercially or in-house kits: AR: Chagatest ELISA lysate (Laboratorios Wiener, Argentina), Chagatest HAI (Laboratorios Wiener, Argentina), Chagatest ELISA recombinant v4.0 (Wiener Lab, Argentina); BO: recombinant and lysate ELISA; BR: conventional in-house ELISA (confirmed by TESA-Blot 60 ); CO: ELISA (Chagatek, Organon, Argentina) and HAI (Chagatest, Weiner, Chile); MX: Bio-Rad Chagascreen Plus v4 (recombinant), Test ELISA para Chagas III (Grupo Bios, Chile), Accutrak Chagas Microelisa Test System (Laboratorio Lemos, Argentina), Accutrack Chagatek ELISA recombinante (Laboratorio Lemos, Argentina); US: Chagas Stat-Pak (Chembio, Medford, NY), Hemagen Chagas’ Kit ELISA (Hemagen Diagnostics Inc., Columbia, MD), and an in-house TESA-Blot 60 . ATL samples were classified using an in-house ELISA based on crude antigen extracts from promastigotes and amastigotes of three different endemic Leishmania species and two reference strains 61 . Two additional panels of samples were used for the validation of antigens and assessment of diagnostic potential.…”

Cutaneous Leishmaniasis: A 2022 Updated Narrative Review into Diagnosis and Management Developments

“…Yet, there are still attempts made to improve the sero-diagnosis of CL. A relative successful approach seems to be the use of crude antigens derived from amastigotes of local Leishmania strains [ 26 ]. In contrast, a systematic review by Zanetti et al revealed that more specific (recombinant) antigens yielded better diagnosis results, that is, sensitivity (93.8–100%) and specificity (82.5–100%) for ATL, compared with crude soluble antigen-based ELISAs [ 27 ].…”

Simple and promising paper-based electrochemical platform for serological detection of American tegumentary leishmaniasis

“…4 Of the approximately 30 species that are part of the Leishmania genus, 5 Leishmania (Viannia) braziliensis has been implicated in 90% of ATL cases in our country. 6 , 7 , 8 Conventional treatment for this disease is relatively toxic, slow, invasive, and expensive, and although this clinical form is not fatal, it causes considerable morbidity in patients; 1 therefore, a rapid diagnosis is crucial for the patient.…”

“…However, this method has some intrinsic limitations: amastigote detection in smears is invasive and has a low sensitivity, particularly in ATL, which requires trained personnel and is time consuming. 6 , 7 , 8 , 9 , 10 , 11 On the other hand, molecular techniques based on polymerase chain reaction (PCR) are not routinely used because of their complexity and difficulty in being implemented in clinical practice, despite presenting good diagnostic performance (high sensitivity and specificity).…”

“…Bracamonte et al 6 performed for the first time a method to diagnose ATL by means of ELISA with a membrane of crude antigens (MCA-ELISA), with very good results. Crude antigenic extracts (CA) were obtained from promastigotes and amastigotes of macrophage cultures of L. (V.) braziliensis , the causative agent of ATL in our region.…”