Towards a Buruli Ulcer Rapid Diagnostic Test That Targets Mycolactone

We report the development of a prototype rapid diagnostic test for Buruli ulcer, an ulcerative necrotizing skin disease caused by M. ulcerans . The test was designed to detect mycolactone, a metabolite unique to M. ulcerans . The chief technical challenge was to develop a simple workflow to extract trace amounts of mycolactone from wound exudates collected with a swab and, after a concentration step, to visualize the mycolactone by means of a lateral flow assay. This was achieved by utilizing a mouse monoclonal antibody specific for mycolactone and magnetic gold nanoshells. The latter are a novel class of reporter particles consisting of a ferrite core, a silica gel middle layer that serves to decrease the overall density of the nanoparticle and facilitate its resuspension in aqueous media, and an outer layer of gold, which provides a dark coloration through plasmon resonance effects. These nanoparticles, once conjugated to the anti-mycolactone antibody, enable the immunomagnetic concentration of the targeted analyte and its detection by lateral flow assay. The test procedure can be conducted within 2 hours with a magnetic rack and no powered instrumentation is required. The test can detect as little as 3.5–7 ng of mycolactone collected on a swab. Author summary Buruli ulcer is a neglected tropical disease that affects the poorest of the poorest in Africa. Even young people can harbor large ulcers, with raw flesh directly exposed. While antibiotic treatments exist, there are no simple methods to diagnose the disease. Here, we attempted to detect mycolactone, a small molecule produced by the bacteria that causes Buruli ulcer. One difficulty was the sample type to be used, as the open wounds are sampled with a swab. The question was how to extract mycolactone from the swab and then detect it by means of a rapid diagnostic test, without using sophisticated equipment. We tried to answer this question by combining monoclonal antibodies specific for mycolactone and novel magnetic nanoparticles to concentrate and visualize mycolactone with a rapid test. ### Competing Interest Statement GP was a member of the WHO Technical Advisory Group on Buruli ulcer and participated in the WHO?FIND meeting on diagnostics for Buruli ulcer in 2018. ### Funding Statement Yes ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: Clinical samples from Ghana were taken at the hospital or health centres as part of the routine diagnostic procedure and case confirmation recommended by WHO, and sent to the NMIMR for analysis. Ethical approval for collection and use of patients’ samples was obtained from the Institutional Review Board of the Noguchi Memorial Institute for Medical Research (NMIMR Study number: NMIMR-IRB CPN 024/18–19). Patients were required to sign an informed consent. To ensure anonymity, personal information such as names, and other personal identifiers associated with patient’s samples were replaced with codes. Clinical Samples from Ivory Coast and Cameroon were collected under IRB approved protocols for the sake of this study and with informed consent. The protocols were cleared by the Comité National d’Ethique des Sciences de la Vie et de la Santé, Ministère de la Santé, Côte d’Ivoire, 2021, IRB 000111917 and the Comité National d’Ethique de La Recherche pour la Santé Humaine, Ministère de la Santé, Cameroun, IRB 2021/06/1367. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All relevant data is included in the manuscript