title: RNA Aptamer That Specifically Binds to Mycolactone and Serves as a Diagnostic Tool for Diagnosis of Buruli Ulcer.
abstract: BACKGROUND
  Buruli ulcer (BU) is a subcutaneous skin disease listed among the neglected tropical diseases by the World Health Organization (WHO). Early case detection and management is very important to reduce morbidity and the accompanied characteristic disfiguring nature of BU. Since diagnosis based on clinical evidence can lead to misdiagnosis, microbiological confirmation is essential to reduce abuse of drugs; since the anti-mycobacterial drugs are also used for TB treatment. The current WHO gold standard PCR method is expensive, requires infrastructure and expertise are usually not available at the peripheral centers where BU cases are managed. Thus one of the main research agendas is to develop methods that can be applied at the point of care. In this study we selected aptamers, which are emerging novel class of detection molecules, for detecting mycolactone, the first to be conducted in a BUD endemic country.
  
  
  METHODS
  Aptamers that bind to mycolactone were isolated by the SELEX process. To measure their affinity and specificity to mycolactone, the selected aptamers were screened by means of isothermal titration calorimetry (ITC) and an enzyme-linked oligonucleotide assay (ELONA). Selected aptamers were assessed by ELONA using swab samples from forty-one suspected BU patients with IS2404 PCR and culture as standard methods. ROC analysis was used to evaluate their accuracy and cutoff-points.
  
  
  RESULTS
  Five out of the nine selected aptamers bound significantly (p< 0.05) to mycolactone, of these, three were able to distinguish between mycolactone producing mycobacteria, M. marinum (CC240299, Israel) and other bacteria whilst two others also bounded significantly to Mycobacterium smegmatis. Their dissociation constants were in the micro-molar range. At 95% confidence interval, the ROC curve analysis among the aptamers at OD450 ranged from 0.5-0.7. Using this cut-off for the ELONA assay, the aptamers had 100% specificity and sensitivity between 0.0% and 50.0%. The most promising aptamer, Apt-3683 showed a discernible cleavage difference relative to the non-specific autocatalysis over a 3-minute time course.
  
  
  CONCLUSIONS
  This preliminary proof-of-concept indicates that diagnosis of BUD with RNA aptamers is feasible and can be used as point of care upon incorporation into a diagnostic platform.
authors: Department of Molecular Medicine, School of Medical Sciences, Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana.|Department of Bacteriology, Noguchi Memorial Institute for Medical Research, University of Ghana, Accra, Ghana.
authors: Samuel A
authors: http://orcid.org/0000-0001-5168-4762
authors: SA
authors: Sakyi
authors: Department of Bacteriology, Noguchi Memorial Institute for Medical Research, University of Ghana, Accra, Ghana.
authors: Samuel Yaw
authors: SY
authors: Aboagye
authors: Department of Bacteriology, Noguchi Memorial Institute for Medical Research, University of Ghana, Accra, Ghana.|Department of Biochemistry, Cell and Molecular Biology, University of Ghana, Accra, Ghana.
authors: Isaac Darko
authors: ID
authors: Otchere
authors: Aptagen LLC, Jacobus, Pennsylvania, United States of America.
authors: Albert M
authors: AM
authors: Liao
authors: Aptagen LLC, Jacobus, Pennsylvania, United States of America.
authors: Thomas G
authors: TG
authors: Caltagirone
authors: Department of Bacteriology, Noguchi Memorial Institute for Medical Research, University of Ghana, Accra, Ghana.|Department of Biochemistry, Cell and Molecular Biology, University of Ghana, Accra, Ghana.
authors: Dorothy
authors: D
authors: Yeboah-Manu
mesh_terms: D052157:Aptamers, Nucleotide / Q000235:genetics / Q000378:metabolism*; D054312:Buruli Ulcer / Q000175:diagnosis* / Q000453:epidemiology / Q000382:microbiology; D057075:Enzyme Assays / Q000379:methods*; D006801:Humans; D007557:Israel / Q000453:epidemiology; D018942:Macrolides / Q000378:metabolism*; D020102:Mycobacterium smegmatis / Q000378:metabolism; D019911:Mycobacterium ulcerans / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D019095:Point-of-Care Systems; D016133:Polymerase Chain Reaction; D012372:ROC Curve; D012680:Sensitivity and Specificity
chemical_list: D052157:Aptamers, Nucleotide; D018942:Macrolides; C117218:mycolactone