- Introduction
- Materials and Methods
- Ethics statement.
- Patients.
- Clinical samples and laboratory confirmation.
- Samples subjected to sequence analysis.
- Amplification of rpoB and rpsL genes.
- Gel extraction of PCR products.
- DNA sequencing.
- Species identification.
- Sequence analysis.
- Comparison of whole-genome extracts with culture extracts.
- Comparison of whole-genome extracts with corresponding culture extracts.
- Statistical analysis.
- Results
- Discussion
ProCite
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A Genotypic Approach for Detection, Identification, and Characterization of Drug Resistance in Mycobacterium ulcerans in Clinical Samples and Isolates from Ghana
Marcus Beissner
Marcus Beissner
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Nana-Yaa Awua-Boateng
Nana-Yaa Awua-Boateng
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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William Thompson
William Thompson
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Willemien A. Nienhuis
Willemien A. Nienhuis
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Erasmus Klutse
Erasmus Klutse
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Pius Agbenorku
Pius Agbenorku
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Joerg Nitschke
Joerg Nitschke
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Karl-Heinz Herbinger
Karl-Heinz Herbinger
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Vera Siegmund
Vera Siegmund
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Erna Fleischmann
Erna Fleischmann
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Ohene Adjei
Ohene Adjei
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Bernhard Fleischer
Bernhard Fleischer
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Tjip S. van der Werf
Tjip S. van der Werf
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Thomas Loscher
Thomas Loscher
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Gisela Bretzel
Gisela Bretzel
Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany; Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology (KNUST), Kumasi, Ghana; Agogo Presbyterian Hospital, Agogo, Ghana; University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands; Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana; Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana; Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany
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Standardized antimycobacterial therapy is considered the treatment of choice for Buruli ulcer disease. To assess the prevalence of drug resistance among clinical Mycobacterium ulcerans isolates in Ghana, we conducted a sequence-based approach to detect mutations associated with drug resistance. We subjected clinical samples to direct DNA sequencing of rpoB and rpsL genes and compared culture and whole-genome extracts regarding the efficiency of sequence analysis; 99.1% (rpoB) and 100% (rpsL) of the patients harbored M. ulcerans wild type. In one isolate (0.9%), a point mutation of the rpoB gene at codon Ser522 leading to an amino acid change was detected. Culture extracts yielded a significantly higher sequencing efficiency than whole-genome extracts. Our data suggest a low level of drug resistance in Ghana. However, mutations associated with drug resistance do occur and require monitoring. Improved techniques are necessary to enhance the efficiency of sequence analysis of whole-genome extracts.
Author Notes
Financial support: This project was supported by the European Commission (Project INCO-CT-2005-015476-BURULICO).
Authors' addresses: Marcus Beissner, Joerg Nitschke, Karl-Heinz Herbinger, Vera Siegmund, Erna Fleischmann, Thomas Loscher, and Gisela Bretzel, Department of Infectious Diseases and Tropical Medicine (DITM), University Hospital, Ludwig-Maximilians University of Munich, Munich, Germany, E-mails: marcus.beissner@lrz.uni-muenchen.de, j.a.nitschke@web.de, herbinger@lrz.uni-muenchen.de, v.siegmund@eurice.de, erna.fleischmann@lrz.uni-muenchen.de, loescher@lrz.uni-muenchen.de, and bretzel@lrz.uni-muenchen.de. Nana-Yaa Awua-Boateng and Ohene Adjei, Kumasi Centre for Collaborative Research in Tropical Medicine (KCCR), Kwame Nkrumah University of Science and Technology, Kumasi, Ghana, E-mails: awua.boateng@kccr.de and oadjei@africaonline.com.gh. William Thompson, Agogo Presbyterian Hospital, Agogo, Ghana, E-mail: wnat@agogohospital.org. Willemien A. Nienhuis and Tjip S. van der Werf, University Medical Centre Groningen (UMCG), University of Groningen, Groningen, The Netherlands, E-mails: wiannix@hotmail.com and t.s.van.der.werf@int.umcg.nl. Erasmus Klutse, Dunkwa Governmental Hospital, Dunkwa-on-Offin, Dunkwa, Ghana, E-mail: eyklutse@yahoo.com. Pius Agbenorku, Reconstructive Plastic Surgery and Burns Unit, Department of Surgery, Komfo Anokye Teaching Hospital, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana, E-mail: pimagben@yahoo.com. Bernhard Fleischer, Bernhard Nocht Institute for Tropical Medicine (BNITM), Hamburg, Germany, E-mail: fleischer@bni-hamburg.de.
Reprints requests: Gisela Bretzel, Department of Infectious Diseases and Tropical Medicine, University Hospital, Ludwig-Maximilians University of Munich, Leopoldstrasse 5, 80802 Munich, Germany, E-mail: bretzel@lrz.uni-muenchen.de.
- Introduction
- Materials and Methods
- Ethics statement.
- Patients.
- Clinical samples and laboratory confirmation.
- Samples subjected to sequence analysis.
- Amplification of rpoB and rpsL genes.
- Gel extraction of PCR products.
- DNA sequencing.
- Species identification.
- Sequence analysis.
- Comparison of whole-genome extracts with culture extracts.
- Comparison of whole-genome extracts with corresponding culture extracts.
- Statistical analysis.
- Results
- Discussion
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Reference Manager
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Zotero
EndNote
-
1.
World Health Organization, 2008. Buruli ulcer: progress report, 2004–2008. Wkly Epidemiol Rec 17: 145–154.
-
2.
World Health Organization, 2004. Provisional Guidance on the Role of Specific Antibiotics in the Management of Mycobacterium ulcerans Disease (Buruli Ulcer). Geneva, Switzerland: World Health Organization.
-
3.
Siegmund V, Adjei O, Nitschke J, Thompson W, Klutse E, Herbinger KH, Thompson R, van Vloten F, Racz P, Fleischer B, Loescher T, Bretzel G, 2007. Dry reagent-based polymerase chain reaction compared with other laboratory methods available for the diagnosis of Buruli ulcer disease. Clin Infect Dis 45: 68–75.
-
4.
Schunk M, Thompson W, Klutse E, Nitschke J, Opare-Asamoah K, Thompson R, Fleischmann E, Siegmund V, Herbinger KH, Adjei O, Fleischer B, Loscher T, Bretzel G, 2009. Outcome of patients with Buruli ulcer after surgical treatment with or without antimycobacterial treatment in Ghana. Am J Trop Med Hyg 81: 75–81.
-
5.
Herbinger KH, Adjei O, Awua-Boateng NY, Nienhuis WA, Kunaa L, Siegmund V, Nitschke J, Thompson W, Klutse E, Agbenorku P, Schipf A, Reu S, Racz P, Fleischer B, Beissner M, Fleischmann E, Helfrich K, van der Werf TS, Loscher T, Bretzel G, 2009. Comparative study of the sensitivity of different diagnostic methods for the laboratory diagnosis of Buruli ulcer disease. Clin Infect Dis 48: 1055–1064.
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