Treatment Outcome of Patients with Buruli Ulcer Disease in Togo

Background Following introduction of antimycobacterial treatment of Buruli ulcer disease (BUD),several clinical studies evaluated treatment outcomes of BUD patients, in particular healing times, secondary lesions and functional limitations. Whereas recurrences were rarely observed, paradoxical reactions and functional limitations frequently occurred. Although systematic BUD control in Togo was established as early as 2007, treatment outcome has not been reviewed to date. Therefore, a pilot project on post-treatment follow-up of BUD patients in Togo aimed to evaluate treatment outcomes and to provide recommendations for optimization of treatment success. Methodology/Principal Findings Out of 199 laboratory confirmed BUD patients, 129 could be enrolled in the study. The lesions of 109 patients (84.5%) were completely healed without any complications, 5 patients (3.9%) had secondary lesions and 15 patients (11.6%) had functional limitations. Edema, category III ulcers >15cm, healing times >180 days and a limitation of movement at time of discharge constituted the main risk factors significantly associated with BUD related functional limitations (P180 days and limitation of movement at discharge constituted the main risk factors for functional limitations in Togolese BUD patients. Standardized treatment plans, patient assessment and follow-up, as well as improved management of medical records are recommended to allow for intensified monitoring of disease progression and healing process, to facilitate implementation of therapeutic measures and to optimize treatment success

Implementation of a National Reference Laboratory for Buruli Ulcer Disease in Togo

Background: In a previous study PCR analysis of clinical samples from suspected cases of Buruli ulcer disease (BUD) from Togo and external quality assurance (EQA) for local microscopy were conducted at an external reference laboratory in Germany. The relatively poor performance of local microscopy as well as effort and time associated with shipment of PCR samples necessitated the implementation of stringent EQA measures and availability of local laboratory capacity. This study describes the approach to implementation of a national BUD reference laboratory in Togo. Methodology: Large scale outreach activities accompanied by regular training programs for health care professionals were conducted in the regions "Maritime'' and "Central,'' standard operating procedures defined all processes in participating laboratories (regional, national and external reference laboratories) as well as the interaction between laboratories and partners in the field. Microscopy was conducted at regional level and slides were subjected to EQA at national and external reference laboratories. For PCR analysis, sample pairs were collected and subjected to a dry-reagent-based IS2404-PCR (DRB-PCR) at national level and standard IS2404 PCR followed by IS2404 qPCR analysis of negative samples at the external reference laboratory. Principal Findings: The inter-laboratory concordance rates for microscopy ranged from 89% to 94%; overall, microscopy confirmed 50% of all suspected BUD cases. The inter-laboratory concordance rate for PCR was 96% with an overall PCR case confirmation rate of 78%. Compared to a previous study, the rate of BUD patients with non-ulcerative lesions increased from 37% to 50%, the mean duration of disease before clinical diagnosis decreased significantly from 182.6 to 82.1 days among patients with ulcerative lesions, and the percentage of category III lesions decreased from 30.3% to 19.2%. Conclusions: High inter-laboratory concordance rates as well as case confirmation rates of 50% (microscopy), 71% (PCR at national level), and 78% (including qPCR confirmation at external reference laboratory) suggest high standards of BUD diagnostics. The increase of non-ulcerative lesions, as well as the decrease in diagnostic delay and category III lesions, prove the effect of comprehensive EQA and training measures involving also procedures outside the laboratory

Effectiveness of Routine BCG Vaccination on Buruli Ulcer Disease: A Case-Control Study in the Democratic Republic of Congo, Ghana and Togo

Background: The only available vaccine that could be potentially beneficial against mycobacterial diseases contains live attenuated bovine tuberculosis bacillus (Mycobacterium bovis) also called Bacillus Calmette-Guerin (BCG). Even though the BCG vaccine is still widely used, results on its effectiveness in preventing mycobacterial diseases are partially contradictory, especially regarding Buruli Ulcer Disease (BUD). The aim of this case-control study is to evaluate the possible protective effect of BCG vaccination on BUD. Methodology: The present study was performed in three different countries and sites where BUD is endemic: in the Democratic Republic of the Congo, Ghana, and Togo from 2010 through 2013. The large study population was comprised of 401 cases with laboratory confirmed BUD and 826 controls, mostly family members or neighbors. Principal Findings: After stratification by the three countries, two sexes and four age groups, no significant correlation was found between the presence of BCG scar and BUD status of individuals. Multivariate analysis has shown that the independent variables country (p = 0.31),sex (p = 0.24),age (p = 0.96),and presence of a BCG scar (p = 0.07) did not significantly influence the development of BUD category I or category II/III. Furthermore, the status of BCG vaccination was also not significantly related to duration of BUD or time to healing of lesions. Conclusions: In our study, we did not observe significant evidence of a protective effect of routine BCG vaccination on the risk of developing either BUD or severe forms of BUD. Since accurate data on BCG strains used in these three countries were not available, no final conclusion can be drawn on the effectiveness of BCG strain in protecting against BUD. As has been suggested for tuberculosis and leprosy, well-designed prospective studies on different existing BCG vaccine strains are needed also for BUD

Effectiveness of Routine BCG Vaccination on Buruli Ulcer Disease: A Case-Control Study in the Democratic Republic of Congo, Ghana and Togo

Background: The only available vaccine that could be potentially beneficial against mycobacterial diseases contains live attenuated bovine tuberculosis bacillus (Mycobacterium bovis) also called Bacillus Calmette-Guerin (BCG). Even though the BCG vaccine is still widely used, results on its effectiveness in preventing mycobacterial diseases are partially contradictory, especially regarding Buruli Ulcer Disease (BUD). The aim of this case-control study is to evaluate the possible protective effect of BCG vaccination on BUD. Methodology: The present study was performed in three different countries and sites where BUD is endemic: in the Democratic Republic of the Congo, Ghana, and Togo from 2010 through 2013. The large study population was comprised of 401 cases with laboratory confirmed BUD and 826 controls, mostly family members or neighbors. Principal Findings: After stratification by the three countries, two sexes and four age groups, no significant correlation was found between the presence of BCG scar and BUD status of individuals. Multivariate analysis has shown that the independent variables country (p = 0.31),sex (p = 0.24),age (p = 0.96),and presence of a BCG scar (p = 0.07) did not significantly influence the development of BUD category I or category II/III. Furthermore, the status of BCG vaccination was also not significantly related to duration of BUD or time to healing of lesions. Conclusions: In our study, we did not observe significant evidence of a protective effect of routine BCG vaccination on the risk of developing either BUD or severe forms of BUD. Since accurate data on BCG strains used in these three countries were not available, no final conclusion can be drawn on the effectiveness of BCG strain in protecting against BUD. As has been suggested for tuberculosis and leprosy, well-designed prospective studies on different existing BCG vaccine strains are needed also for BUD

Inclusion and exclusion criteria.

<p>Table 1 shows inclusion and exclusion criteria for study participants.</p><p>Inclusion and exclusion criteria.</p

Results of external quality assurance for microscopy.

<p><a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002011#pntd-0002011-t003" target="_blank">Table 3</a> shows results of Ziehl-Neelsen microscopy conducted at CHR and corresponding results of EQA conducted at INH and DITM during the initial (phase I, September 2010 through December 2010) and transitional (phase II, January 2011 through April 2012) phases. N/A, not applicable.</p>a<p>Swab, slides were prepared as direct smears from swab samples.</p>b<p>FNA, slides were prepared as direct smears from fine-needle aspirate samples.</p>c<p>Positivity rate, number of positive samples divided by the total number of samples tested.</p>d<p>Rate of false negative results at CHR and INH, respectively, compared to DITM results.</p>e<p>Rate of false positive results at CHR and INH, respectively, compared to DITM results.</p>f<p>Rate of false negative and false positive results at CHR and INH, respectively, compared to DITM results.</p>g<p>Rate of concordant results between CHR/INH, CHR/DITM and INH/DITM.</p>h<p>Confirmation rate, number of laboratory confirmed BUD patients divided by the total number of suspected BUD cases.</p

Clinical samples analyzed by microscopy for <i>M. ulcerans</i>.

<p><a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002011#pntd-0002011-t002" target="_blank">Table 2</a> indicates all slides prepared from swab or FNA samples and subjected to Ziehl-Neelsen staining at “Centre Hospitalier Régional” (CHR) for the detection of acid fast bacilli. Slides were analyzed consecutively at CHR and the Department of Infectious Diseases and Tropical Medicine (DITM), Ludwig-Maximilians-University during initial phase (phase I) or CHR, at the “Institut National d'Hygiène” (INH) and DITM during transitional phase (phase II). N/A, not applicable.</p>a<p>MIC, microscopic detection of acid fast bacilli.</p>b<p>Total, number of slides prepared from swab and FNA samples and subjected to reading at CHR/DITM or CHR/INH/DITM.</p>c<p>Phase I, initial phase of implementation of the national reference laboratory at INH from September 2010 through December 2010; slides were read at CHR and forwarded via DAHWT to DITM for EQA.</p>d<p>Phase II, transitional phase of implementation of the national reference laboratory at INH from January 2011 through April 2012; slides were read at CHR, followed by blinded re-reading at INH and DITM.</p

Stepwise approach to implementation of diagnostic laboratory facilities at INH.

<p><a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002011#pntd-0002011-g001" target="_blank">Figure 1</a> describes the process of implementation of diagnostic laboratory facilities at INH in three phases and the flow of samples as well as the flow of feedback between the Department for Infectious Diseases and Tropical Medicine (DITM), Ludwig-Maximilians-University, Munich, Germany, the “Institut National d'Hygiène” (INH), Lomé, Togo, the “Centre Hospitalier Régional Maritime” (CHR), Tsévié, Togo, and field staff. BUD, Buruli ulcer disease; CLT, “Contrôleur Lèpre-TB-Buruli” – district controllers; DRB-PCR, dry-reagent-based IS<i>2404</i> PCR; EQA, external quality assurance; MIC, microscopic detection of acid fast bacilli by Ziehl-Neelsen staining; PCR, polymerase chain reaction; qPCR, IS<i>2404</i> quantitative real-time PCR; standard PCR, conventional gel-based IS<i>2404</i> PCR; USP, “Unité de Soins Périphérique” – peripheral health posts.</p

Clinical samples analyzed by PCR for <i>M. ulcerans</i>.

<p><a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002011#pntd-0002011-t004" target="_blank">Table 4</a> indicates all samples tested by PCR at IHN and DITM. During the initial phase (phase I) samples were analyzed by standard gel-based IS<i>2404</i> PCR at DITM. During the second phase (phase II) parallel samples were subjected to IS<i>2404</i> dry-reagent based (DRB) PCR at INH and standard IS<i>2404</i> PCR at DITM. During both phases all samples tested negative in standard PCR were subjected to re-testing by IS<i>2404</i> quantitative real-time PCR (qPCR) at DITM.</p>a<p>Swab, DNA extracts prepared from swab samples.</p>b<p>FNA, DNA extracts prepared from fine-needle aspirate samples.</p>c<p>Punch, DNA extracts prepared from 3 mm punch biopsy samples.</p>d<p>Phase I, initial phase of implementation of the national reference laboratory at INH from September through December 2010.</p>e<p>Only samples tested negative in standard IS<i>2404</i> PCR were subjected to IS<i>2404</i> qPCR at DITM.</p>f<p>Total amount of samples tested by DRB- and Standard PCR during the corresponding phases.</p>g<p>Phase II, transitional phase of implementation of the national reference laboratory at INH from January 2011 through April 2012.</p

Age distribution of laboratory confirmed BUD patients.

<p>Age distribution of 64 laboratory confirmed BUD patients recruited from September 2010 through April 2012. The age of all patients was known and 48.4% (31/64) were in age group 5–14 years. The age range was 2–68 years with a mean of 18.1 years and a median of 13 years.</p