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ABSTRACT |
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The name BU suggests that the disease appears only as ulcers of the skin; however, the clinical evolution of the disease is quite variable. Most authorities divide lesions of BU in the skin to three forms.9 The first is the non-ulcerative form, which appears as subcutaneous nodules, limited indurated plaques, or widespread edematous indurations. These lesions are usually not painful but may itch. Systemic inflammatory responses are minimal or absent. The second is the ulcerative form, (Figure 1
), which is characterized by ulcers of widely varying sizes with undermined edges that tend to extend, but are well defined. Advanced or long-standing indurated plaques and edematous forms may ulcerate, but such ulcers are ragged and poorly defined. Secondary infections may provoke pain. The third is the end stage or healing forms. Healing may take many months and sometimes is punctuated by local recurrences of active disease. Healing usually begins in the nondependent area of the ulcer. Lesions tend to resolve spontaneously but the resulting scars often lead to disabling or disfiguring sequelae (Figure 1). Bone lesions also exist in BU. The damaged bone can be situated just under the lesion (contact bone lesion) or remote from the ulcer (metastatic bone lesion).
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In Cameroon, BU was described for the first time in 1969 in 47 cases, all originating from a well-circumscribed area in the valley of the Nyong River in the Center Province between the villages of Ayos and Akonolinga.13–15 After its first description, this focus did not arouse any particular interest among either health authorities or public health professionals. Recently, however, District Leprosy Officers noted large, necrotizing chronic skin ulcers resistant to therapy in several children and adults in the same region. Enquiries among the local health personnel and the local population showed that they knew the clinical appearance and development of BU lesions very well. They were referred to in the local dialect as "Atom". The great majority of patients with "Atom," which was supposed to be caused by malediction, were under treatment by traditional healers. These findings suggested that BU, already described in the late 1960s in this area, had continued to exist or had reappeared. In this report, we present the results of a cross-sectional survey performed in August 2001 assessing the magnitude of the BU problem in the Nyong River basin in the Center Province of Cameroon.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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) with a population of approximately 98,500, was initially identified as the endemic region. The area has a population density of less than 50 people per km2 and many villages have fewer than 2,000 residents. The impoverished population of the area lives mainly by subsistence agriculture and informal commerce.
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Case definition. For the purpose of the survey, cases were defined clinically as active BU and inactive BU. Active BU was defined as 1) a nodule or plaque or edema consistent with BU, 2) chronic ulcers with subcutaneous tissue necrosis, undermined edges and peripheral induration, which were painless unless secondarily infected, with no constitutional symptoms, except when osteomyelitis was present, or 3) healing ulcers with a history and healing aspect consistent with BU. Inactive (healed) BU were defined as scar tissue on a former lesion deeply sunk into surrounding healthy skin (stellate aspect), with or without complications (e.g., contractures and/or ankylosis) leading to invalidity. A new case was defined as a patient with no previous history of or treatment of BU, and a recurrent case was defined as a patient presenting within one year from the end of the last treatment with a new lesion at the same or a different site.
Data collection. During a two-week survey, all settlements and villages identified in the first enquiry as having possible cases of BU were visited. Records were made of each patient after informed consent was given on a pre-established data form adapted from the Buruli ulcer form recommended by WHO.9 The study was reviewed and approved by the Ministry of Health in Cameroon. Information was collected on identity, residence, presence of other cases in the family or among people who had lived or were living in the same household, duration of lesions, and previous treatment. Possible risk factors and protective factors, such as distance of residence from the Nyong River, sources of water, and vaccination with bacilli Calmette-Guérin (BCG) were also recorded. The patients were thoroughly examined and the lesions were recorded according to the data form. Lesions were defined as active or inactive, and the location of the lesion was marked on a drawing of a human body. The largest diameter of ulcerative lesions was measured. The presence of complications was also reported. A photograph of the affected part of the body was taken with the patient’s permission.
Sample collection. Swab specimens were taken from all cases with active BU excluding pre-ulcerative lesions and ulcers in an advanced stage of healing. Samples were taken with a cotton swab from under the undermined edges of the ulcers or areas of drainage. Smears were sent twice a week to the Centre Pasteur du Cameroun for Ziehl-Neelsen staining and microscopic examination to identify acid-fast bacilli. Samples for polymerase chain reaction (PCR) detection of M. ulcerans DNA were stored in a cool box and maintained at 4°C until they were sent to the Institute of Tropical Medicine in Ant-werp and/or to the Swiss Tropical Institute in Basel.
IS2404 PCR. DNA was released from the dry swabs essentially as described previously using heat (15 minutes at 95°C) and alkaline lysis (0.2% sodium dodecyl sulfate, 0.05 M NaOH).9 Additional proteinase K digestion and vortexing with glass beads was found to improve the release of DNA. After extraction and precipitation of total DNA, IS2404 PCR amplification was done for 35 cycles essentially as described previously using HotStart DNA Polymerase (Qiagen, Basel, Switzerland).16 The three-room PCR principal was applied and negative controls were included to avoid or detect potential cross-contaminations.9 Inhibition of the PCR was overcome in a number of cases by repeating the PCR using a 1:10 dilution of the template solution.9 The size of the amplification products with a predicted size of 568 basepairs was analyzed by comparison with positive controls using visualization of DNA fragments after electrophoresis by ultraviolet transillumination of gels stained with ethidium bromide. Identity of amplification products was verified further by Southern blotting using an IS2404 probe.
Statistical analysis. Data were collected and analyzed using the Epi-Info software program (version 6.04fr, Centers for Disease Control and Prevention, Atlanta, GA). The chi-square test was used to elicit associations between dichotomous variables. Continuous variables were analyzed using the Student’s t-test or analysis of variance as appropriate. Two-tailed tests were used, with P < 0.05 considered significant.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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A subsequent cross-sectional survey performed in August 2001 identified on clinical grounds 436 BU cases in 83 different villages and settlements. Of these 436 cases 202 (46%) presented with active lesions while 234 (54%) were inactive cases. Twenty-five of 202 active cases (12.4%) were recurrent cases. While the overall prevalence of active and inactive BU in the surveyed area was 0.44%, the highest prevalence for active cases found in a particular settlement was 8%. Disease prevalence for active and/or inactive cases was higher in villages closer to the Nyong River (Figure 2
). It was principally the rural, impoverished part of the population with limited geographic and economic access to health facilities that was affected. Sixty-six (28.2%) of the 234 cases with healed ulcers had severe chronic functional disabilities as a result of contraction deformities. In one case, amputation of a finger was observed.
The age and sex distribution of the 202 cases with clinically diagnosed active BU are shown in Figure 3. A total of 115 (56.9%) of these were males and 87 (43.1%) were females, resulting in a male:female sex ratio of 1.3:1. The age of patients with active BU ranged from 2 to 90 years with a median age of 14.5 years. Fifty percent of the cases were less than 15 years of age, but no case was observed in children less than two years of age. The rate of illness did not differ significantly between the sexes or between different age groups.
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. Most lesions (93%) were located on the extremities, with the lower limbs being involved twice as often as the upper limbs. The distribution of lesions by site of localization on the body did not show any significant differences with respect to age or sex. The right or left distribution of lesions was similar both in the upper and the lower limbs (P = 0.20 and P = 0.12, respectively).
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A BCG vaccination scar was observed in 105 (52.0%) of the 202 active cases. Multiple lesions defined as lesions present simultaneously on different parts of the body were observed more often in children (
15 years old) without a BCG scar compared with those with the scar (7 of 20 versus 1 of 30; P < 0.01) suggesting a protective effect of this vaccination against more severe forms of BU in this age group. No association was found for bone involvement in BU even after stratifying patients by age groups.
One hundred seventeen cases (57.9%) said that a family member or relative had previously had or currently had BU. In one family, three children had active BU at the same time. This also occurred in two children each in three other families. Relatives of patients with active BU, or members of their households who had had BU, were asked when the illness had occurred. Seventy-five percent reported that it was within the last three years, and 5% reported that it occurred more than 10 years ago. The different types of sources of (drinking) water (water tap, river, well, spring) were not significantly associated with the presence of BU. Patients probably went to the swamps for other domestic activities.
An analysis of the health-seeking behavior of the cases revealed that more than two-thirds (67.3%) of them were exclusively under treatment given by traditional healers and less than 10% of them were being treated exclusively by trained health personnel.
Samples for laboratory confirmation of the presence of M. ulcerans in the lesions were taken from 164 (81.1%) of the 202 clinically diagnosed active cases. Samples could not be taken from the other 38 cases either because they presented with non-ulcerative lesions such as nodules, plaques, or edema (n = 15) or because they presented with ulcers in an advanced stage of healing (n = 23). Direct microscopic examination of the 164 samples stained by the Ziehl-Neelsen technique revealed acid-fast bacilli in 38 (23.2%) cases. A total of 162 samples were analyzed by the IS2404 PCR technique (two PCR samples were lost). One hundred thirty-five (83.3%) of these samples were positive for M. ulcerans DNA. In 34 cases, both Ziehl-Neelsen staining and the PCR were positive.
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DISCUSSION |
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The differential diagnosis of the clinically identified 15 pre-ulcerative lesions (i.e., four cases with nodules, seven with plaques, and four with edema) was most problematic since it had to be done without any laboratory reconfirmation. All cases with inactive (healed) BU also had to be diagnosed exclusively clinically. Thus, diagnostic misclassification may have overestimated the overall prevalence of BU. Conversely, it is generally admitted that the clinical diagnosis of ulcerating BU by an experienced clinician is relatively straightforward in a known endemic area.17 Other common chronic ulcerative lesions encountered in Cameroon (tropical and venous ulcers, ulcers due to burns) can be differentiated from BU by their clinical aspects and case history. We therefore included all the 202 clinically diagnosed cases with active lesions in the analyses of the clinical presentation and of possible risk and protective factors.
If one supposes that clinical diagnosis together with partial confirmation by laboratory procedures was mostly correct, we may ask, alternatively, whether it was likely that we missed a substantial number of cases. The delimitation of the survey area was empirical, depending substantially on the ability of the local population to identify the disease. However, the population and peripheral local health personal appeared to identify BU even in its early, pre-ulcerative forms surprisingly well. Conversely, we may have missed cases because presentation was voluntary and this could have resulted in a selection bias. Therefore, the results of the survey may rather have underestimated, rather than overestimated, prevalence. On various occasions, we even noticed traditional healers forbidding their patients to present themselves to the survey team. People with inactive BU were not included in the analysis of the clinical presentation and possible risk and protective factors for two reasons. First, we assumed that the presentation of cases on a voluntary basis would induce a selection bias towards active cases. Second, case histories from people with lesions were likely to be less accurate when the disease was acute some years ago.
The endemic area had characteristics similar to those found in foci in other African countries,4–7 and the clinical characteristics of active cases notified resemble what has been found in similar situations.14 The endemic area is located in the swampy banks along the slow-flowing Nyong River and some of its tributaries, and is isolated. The frequency of identified cases decreased with the distance to the river. Similar to findings in Côte d’Ivoire,18 more than half of the cases were children 15 years old, with the highest rate of infection in those 10–14 years old. No preponderance of women in the group of adult (
15 years old) patients was found, as reported in earlier descriptions of this focus and in other studies.6–13 The ulcerative forms predominated in prevalent active BU cases; our finding of 92.8% is consistent with the 89.5% or 96.4% found in prevalence studies in Côte d’Ivoire and in Benin, respectively.5,6 However, in a more recent study performed with 1,700 patients from Benin (from 1997 to 2001), there was no statistically significant difference between the percentages of ulcerated and non-ulcerated forms. The increasing number of non-ulcerated forms detected from 1997 to 2001 is a result of increasing activity of public health programs that raised awareness of BU clinical forms (Debacker M and others, unpublished data).
Our study showed a relatively high frequency (14.9%) of bone involvement, although similar frequencies have been reported in other studies.19 This high frequency may have been due to a selection bias resulting from our case-finding method and use of clinical diagnosis. Lesions on the limbs were predominant. In the first, restricted case series of the Nyong River focus, upper limbs were predominantly affected, but our findings confirm the general finding of lower limbs being most affected.13 Even after stratification for sex and age, no unequal right-left distribution of lesions on limbs was found, as reported in some other studies.4,17,18 Similarly, in a recent study in Benin, there were no significant differences between the number of lesions on the right or left limbs (Debacker M and others, unpublished data).
In 1969, the Uganda Buruli Group demonstrated in a controlled trial that BCG vaccination confers partial protection against BU for at least six months.20 This finding was confirmed by Smith and others in 1976.21 In Benin, BCG vaccination appeared to partially protect children less than 15 years of age with confirmed BU against the osseous forms of BU.22 In our study population, children not vaccinated with BCG appeared to have a significantly higher risk of multiple lesions, confirming the partial protective effect of BCG vaccination against severe forms of BU in children.
Buruli ulcer tends to affect several members of the same family, which was previously observed in early studies of the disease in Uganda.2 Reported figures of the proportion of households with several BU cases are scarce, varying between 0% and 5%.13,23 Our figure of approximately 58% of a history of cases in the family or among people living in the same compound appears high. However, it is not comparable with figures reported in other studies because our questionnaire did not differentiate between other family members currently with BU and family members known to have had BU in the past. The data suggest that within an endemic area, scattered disease foci of various importance subsist over time.
Even with only partial laboratory reconfirmation, we conclude on the basis of clinical and epidemiologic evidence that the area surveyed constitutes a focus of emergent endemic BU of considerable magnitude. The accumulation of family cases during the past three years suggests that BU has recently re-emerged on a larger scale in the survey area. However, this focus has probably existed without interruption, especially if one considers the distribution of active and inactive cases without major transposition of the different individual foci, for more than 30 years in a well-circumscribed area without triggering off any intervention by health authorities. Further support was given to the idea that the disease had never really disappeared from the area by the fact that it was evidently well known and described by the local population.
The preference of patients for traditional medicine reflects a situation in which the health system has had nothing to offer to assist these patients and, consequently, where the population has lost confidence in the ability of the health system to provide assistance. Currently, surgery is the only proven effective treatment of M. ulcerans disease.8 Limiting factors include inadequate surgical facilities, the need for a prolonged stay in the hospital, and high treatment costs. If one estimates that the mean duration of the disease without medical intervention is one year, the annual incidence could be estimated to be 100–150 new cases of BU in an area largely identical with the survey area. With more than 25% of these patients being at high risk of developing severe long-term disabilities, a comprehensive public health intervention by health authorities and the health districts concerned is urgently needed. Control strategies promoted by the Global Buruli Ulcer Initiative include strengthening of the health care capacity in BU-endemic areas by upgrading surgical facilities, health education, and staff training in the communities to promote early detection and rapid referral, development of motivational strategies, and rehabilitation of those already deformed by this disease.8
Received February 8, 2003. Accepted for publication May 29, 2003.
Acknowledgments: We thank Dr. Ayissi Christopher (Delegate of Public Health of the Centre Province) for his assistance. We are also grateful to the late Simon Bitoto and Daniel Ze Bekolo (Leprosy District Officers for Ayos and Akonolinga Health Districts), as well as to Petrus Nkamsse and Arlette Ngassam for the invaluable help they gave us in different aspects of this study. We also thank Dr. Thomas A. Smith for helping with the design of a map of the survey area showing prevalence by location of the survey area and Jeniffer Jenkins for editing the manuscript.
Financial support: This study was funded by Aide aux Lépreux Emmaüs-Suisse and Médecins Sans Frontières Switzerland. Françoise Portaels was supported by the Fund for Scientific Research, Flanders (Belgium) (FWO-Vlaanderen contract no. G-0301-01).
Authors’ addresses: Jürgen Noeske, Kolveniersstraat 6, Bus 7, B 2000 Antwerp, Belgium. Christopher Kuaban and Josephine Mbuagbaw, Centre Pasteur du Cameroun, BP 1274, Yaounde, Cameroon. Simona Rondini and Gerd Pluschke, Molecular Immunology, Swiss Tropical Institute, Socinstrasse 57, CH 4002, Basel, Switzerland, Telephone: 41-61-284-8235, Fax.: 41-61-271-8654, E-mail: Gerd.Pluschke{at}unibas.ch. Philippe Sorlin, Centre Pasteur du Cameroun, Yaounde, Cameroon. Laura Ciaffi, BP 30139, Yaoundé, Cameroon. Françoise Portaels, Mycobacteriology Unit, Institute of Tropical Medicine, Nationalestraat 155, B 2000 Antwerp, Belgium.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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