Molecular detection of Mycobacterium ulcerans in the environment and its relationship with Buruli ulcer occurrence in Zio and Yoto districts of maritime region in Togo (original) (raw)
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PLOS Neglected Tropical Diseases, 2008
Mycobacterium ulcerans, the causative agent of Buruli ulcer, is an emerging environmental bacterium in Australia and West Africa. The primary risk factor associated with Buruli ulcer is proximity to slow moving water. Environmental constraints for disease are shown by the absence of infection in arid regions of infected countries. A particularly mysterious aspect of Buruli ulcer is the fact that endemic and non-endemic villages may be only a few kilometers apart within the same watershed. Recent studies suggest that aquatic invertebrate species may serve as reservoirs for M. ulcerans, although transmission pathways remain unknown. Systematic studies of the distribution of M. ulcerans in the environment using standard ecological methods have not been reported. Here we present results from the first study based on random sampling of endemic and non-endemic sites. In this study PCR-based methods, along with biofilm collections, have been used to map the presence of M. ulcerans within 26 aquatic sites in Ghana. Results suggest that M. ulcerans is present in both endemic and non-endemic sites and that variable number tandem repeat (VNTR) profiling can be used to follow chains of transmission from the environment to humans. Our results suggesting that the distribution of M. ulcerans is far broader than the distribution of human disease is characteristic of environmental pathogens. These findings imply that focal demography, along with patterns of human water contact, may play a major role in transmission of Buruli ulcer.
Internal Medicine: Open Access, 2017
Buruli ulcer is a neglected skin disease caused by Mycobacterium ulcerans (MU), and affects 1000 people each year worldwide and particularly in African countries. Eradication of Buruli ulcer is difficult because of the lack of early diagnostic in rural endemic regions, and the unknown of the disease in national health care system. In the rural wetlands and marshes in Central and West Africa, children are most affected. MU belongs to environmental Mycolactone Producing Mycobacteria (MPMs) and presents high genetic diversity of the circulating strains. Diagnoses were applied by culture and genome detection by Polymerase Chain Reaction (PCR). The PCR became a gold method to confirm clinical and environmental samples for MU. The MIRU-VNTR method and the Restriction Fragment Length Polymorphism (RFLP) combined with MIRU-VNTR markers were used to discriminate Mycobacteria from different sources. The aim of this study was to investigate the molecular diversity of different mycobacteria sources, from environment, culture strains, and clinical samples by using combined MIRU-VNTR-Typing and RFLP. A total of 26 samples (water, sediment, mycobacteria strains, and swab) from endemic sites were first confirmed by IS2404 or Ziehl-Neelsen staining. The samples were analyzed by PCR typing for 4 specific markers (MIRU-1, VNTR6, VNTR19, ST-1) and the amplicons were digested with MspI restriction endonuclease and separated by 3% agarose gel electrophoresis for PCR-RFLP analysis. Our results showed different amplification by VNTR-MIRU-typing. For environmental samples a low amplification was detected by 25% for PCR-MIRU-1. Culture strains and clinical samples had amplification rate of 35.7% and 62.5% respectively. ST-1 had the best amplified marker for culture strains by 71.4%, while clinical samples have good amplification rate by 62.5 % for all markers. PCR-RFLP-profiles of clinical samples were identical, while environmental and mycobacteria strains showed different PCR-RFLP-profiles. We have developed a PCR-RFLP sensitive, easier and inexpensive approach to confirm genotyping of nontuberculosis mycobacteria in endemic countries for environmental screening. We suggest mutation in repeat loci of VNTR-MIRU sequence and the adaptation of mycobacteria from environment to human. This study confirms the circulation of several genotypes of mycobacteria in Ivory Coast.
Evaluation of real-time PCR for Mycobacterium ulcerans in endemic region in Côte d'Ivoire
African Journal of …, 2011
Mycobacterium ulcerans. The events of BU are the skin lesions. The lack of early diagnosis and treatment cause severe disability. Today the emergence to BU in Africa and particularly in Côte d'Ivoire needs faster diagnosis to control and to prevent the infection by M. ulcerans. The surveillance of BU is difficult, because the transmission of M. ulcerans occurs in rural regions where the transport of fresh collected sample is long, and the detection with culture technique needs several months. This study has allowed the application of polymerase chain reaction (PCR) technique in real time with two targets for molecular diagnosis of BU in Côte d' Ivoire. 63 samples (clinical, environmental, local strains and reference strains) were analyzed in realtime PCR by comparing the target of the Insertion Sequence (IS) 2404 and the sequence Ketoreductase-B (KR-B), located respectively on the chromosome and on the virulence plasmid. 49 samples (76%) were positive in real-time for both targets. The sensitivity of the PCR shows a detection limit of 0.25 genome copy for both targets. The capacity, speed and sensitivity of real-time PCR assays improve the diagnosis and contribute to strengthening the eradication of BU in Côte d'Ivoire.
Detection of Mycobacterium ulcerans in the Environment Predicts Prevalence of Buruli Ulcer in Benin
PLoS Neglected Tropical Diseases, 2012
Background: Mycobacterium ulcerans is the causative agent of Buruli ulcer (BU). In West Africa there is an association between BU and residence in low-lying rural villages where aquatic sources are plentiful. Infection occurs through unknown environmental exposure; human-to-human infection is rare. Molecular evidence for M. ulcerans in environmental samples is well documented, but the association of M. ulcerans in the environment with Buruli ulcer has not been studied in West Africa in an area with accurate case data.
Source Tracking Mycobacterium ulcerans Infections in the Ashanti Region, Ghana
PLoS neglected tropical diseases, 2015
Although several studies have associated Mycobacterium ulcerans (MU) infection, Buruli ulcer (BU), with slow moving water bodies, there is still no definite mode of transmission. Ecological and transmission studies suggest Variable Number Tandem Repeat (VNTR) typing as a useful tool to differentiate MU strains from other Mycolactone Producing Mycobacteria (MPM). Deciphering the genetic relatedness of clinical and environmental isolates is seminal to determining reservoirs, vectors and transmission routes. In this study, we attempted to source-track MU infections to specific water bodies by matching VNTR profiles of MU in human samples to those in the environment. Environmental samples were collected from 10 water bodies in four BU endemic communities in the Ashanti region, Ghana. Four VNTR loci in MU Agy99 genome, were used to genotype environmental MU ecovars, and those from 14 confirmed BU patients within the same study area. Length polymorphism was confirmed with sequencing. MU w...
PLoS Neglected Tropical Diseases, 2010
Buruli ulcer (BU) is an emerging necrotizing disease of the skin and subcutaneous tissue caused by Mycobacterium ulcerans. While proximity to stagnant or slow flowing water bodies is a risk factor for acquiring BU, the epidemiology and mode of M. ulcerans transmission is poorly understood. Here we have used high-throughput DNA sequencing and comparisons of the genomes of seven M. ulcerans isolates that appeared monomorphic by existing typing methods. We identified a limited number of single nucleotide polymorphisms (SNPs) and developed a real-time PCR SNP typing method based on these differences. We then investigated clinical isolates of M. ulcerans on which we had detailed information concerning patient location and time of diagnosis. Within the Densu river basin of Ghana we observed dominance of one clonal complex and local clustering of some of the variants belonging to this complex. These results reveal focal transmission and demonstrate, that micro-epidemiological analyses by SNP typing has great potential to help us understand how M. ulcerans is transmitted.
First Molecular Characterisation of Clinical Strains of Mycobacterium ulcerans in Togo (West Africa)
Microbiology Research Journal International
Background: Buruli ulcer is the third most common mycobacterial disease worldwide. Cases most occur in 30 countries but severe cases occur in West Africa countries such as Benin, Cote d’Ivoire and Togo mainly in rural regions. Early diagnosis may prevent severe disability. The molecular technique seems the best solution and new Mycobacterial Interspersed Repetitive Units (MIRU) and variable number tandem repeats (VNTR) typing method are themost reproducible in this regard. They propose geographical, inter and intraspecies differentiation and can be used as a diagnosis tool. Objective: The objective of this study was to investigate the molecular diversity by using MIRUVNTR typing in clinical samples of BU patients in Togo. Study Design: 64 DNA extracts from clinical samples were collected from BU patients in the two principal endemics districts in Togo (Yoto and Zio) with three less endemic districts (Bas Mono, Lacs and Vo). First, IS2404 and KR real-time PCR plus IS2606 conventional...
2019
Introduction: Buruli ulcer (BU) is a serious skin disease caused by Mycobacterium ulcerans. According to WHO, 70% of BU cases should be confirmed by the PCR-IS2404 gene amplification. The objective of this study is to validate a real-time PCR for the detection of M. ulcerans in clinical and environmental samples. Methodology: A total of 70 clinical samples, 10 M. ulcerans strains and 15 environmental samples were tested by Ziehl-Neelsen staining technique, conventional PCR, real time qPCR and culture. The proportion of positive cases of M. ulcerans detection between the different tests was compared by the Chi-square test. The difference was statistically significant for a P-value ≤ 0.05. Results: Out of 33/80 samples were cultured positive (41.25%) to M. ulcerans, 41/80 ZN staining were positive (51.25%) to AFB at the microscopy, 55/80 (68.75%) and 64/80 samples (80%) were positive to the IS2404 insertion sequence from conventional PCR and qPCR respectively. Both PCR techniques show...