A combined effort of 11 laboratories in the WHO African region to improve quality of Buruli ulcer PCR diagnosis: The “BU-LABNET”

Abstract

Buruli ulcer is one of the 20 neglected tropical diseases in the world. This necrotizing hypodermitis is a chronic debilitating disease caused by an environmental Mycobacterium ulcerans. At least 33 countries with tropical, subtropical and temperate climates have reported Buruli ulcer in African countries, South America and Western Pacific regions. Majority of cases are spread across West and Central Africa. The mode of transmission is unclear, hindering the implementation of adequate prevention for the population. Currently, early diagnosis and treatment are crucial to minimizing morbidity, costs and preventing long-term disability. Biological confirmation of clinical diagnosis of Buruli ulcer is essential before starting chemotherapy. Indeed, differential diagnosis are numerous and Buruli ulcer has varying clinical presentations. Up to now, the gold standard biological confirmation is the quantitative PCR, targeting the insertion sequence IS2404 of M. ulcerans performed on cutaneous samples. Due to the low PCR confirmation rate in endemic African countries (under 30% in 2018) for numerous identified reasons within this article, 11 laboratories decided to combine their efforts to create the network “BU-LABNET” in 2019. The first step of the network was to harmonize the procedures and ship specific reagents to each laboratory. With this system in place, implementation of these procedures for testing and follow-up was easy and the laboratories were able to carry out their first quality control with a very high success rate. It is now time to integrate other neglected tropical diseases to this platform, such as yaws or leprosy.

An Overview of 10 Years of Activity of a Molecular Laboratory for Buruli Ulcer Diagnosis at a Field Hospital in Benin

“…Between 2012 and 2022, the CDTLUB laboratory received and analyzed diverse samples consisting of swabs, FNA, and skin biopsy specimens. The laboratory received dried swabs and biopsy specimens or FNA resuspended in water, always transported in an icebox, as recommended by the Standard Operating Protocols of the BU-LabNet ( 5 ). Each sample underwent Ziehl-Neelsen staining and qPCR for the IS2404 sequence.…”

“…DNA is extracted using the Genolyse kit (Hain LifeScience GmbH, Nehren, Germany), which uses a similar method to the previous one based on an alkaline lysis and heat. Also, quantification is performed using a plasmid instead of purified M. ulcerans DNA ( 5 ).…”

“…Two methods are currently used in the field to confirm a clinical diagnosis of BU: Ziehl-Neelsen staining to detect acid-fast bacilli and qPCR for the detection of pathogen-specific DNA ( 5 – 8 ). Quantitative PCR for the IS2404 sequence has proven the most sensitive and specific of the established tests for detecting M. ulcerans , and is therefore currently considered the gold standard for the confirmation of clinical diagnosis ( 5 , 9 , 10 ). The WHO recommends PCR confirmation before the start of treatment to rule out differential diagnoses because this approach can facilitate identification of patients with non-BU chronic ulcers ( 8 ).…”

“…The results obtained were good, and, since 2014, the qPCR has been performed at the CDTLUB laboratory in Benin only. Since its creation, this laboratory has participated in different available external quality assessment programs ( 5 , 14 ). Here, we provide a report of the first 10 years of M. ulcerans diagnostic activities at the CDTLUB laboratory, together with our drawn conclusions and recommendations.…”

Unlocking Opportunities for Mycobacterium leprae and Mycobacterium ulcerans

“…Dedicated research efforts are required in this neglected mycobacterial infection domain, or else a comparable situation with newly emerging M. leprae and M. ulcerans drug-resistant strains may occur sooner rather than later. This emphasizes the urgency of taking immediate action to halt the spread of these drug-resistant strains and, in particular, to support The Global Leprosy Strategy 2021–2030 “Towards zero leprosy”, which is being carried out with the goal of the targeted reduction of new patients, and the BU_LABNET initiatives, which have already been taken up to improve the quality of BU PCR diagnosis by 11 laboratories in the WHO African region; however, a substantial number of infections are still occurring. BCG vaccination has huge implications for mycobacterial disease research moving forward and public health, while the clinical trial of LepVax indicates that it is safe and immunogenic in healthy subjects, which is very encouraging for its clinical development …”