Case Report of Rhodococcus erythropolis Contamination in Buruli Ulcer Lesions: Effects on Mycobacterium ulcerans Diagnosis.

Abstract

Introduction: We describe cases of contamination of Mycobacterium ulcerans infections (Buruli ulcer) with Rhodococcus erythropolis, a bacterium of environmental origin that is rarely associated with human infection.

Case presentation: The infectious pathogen of Buruli ulcer, Mycobacterium ulcerans, was detected and cultured in vitro from two lesion swabs taken from clinically described Buruli ulcer-like patients (4 and 34 years old). Infection by M. ulcerans was confirmed using the WHO-recommended IS2404/IPC-qPCR multiplex analysis of DNA extracts from patient samples, which were subsequently categorized as positive. PCR-positive samples were incubated in Löwenstein-Jensen (LJ) culture media, and colony phenotypes characteristic of M. ulcerans were observed 14 days postincubation. However, analysis of culture suspensions by qPCR revealed no M. ulcerans DNA, while Ziehl-Neelsen (ZN) staining showed phenotypes closely related to acid-fast bacilli (AFB) of M. ulcerans. Detected AFBs were not clustered after ZN staining. Further microbial identification and characterization by MALDI-TOF MS and Gram staining revealed the presence of Rhodococcus erythropolis. The identification was confirmed by whole-genome sequencing (WGS) to establish the genomic link between this originally called Mycobacterium erythropolis and M. ulcerans. Analysis of short reads from WGS confirmed the organism as R. erythropolis. When employing the M. ulcerans Agy99 reference chromosome, comparative analysis of whole-genome sequences revealed little genomic relatedness between the two organisms, with an average genome coverage of 5.72%.

Conclusion: The study reports the first contamination cases of M. ulcerans-infected lesions with R. erythropolis. Although R. erythropolis did not interfere in the detection specificity of M. ulcerans by IS2404/IPC-qPCR, it completely inhibited M. ulcerans growth in recommended LJ culture media, complicating routine biological diagnosis by culture and microscopy. Hence, Buruli ulcer is likely to be underdiagnosed due to lesion contamination by R. erythropolis and difficulties in M. ulcerans identification in the routine clinical diagnosis procedure.