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Mycolactone, the amphiphilic macrolide toxin secreted by Mycobacterium ulcerans, plays a significant role in the pathology and manifestations of Buruli ulcer (BU). Consequently, it follows that the toxin is a suitable target for the development of diagnostics and therapeutics for this disease. Yet, several challenges have deterred such development. For one, the lipophilic nature of the toxin makes it difficult to handle and store and contributes to variability associated with laboratory experimentation and purification yields. In this manuscript, we have attempted to incorporate our understanding of the lipophilicity of mycolactone in order to define the optimal methods for the storage, handling, and purification of this toxin. We present a systematic correlation of variability associated with measurement techniques (thin-layer chromatography (TLC), mass spectrometry (MS), and UV-Vis spectrometry), storage conditions, choice of solvents, as well as the impact of each of these on toxin function as assessed by cellular cytotoxicity. We also compared natural mycolactone extracted from bacterial culture with synthesized toxins in laboratory (solvents, buffers) and physiologically relevant (serum) matrices. Our results point to the greater stability of mycolactone in organic, as well as detergent-containing, solvents, regardless of the container material (plastic, glass, or silanized tubes). They also highlight the presence of toxin in samples that may be undetectable by any one technique, suggesting that each detection approach captures different configurations of the molecule with varying specificity and sensitivity. Most importantly, our results demonstrate for the very first time that amphiphilic mycolactone associates with host lipoproteins in serum, and that this association will likely impact our ability to study, diagnose, and treat Buruli ulcers in patients.
Jessica Z. Kubicek-Sutherland; Dung M. Vu; Aaron S. Anderson; Timothy C. Sanchez; Paul J. Converse; Ricardo Martí-Arbona; Eric L. Nuermberger; Basil I. Swanson; Harshini Mukundan. Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone. Toxins 2019, 11, 202 .
AMA StyleJessica Z. Kubicek-Sutherland, Dung M. Vu, Aaron S. Anderson, Timothy C. Sanchez, Paul J. Converse, Ricardo Martí-Arbona, Eric L. Nuermberger, Basil I. Swanson, Harshini Mukundan. Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone. Toxins. 2019; 11 (4):202.
Chicago/Turabian StyleJessica Z. Kubicek-Sutherland; Dung M. Vu; Aaron S. Anderson; Timothy C. Sanchez; Paul J. Converse; Ricardo Martí-Arbona; Eric L. Nuermberger; Basil I. Swanson; Harshini Mukundan. 2019. "Understanding the Significance of Biochemistry in the Storage, Handling, Purification, and Sampling of Amphiphilic Mycolactone." Toxins 11, no. 4: 202.
Novel regimens combining bedaquiline and pretomanid with either linezolid (BPaL regimen) or moxifloxacin and pyrazinamide (BPaMZ regimen) shorten the treatment duration needed to cure TB in BALB/c mice compared to the first-line regimen and have yielded promising results in initial clinical trials. However, the independent contribution of the investigational new drug pretomanid to the efficacy of BPaMZ has not been examined and its contribution to BPaL has been examined only over the first 2 months of treatment. In the present study, the addition of pretomanid to BL increased bactericidal activity, prevented emergence of bedaquiline resistance, and shortened the duration needed to prevent relapse with drug-susceptible isolates by at least 2 months in BALB/c mice. Addition of pretomanid to BMZ resulted in a 1 log10 greater CFU reduction after 1 month of treatment and/or reduced the number of mice relapsing in each of 2 experiments in BALB/c mice and in immunocompromised nude mice. Bedaquiline-resistant isolates were found at relapse in only one BMZ-treated nude mouse. Treatment of infection with a pyrazinamide-resistant mutant in BALB/c mice with BPaMZ prevented selection of bedaquiline-resistant mutants and reduced the proportion of mice relapsing compared to BMZ alone. Among severely ill C3HeB/FeJ mice with caseous pneumonia and cavitation, BPaMZ increased median survival (≥60 vs. 21 days) and reduced median lung CFU by 2.4 log10 at 1 month compared to BMZ. In conclusion, in 3 different mouse models, pretomanid contributed significantly to the efficacy of the BPaMZ and BPaL regimens, including restricting the selection of bedaquiline-resistant mutants.
Jian Xu; Si-Yang Li; Deepak V. Almeida; Rokeya Tasneen; Kala Barnes-Boyle; Paul J. Converse; Anna M. Upton; Khisimuzi Mdluli; Nader Fotouhi; Eric L. Nuermberger. Contribution of pretomanid to novel regimens containing bedaquiline with either linezolid or moxifloxacin and pyrazinamide in murine models of tuberculosis. 2019, 514661 .
AMA StyleJian Xu, Si-Yang Li, Deepak V. Almeida, Rokeya Tasneen, Kala Barnes-Boyle, Paul J. Converse, Anna M. Upton, Khisimuzi Mdluli, Nader Fotouhi, Eric L. Nuermberger. Contribution of pretomanid to novel regimens containing bedaquiline with either linezolid or moxifloxacin and pyrazinamide in murine models of tuberculosis. . 2019; ():514661.
Chicago/Turabian StyleJian Xu; Si-Yang Li; Deepak V. Almeida; Rokeya Tasneen; Kala Barnes-Boyle; Paul J. Converse; Anna M. Upton; Khisimuzi Mdluli; Nader Fotouhi; Eric L. Nuermberger. 2019. "Contribution of pretomanid to novel regimens containing bedaquiline with either linezolid or moxifloxacin and pyrazinamide in murine models of tuberculosis." , no. : 514661.
Until 2004, the skin disease known as Buruli ulcer, caused by Mycobacterium ulcerans, could only be treated by surgery and skin grafting. Although this worked reasonably well on early lesions typically found in patients in Australia, the strategy was usually impractical on large lesions resulting from diagnostic delay in patients in rural West Africa. Based on promising preclinical studies, treatment trials in West Africa have shown that a combination of rifampin and streptomycin administered daily for 8 weeks can kill M. ulcerans bacilli, arrest the disease, and promote healing without relapse or reduce the extent of surgical excision. Improved treatment options are the focus of research that has increased tremendously since the WHO began its Global Buruli Ulcer Initiative in 1998.
Paul J Converse; Eric L Nuermberger; Deepak V Almeida; Jacques H Grosset. TreatingMycobacterium ulceransdisease (Buruli ulcer): from surgery to antibiotics, is the pill mightier than the knife? Future Microbiology 2011, 6, 1185 -1198.
AMA StylePaul J Converse, Eric L Nuermberger, Deepak V Almeida, Jacques H Grosset. TreatingMycobacterium ulceransdisease (Buruli ulcer): from surgery to antibiotics, is the pill mightier than the knife? Future Microbiology. 2011; 6 (10):1185-1198.
Chicago/Turabian StylePaul J Converse; Eric L Nuermberger; Deepak V Almeida; Jacques H Grosset. 2011. "TreatingMycobacterium ulceransdisease (Buruli ulcer): from surgery to antibiotics, is the pill mightier than the knife?" Future Microbiology 6, no. 10: 1185-1198.