Paradoxical Hypersusceptibility of Drug-resistant Mycobacteriumtuberculosis to β-lactam Antibiotics

• Paradoxical hypersusceptibility is observed drug susceptibility despite innate resistance in the wild type state.
• Many MDR and XDR M. tuberculosis strains are susceptible to amoxicillin/clavulanate.
• Whole-genome sequencing identified mutations associated with paradoxical hypersusceptibility.
• An expanded role for β-lactams in drug-resistant M. tuberculosis is supported.The global increase in drug-resistant tuberculosis has prompted a search for alternative therapies, including repurposing existing antibiotics. β-lactam antibiotics are safe drugs, however, they have previously been thought to be of limited use for tuberculosis due to innate resistance to this drug class. In this study, the authors found many drug-resistant tuberculosis isolates from South Africa to be susceptible to a β-lactam and β-lactamase combination, amoxicillin/clavulanate. With the use of comparative genomics, multiple genetic mutations were identified to be associated with this hypersusceptible phenotype. These findings support an expanded role of β-lactam/β-lactamase inhibitor combinations for treatment of drug-resistant TB.

Mycobacterium tuberculosis (M. tuberculosis) is considered innately resistant to β-lactam antibiotics. However, there is evidence that susceptibility to β-lactam antibiotics in combination with β–lactamase inhibitors is variable among clinical isolates, and these may present therapeutic options for drug-resistant cases. Here we report our investigation of susceptibility to β-lactam/β–lactamase inhibitor combinations among clinical isolates of M. tuberculosis, and the use of comparative genomics to understand the observed heterogeneity in susceptibility. Eighty-nine South African clinical isolates of varying first and second-line drug susceptibility patterns and two reference strains of M. tuberculosis underwent minimum inhibitory concentration (MIC) determination to two β-lactams: amoxicillin and meropenem, both alone and in combination with clavulanate, a β–lactamase inhibitor. 41/91 (45%) of tested isolates were found to be hypersusceptible to amoxicillin/clavulanate relative to reference strains, including 14/24 (58%) of multiple drug-resistant (MDR) and 22/38 (58%) of extensively drug-resistant (XDR) isolates. Genome-wide polymorphisms identified using whole-genome sequencing were used in a phylogenetically-aware linear mixed model to identify polymorphisms associated with amoxicillin/clavulanate susceptibility. Susceptibility to amoxicillin/clavulanate was over-represented among isolates within a specific clade (LAM4), in particular among XDR strains. Twelve sets of polymorphisms were identified as putative markers of amoxicillin/clavulanate susceptibility, five of which were confined solely to LAM4. Within the LAM4 clade, ‘paradoxical hypersusceptibility’ to amoxicillin/clavulanate has evolved in parallel to first and second-line drug resistance. Given the high prevalence of LAM4 among XDR TB in South Africa, our data support an expanded role for β-lactam/β-lactamase inhibitor combinations for treatment of drug-resistant M. tuberculosis.