Evolutionary Genomics and Fitness Landscapes of Beta-Lactamase Variants in Gram-Negative Clinical Pathogens Across Four Nigerian Teaching Hospitals

Beta-lactamase enzyme evolution is a primary driver of expanding antibiotic resistance in gram-negative pathogens, yet the evolutionary genomics of beta-lactamase variants circulating in Nigerian clinical environments have not been characterised through combined structural, functional, and phylogenomic approaches. This dissertation investigates the evolutionary genomics and fitness landscapes of beta-lactamase variants in clinical gram-negative pathogens at teaching hospitals in Lagos (South West), Abuja (North Central), Calabar (South South), and Gombe (North East). A genomic collection of 240 gram-negative clinical isolates, predominantly Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae, underwent Illumina and MinION hybrid whole genome sequencing. Beta-lactamase variant cataloguing used ResFinder and CARD databases. Protein structural modelling of novel or divergent variants was performed using AlphaFold2 and molecular docking simulations with ceftazidime, cefotaxime, and meropenem substrates. Fitness cost of resistance gene carriage was quantified in isogenic competition experiments. One hundred and twelve distinct beta-lactamase enzyme variants were identified, including three novel CTX-M variants not deposited in existing databases and a novel NDM-type variant exhibiting broadened substrate spectrum by structural modelling. Phylogenomic analysis assigns the novel NDM variant to a distinct evolutionary clade originating within the North East hospital network. Fitness experiments show that novel CTX-M variants impose minimal growth rate penalties, predicting strong fixation potential in clinical environments without antibiotic pressure. Keywords: beta-lactamase, evolutionary genomics, fitness landscape, gram-negative pathogens, Nigeria