Genome-wide discovery of epistatic loci affecting antibiotic resistance using evolutionary couplings

In theory, whole genome sequencing should allow the discover of interdependent loci that cause antibiotic resistance. In practice, however, identifying this epistasis remains a challenge as the vast number of possible interactions erodes statistical power. To solve this problem, we infer the most strongly coupled loci using a method that has been successfully used to identify epistatic residues in proteins and here applied to whole genomes. Our method reduces the number of tests required for an epistatic genome-wide association study and increases the likelihood of identifying causal epistasis. We discover 45 loci and 291 epistatic pairs that influence the dose needed to inhibit growth for five different antibiotics on 1102 isolates of Neisseria gonorrhoeae, confirmed in an independent dataset of 495 isolates. Many of the epistatic pairs contain at least one locus or gene previously associated with antibiotic resistance, including interactions between gyrA and parC associated with ciprofloxacin resistance and between penA and murE with cefixime resistance. Our work provides a systematic identification of epistasis pairs in N. gonorrhoeae resistance and a generalizable method for epistatic genome-wide association studies.