Using Whole Genome Sequencing to Track Colibacillosis on Saskatchewan Broiler Flocks

Abstract

Colibacillosis is a systemic infection caused by Escherichia coli resulting in significant morbidity and mortality in broiler flocks worldwide. Little is known about the group of E. coli that cause colibacillosis, collectively termed avian pathogenic E. coli (APEC). My MSc research focused on determining how APEC differ from resident E. coli that live in the chicken gut but do not cause disease. I hypothesized that systemic and cecal E. coli are genetically distinct, and E. coli that cause colibacillosis are virulent outbreak strains. My objectives were to isolate E. coli from Saskatchewan broilers, sequence their genomes using Nanopore and Illumina technology, and screen them for virulence, antimicrobial resistance, and disinfectant resistance. I developed a pipeline to isolate and sequence E. coli from Saskatchewan colibacillosis outbreaks, selecting isolates based on outbreak, disease status, and biofilm profiles. I sequenced 96 E. coli isolates, consisting of 58 from diseased broilers with confirmed colibacillosis (systemic E. coli), and 38 from the cecal contents of healthy broilers in the same flocks (cecal E. coli). Our initial experiments were optimized for whole genome assembly and excluded DNA fragments under 500bp; therefore, we likely missed plasmids present in E. coli isolates. I tested six plasmid kits and two sequencing protocols to develop a methodology to capture missed plasmids in avian E. coli isolates and successfully identified new plasmids in both types of isolates. Systemic E. coli were more drug-resistant than cecal E. coli against a panel of 27 antimicrobial agents and possessed significantly more plasmids than cecal E. coli. plasmids contained multiple virulence and antimicrobial resistance genes that may contribute to disease. Since biofilms can provide protection from antibiotics and disinfectants, I quantified biofilm formation in three different medias. Systemic isolates were significantly more likely to form biofilms in rich media, but there was no correlation between biofilm formation and antimicrobial resistance. My characterization led us to conclude that systemic and cecal E. coli represent two different populations of strains. This will need to be confirmed with the analysis of more isolates. Characterization of avian pathogenic E. coli will help us understand how these isolates cause disease.