Draft genome sequences of 12 Escherichia coli co-isolated with Enterococcus spp. from dogs with polybacterial bacteriuria at a veterinary hospital

ABSTRACT Escherichia coli are frequently co-isolated with Enterococcus spp. from urine cultures of dogs with urinary tract infections (UTIs). Uropathogenic E. coli (UPEC) are augmented by Enterococcus in polymicrobial UTIs. We report the draft genome sequences of 12 UPEC co-isolated with Enterococcus spp. from canine urinary tract infections.

KEYWORDS E. coli, UPEC, canine urinary tract infection, polymicrobial infections E scherichia coli are the most common organisms isolated from urine cultures of dogs with bacteriuria, and uropathogenic E. coli (UPEC) are implicated in cases of E. coli bacteriuria that are concurrent with clinical signs of urinary tract infections (UTIs) (1). Interestingly, UPEC are commonly co-isolated with Enterococcus spp. in dogs with polymicrobial bacteriuria (2). We have found that dogs with polymicrobial bacteriuria consisting of E. coli and Enterococcus spp. had worse clinical outcomes compared to dogs with single-species bacteriuria (3). Furthermore, dogs are a reservoir for these uropatho gens that have the potential to be zoonotically transmitted (4). Published genomes of E. coli isolated from diagnosed polymicrobial UTIs are limited despite their clinical and epidemiological significance.
Twelve E. coli isolates from urine cultures of dogs diagnosed with polymicrobial E. coli and enterococcal UTIs were sequenced. All E. coli strains were obtained from diagnostic aerobic cultures of aseptically collected urine samples on trypticase soy agar with 5% sheep blood that contained mixed growth of only E. coli and Enterococcus from 2016 to 2020 at the NC State University Veterinary Hospital in Raleigh, NC (3). Sequenced E. coli and the co-isolated Enterococcus strains were confirmed to the species level with matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry using standard methods (3). Institutional ethics approvals were not required for this work.
E. coli genomic DNA was isolated from overnight cultures grown on trypticase soy agar with 5% sheep blood. The QIAGEN DNeasy PowerLyzer Microbial Kit (QIAGEN, Germantown, MD, USA) was used for DNA isolation. A Qubit 4.0 Fluorometer and the dsDNA high-sensitivity assay kit (ThermoFisher Scientific, Waltham, MA, USA) were utilized to assess DNA quality and quantity. The Nextera DNA Flex Library Prep Kit (Illumina, San Diego, CA, USA) was used to prepare pooled libraries with an insert size of ~350 bp. Sequencing was completed on the Illumina MiSeq platform using a MiSeq v2 Reagent Kit (500 cycles) with paired-end 150-nucleotide reads and V2 chemistry according to the manufacturer's instructions. MicroRunQC version 3 workflow within GalaxyTrakr (5)   Enterococcus was the only other bacterial species co-isolated with sequenced E. coli strains and was identified at the species level with MALDI-TOF mass spectrometry using standard methods (3). b Contigs were split into scaffolds prior to annotation.
c Determined by the National Center for Biotechnology Information (NCBI) Prokaryotic Genome Annotation Pipeline (PGAP) (6).
parameters. Sequences were then annotated using the National Center for Biotechnol ogy Information (NCBI) Prokaryotic Genome Annotation Pipeline (PGAP) version 6.3 (6). E. coli genome coverage ranged from 69 to 220.1× (average: 118.8×), and contig N 50 values ranged from 57,645 to 392,281 bp (average: 229,541 bp). Assembled genomes were uploaded to the Center for Genomic Epidemiology virulence finder database version 2.0.3 (7-9) to detect the UPEC-associated fimH gene (10) in all strains using a 95% identity threshold and a minimum gene coverage of 60%. Subsequent fimH typing was completed using FimTyper for E. coli version 1.0 (11). Unless otherwise stated, default parameters were used for all sequence processing and analysis software. Isolate details, sequencing and assembly statistics, and GenBank accession numbers for raw reads and draft genome assemblies are presented in Table 1.