Draft Genome Sequences of 27 Staphylococcus aureus Strains and 3 Staphylococcus Species Strains Isolated from Bovine Intramammary Infections

Staphylococcus aureus is a major pathogen that is responsible for both clinical and subclinical contagious mastitis in dairy cows. Dairy cows with active intramammary infection often have milk yield reductions. Persistent intramammary infections are a common cause of culling in dairy herds, which results in economic losses for the dairy industry (1). Although S. aureus-associated bovine mastitis cases have decreased since the implementation of mastitis control programs, S. aureus remains a challenge for dairy farmers and veterinarians (2, 3). In bovine mastitis control programs, prevention is the best strategy for reducing the burdens in the dairy industry. Although thousands of S. aureus isolates have been sequenced and reported, relatively few are from bovine intramammary infections. The availability of S. aureus isolates from intramammary infections will facilitate understanding of the molecular basis of pathogenic S. aureus characteristics associated with bovine mastitis. The Mastitis Network maintains a culture collection of mastitis isolates from Canada (4); each strain sequenced in this project was obtained from that collection.

Here, we present the draft genome sequences of 27 S. aureus isolates and 3 Staphylococcus sp. isolates from bovine intramammary infections in Canada. The isolates were initially identified at the species level using matrix-assisted laser desorption ionization–time of flight (MALDI) mass spectrometry, as described previously (5). Each isolate was cultivated from the –80°C stock on a tryptic soy agar plate, which was incubated overnight at 37°C. A single well-isolated colony was used to inoculate tryptic soy broth, which was incubated overnight at 37°C with agitation at 200 rpm. A 1.5-ml aliquot of the liquid culture was used for DNA extraction with the DNAzol reagent (Invitrogen) and lysostaphin (Sigma-Aldrich) according to the manufacturers’ instructions. Briefly, sequencing libraries were prepared as paired-end libraries with the Nextera Flex DNA library preparation kit (Illumina, San Diego, CA) and Nextera DNA CD indexes (96 indexes, 96 samples), and the libraries were sequenced on a MiSeq benchtop sequencer (Illumina) for 301 cycles in each direction. The reads were assembled de novo into high-quality draft genomes with ProkaryoteAssembly version 0.1.6 (https://github.com/bfssi-forest-dussault/ProkaryoteAssembly). Default parameters were used throughout, with the exception of the trimming step, for which the command trimq=20 2> {stats_out} was used to trim low-quality sequences with a Q score of <20. This assembly resulted in nonoverlapping contiguous sequences being generated for each genome (Table 1). Gene predictions and annotations were performed using the National Center for Biotechnology Information (NCBI) Prokaryotic Genome Annotation Pipeline (PGAP).

This work was funded by Op+lait through a Concours Nouvelles Initiatives de Recherche grant awarded to S.D. and J.R. S.P. and D.J. received scholarship funding through the Op+lait Complements de Bourse program, as well as through the NSERC CREATE in Milk Quality program.