Here, we report the draft genome sequences of 36 Enterococcus faecalis and 7 Enterococcus faecium isolates recovered from a beef processing facility and retail ground beef. The beef processing facility samples were collected from beef carcasses, conveyor belts, and ground product.
ABSTRACT
Here, we report the draft genome sequences of 36 Enterococcus faecalis and 7 Enterococcus faecium isolates recovered from a beef processing facility and retail ground beef. The beef processing facility samples were collected from beef carcasses, conveyor belts, and ground product.
ANNOUNCEMENT
Enterococcus faecalis and Enterococcus faecium are commensal microorganisms present in the gastrointestinal tract of both humans and cattle; therefore, enterococci are often used as an indicator of fecal contamination. We collected samples from four different locations in a commercial beef processing facility in Alberta, Canada, over an 18-month period. Enterococcus spp. were collected from a randomly selected 10- by 10-cm area on carcasses after hide removal (H) and final washing (W), as well as from conveyer belts (C) using a 2- by 2-cm sterile gauze swab (n = 150 each). Ground product (n = 150) and retail ground beef (n = 60) samples (25 g each) were also collected during the same time period. The swabs and ground samples were homogenized and preenriched in a Stomacher 400 circulator (Seward, Norfolk, UK) at 260 rpm for 2 min in 10 ml or 225 ml of buffered peptone water, respectively, and incubated overnight at 37°C. A 1-ml aliquot of this mixture was then added to 9 ml of Enterococcosel broth (BD, Mississauga, Ontario, Canada) and incubated overnight at 37°C to enrich for Enterococcus spp. Enterococcosel broth tubes displaying evidence of esculin hydrolysis (black) were streaked onto Enterococcosel agar and incubated at 37°C. After 48 h, the plates were examined for colonies with black zones, which is indicative of esculin hydrolysis. The groES-EL spacer region of presumptive enterococcal colonies was amplified using the EntES-211-233-F and Ent-EL-74-95-R primers (1), and the product was sequenced using an ABI Prism 3130xl genetic analyzer (Thermo Fisher Scientific, Inc., Mississauga, Ontario, Canada) for confirmation and species identification. From the confirmed Enterococcus spp., 36 E. faecalis and 7 E. faecium isolates were selected for whole-genome sequencing.
Briefly, genomic DNA was extracted using the DNeasy blood and tissue kit (Qiagen, Mississauga, Ontario, Canada) with the modification that cells were incubated with agitation (150 rpm) for 45 min at 37°C in 280 μl of lysis buffer (20 mM Tris-HCl [pH 8.0], 2 mM sodium EDTA, 1.2% Triton X-100, and 20 mg/ml lysozyme; Sigma-Aldrich Canada, Toronto, Ontario, Canada). The Nextera XT DNA library preparation kit (Illumina, Inc., San Diego, CA, USA) was used to prepare sequencing libraries that were sequenced on a MiSeq instrument (Illumina, Inc.) with the MiSeq reagent kit v3 (Illumina, Inc.; 600 cycles) as per the manufacturer’s instructions.
FastQC v0.11.5 (2) was used to assess read quality before (678,235 ± 23,253 [standard error of the mean {SEM}] reads per isolate) and after (596,916 ± 21,456 reads per isolate) quality filtering. Sequencing adapters, reads with a quality score of less than 15 over a 4-bp sliding window, and reads that were less than 50 bp in length were removed with Trimmomatic v0.38 (3). The paired-end reads were assembled with SPAdes v3.11.1 (4), with the default parameters in the “careful” mode, and the quality of the assemblies was determined using QUAST v5.0.1 (5). Contigs less than 500 bp in length were removed prior to confirming the taxonomy of each assembly with Kraken 2 v2.0.7-beta and the minikraken2 database v2 (6). The assemblies were then annotated with Prokka v1.13.3 (7) using the default parameters. Multilocus sequence typing (MLST) was done using the Enterococcus faecalis MLST website (https://pubmlst.org/efaecalis/) (8) and the Enterococcus faecium MLST website (https://pubmlst.org/efaecium/) (9). The assembly statistics, GenBank and SRA accession numbers, and MLST results for each isolate are presented in Table 1.
TABLE 1.
Assembly statistics for Enterococcus faecalis and Enterococcus faecium isolates from a beef processing facility and retail ground beefa
Isolate name | Species | GenBank accession no. | SRA accession no. | No. of contigs | No. of reads | Genome size (bp) | N50 value (bp) | Coverage (×) | No. of coding sequences | G+C content (%) | MLST |
---|---|---|---|---|---|---|---|---|---|---|---|
C112 | E. faecalis | GCA_006541215 | SRR9321129 | 26 | 722,884 | 2,988,147 | 605,310 | 73 | 2,886 | 37.43 | Unknown |
C138 | E. faecalis | GCA_006541395 | SRR9321128 | 30 | 899,074 | 2,695,188 | 213,629 | 100 | 2,557 | 37.59 | 228 |
C144 | E. faecalis | GCA_006541075 | SRR9321127 | 34 | 689,845 | 2,697,493 | 222,684 | 77 | 2,519 | 37.58 | 228 |
C146 | E. faecalis | GCA_006541305 | SRR9321126 | 28 | 502,816 | 2,697,692 | 351,123 | 56 | 2,567 | 37.59 | 228 |
G109 | E. faecalis | GCA_006541225 | SRR9321133 | 65 | 463,324 | 2,853,332 | 140,707 | 49 | 2,739 | 37.57 | Unknown |
G127E | E. faecalis | GCA_006541295 | SRR9321132 | 105 | 633,257 | 3,053,018 | 80,851 | 62 | 2,997 | 37.48 | Unknown |
G138E | E. faecalis | GCA_006541335 | SRR9321131 | 119 | 564,994 | 3,016,964 | 86,924 | 56 | 2,917 | 37.45 | 21 |
G149 | E. faecalis | GCA_006541345 | SRR9321130 | 125 | 446,360 | 2,994,467 | 59,749 | 45 | 2,854 | 37.42 | 40 |
G42 | E. faecalis | GCA_006541405 | SRR9321125 | 97 | 630,933 | 2,872,254 | 73,537 | 66 | 2,712 | 37.56 | 21 |
G69E | E. faecalis | GCA_006541355 | SRR9321124 | 17 | 529,454 | 2,959,381 | 645,373 | 54 | 2,784 | 37.51 | 202 |
G81 | E. faecalis | GCA_006541905 | SRR9321137 | 59 | 859,935 | 2,835,934 | 144,986 | 91 | 2,697 | 37.55 | Unknown |
G85 | E. faecalis | GCA_006541465 | SRR9321136 | 64 | 640,416 | 2,876,123 | 124,461 | 67 | 2,732 | 37.52 | 76 |
H102 | E. faecalis | GCA_006541755 | SRR9321139 | 106 | 416,566 | 2,830,742 | 73,730 | 44 | 2,747 | 37.6 | 147 |
H112E | E. faecium | GCA_006541175 | SRR9321138 | 42 | 512,378 | 2,655,194 | 141,400 | 58 | 2,519 | 38.05 | 212 |
H134E | E. faecium | GCA_006541535 | SRR9321141 | 66 | 535,680 | 2,507,908 | 101,754 | 64 | 2,365 | 38.07 | 29 |
H136 | E. faecalis | GCA_006541265 | SRR9321140 | 22 | 883,588 | 3,052,803 | 378,930 | 87 | 3,001 | 37.34 | Unknown |
H22 | E. faecalis | GCA_006541095 | SRR9321143 | 35 | 657,288 | 2,786,536 | 250,863 | 71 | 2,564 | 37.49 | 16 |
H4 | E. faecalis | GCA_006541805 | SRR9321142 | 59 | 641,369 | 2,996,880 | 146,743 | 64 | 2,860 | 37.42 | 40 |
H44 | E. faecalis | GCA_006541185 | SRR9321135 | 46 | 649,040 | 2,816,920 | 228,593 | 69 | 2,629 | 37.57 | 76 |
H74 | E. faecalis | GCA_006541115 | SRR9321134 | 38 | 769,279 | 2,733,857 | 166,527 | 84 | 2,575 | 37.57 | Unknown |
H96E | E. faecalis | GCA_006541255 | SRR9321160 | 32 | 664,950 | 2,847,191 | 191,075 | 70 | 2,644 | 37.53 | 708 |
R2 | E. faecium | GCA_006541615 | SRR9321161 | 145 | 784,155 | 2,743,995 | 46,108 | 86 | 2,682 | 38.24 | 76 |
R20 | E. faecalis | GCA_006541155 | SRR9321162 | 163 | 623,226 | 2,985,199 | 78,020 | 63 | 2,859 | 37.52 | 260 |
R26E | E. faecium | GCA_006541645 | SRR9321163 | 237 | 261,875 | 2,688,512 | 23,732 | 29 | 2,594 | 38.38 | 76 |
R29 | E. faecalis | GCA_006541545 | SRR9321156 | 45 | 468,536 | 2,911,452 | 206,037 | 48 | 2,785 | 37.47 | 260 |
R30 | E. faecalis | GCA_006541705 | SRR9321157 | 49 | 505,869 | 2,647,103 | 84,312 | 57 | 2,488 | 37.7 | 228 |
R37 | E. faecalis | GCA_006541775 | SRR9321158 | 70 | 482,350 | 2,929,864 | 135,712 | 49 | 2,769 | 37.39 | 260 |
R43E | E. faecalis | GCA_006541085 | SRR9321159 | 73 | 591,636 | 2,833,916 | 87,778 | 63 | 2,734 | 37.49 | Unknown |
R49 | E. faecalis | GCA_006541715 | SRR9321154 | 36 | 390,483 | 3,029,112 | 153,103 | 39 | 2,921 | 37.37 | 192 |
R4E | E. faecium | GCA_006541685 | SRR9321155 | 100 | 667,395 | 2,738,432 | 75,527 | 73 | 2,683 | 38.24 | Unknown |
R50 | E. faecalis | GCA_006541785 | SRR9321147 | 113 | 519,617 | 2,976,798 | 77,751 | 52 | 2,863 | 37.46 | 21 |
R51 | E. faecalis | GCA_006541855 | SRR9321146 | 62 | 463,047 | 3,005,276 | 121,849 | 46 | 2,957 | 37.45 | 84 |
R52 | E. faecalis | GCA_006541795 | SRR9321145 | 66 | 449,634 | 3,046,695 | 119,574 | 44 | 2,931 | 37.37 | 260 |
R53 | E. faecalis | GCA_006541655 | SRR9321144 | 79 | 596,560 | 3,030,982 | 119,130 | 59 | 2,966 | 37.42 | 260 |
R5E | E. faecalis | GCA_006541525 | SRR9321151 | 76 | 611,462 | 2,934,492 | 97,444 | 63 | 2,785 | 37.39 | Unknown |
R7 | E. faecalis | GCA_006541595 | SRR9321150 | 51 | 507,624 | 2,911,924 | 184,949 | 52 | 2,821 | 37.47 | 260 |
W100 | E. faecalis | GCA_006541515 | SRR9321149 | 55 | 452,646 | 2,961,448 | 130,717 | 46 | 2,832 | 37.42 | 260 |
W133 | E. faecalis | GCA_006541445 | SRR9321148 | 364 | 678,090 | 2,944,917 | 14,728 | 69 | 2,799 | 37.19 | Unknown |
W141 | E. faecium | GCA_006541625 | SRR9321153 | 82 | 626,699 | 2,761,265 | 123,573 | 68 | 2,695 | 38.22 | 76 |
W148E | E. faecium | GCA_006541485 | SRR9321152 | 96 | 501,112 | 2,758,663 | 89,896 | 54 | 2,687 | 38.23 | 76 |
W19 | E. faecalis | GCA_006541875 | SRR9321122 | 76 | 865,657 | 2,956,998 | 112,037 | 88 | 2,846 | 37.5 | Unknown |
W84 | E. faecalis | GCA_006541885 | SRR9321123 | 99 | 564,979 | 3,018,223 | 89,324 | 56 | 2,896 | 37.45 | Unknown |
W97 | E. faecalis | GCA_006541435 | SRR9321121 | 61 | 741,306 | 3,050,168 | 155,000 | 73 | 2,961 | 37.37 | 40 |
The number of coding sequences is based on Prokka annotations. Annotations in GenBank are based on the Prokaryotic Genome Annotation Pipeline (PGAP).
The genome assemblies were also screened for the presence of antimicrobial resistance (AMR) genes using BLASTn 2.6.0+ and the Comprehensive Antibiotic Resistance Database (CARD; v3.0.2) (10), with a minimum identity of 90%. The most prevalent AMR genes in the E. faecalis genomes were tet(M) (25%) and erm(B) (8%); in the E. faecium assemblies, msrC (100%), aac(6′)-II (100%), and tet(M) (29%) were most frequently detected.
Data availability.
All sequences and draft genome assemblies have been deposited in the Sequence Read Archive and GenBank, respectively, under the accession numbers listed in Table 1.
ACKNOWLEDGMENTS
This project was financially supported by the Beef Cattle Research Council and Agriculture and Agri-Food Canada.
We thank Cara Service, Scott Hrycauk, and Nicole Lassel for their valuable technical assistance.
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Associated Data
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Data Availability Statement
All sequences and draft genome assemblies have been deposited in the Sequence Read Archive and GenBank, respectively, under the accession numbers listed in Table 1.