Draft Genome Sequences of Tetracycline-Resistant Shiga Toxin-Producing Escherichia coli Isolates from Food

ABSTRACT

Shiga toxin-producing Escherichia coli (STEC) is a foodborne pathogen transmitted from animal to humans through contaminated food. Here, we report the draft genome sequences of six STEC isolates (six serotypes) from food (cheese, coriander, and pea protein pellets) in different countries; these isolates were resistant to tetracycline, with MIC values ranging from <1.5 to 256 μg/mL.

ANNOUNCEMENT

Shiga toxin-producing Escherichia coli (STEC) can cause bloody diarrhea, hemolytic uremic syndrome (HUS), and hemorrhagic and even fatal colitis in human. It causes around 2.5 million acute illnesses each year globally, and the consumption of contaminated foods (such as meat, beef, milk, fruits, and vegetables) is the primary source of STEC transmission, contributing 50% of all STEC infections (1, 2). Although E. coli O157:H7 is the most important STEC serotype posing a public health threat, non-O157:H7 STEC serotypes, such as O26, O111, O103, and O145, have frequently been involved in sporadic cases and outbreaks in the United States and other countries (3). In addition, more than 400 serotypes of non-O157:H7 STEC have been reported, and there were at least 42 STEC outbreaks in the United States from 2006 to 2021 (4, 5). Recently, STEC has been elevated to a major global public health concern because of the growing number of infections and major outbreaks with multidrug-resistant (MDR) strains.

Here, we announce six draft genome sequences of STEC isolates sourced from cheese, coriander, and pea protein pellets (Table 1). They were obtained following the U.S. Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) (6). All bacterial isolates were grown from frozen stocks, streaked onto MacConkey agar (FDA BAM media M91), and incubated overnight at 37 ± 1°C. The isolates were serotyped as O2:H1, O146:H21, NEG:+, O6:H10, O8:H30, and O103:H2 at the E. coli Reference Center (Department of Food Science) at the Pennsylvania State University (PA, USA), following the traditional agglutination-based method (7). The MICs were interpreted according to the Clinical and Laboratory Standards Institute (CLSI) (8). The MIC data revealed that these strains were resistant to tetracycline; two isolates, 872416 and 886340, were categorized as highly resistant to tetracycline, with MIC values of 64 and 256 μg/mL, respectively (Table 1).

TABLE 1.

Genome features of Shiga toxin-producing Escherichia coli isolates sequenced in this project

Isolate	Source	Serotype	MIC of tetracycline (μg/mL)	No. of reads	No. of contigs	Contig N50 (bp)	Coverage (×)	G+C content (%)	NCBI accession no.			Illumina sequencer
									BioSample	SRA	GenBank
879916	Cheddar cheese, USA	O2:H1	2	4,803,343	63	200,856	50	50.59	SAMN16454162	SRR12904588	JADDRX000000000	NextSeq 500
875221	Coriander, Mexico	O146:H21	<1.5	5,071,407	187	103,993	50	50.58	SAMN16454163	SRR12904587	JADDRW000000000	NextSeq 500
927095	Blue cheese, France	NEG:+	<1.5	4,680,969	121	114,771	50	50.57	SAMN16454164	SRR12904586	JADDRV000000000	NextSeq 500
872416	Roquefort cheese, France	O6:H10	64	4,955,756	257	61,625	50	50.68	SAMN16454165	SRR12904585	JADDRU000000000	NextSeq 500
886340	Roquefort cheese, France	O8:H30	256	5,349,033	247	65,023	50	50.55	SAMN16454166	SRR12904584	JADDRT000000000	MiSeq
932783	Pea protein pellets, Norway	O103:H2	<2	5,099,373	310	90,199	50	50.51	SAMN16454167	SRR12904583	JADDRS000000000	NextSeq 500

A single STEC colony isolated from a MacConkey agar plate was inoculated into LB (Luria-Bertani) broth (Becton, Dickinson and Company, Franklin Lakes, NJ, USA) and incubated at 37 ± 1°C overnight for genomic DNA preparation. Genomic DNA was extracted using a DNeasy blood and tissue kit (Qiagen, Valencia, CA, USA) according to the manufacturer’s instructions. DNA libraries were constructed using a Nextera XT DNA library kit (Illumina, San Diego, CA, USA). The library concentration was assessed using a high-sensitivity (HS) kit (Agilent Technologies, Santa Clara, CA, USA) with a Qubit 4.0 fluorometer (Life Technologies, Carlsbad, CA, USA). The quality of the libraries was analyzed using a 2100 Bioanalyzer instrument (Agilent Technologies). The pooled libraries for all isolates were loaded onto a flow cell and sequenced using the Illumina NextSeq 500 platform (Illumina) with a NextSeq reagent kit v2.5 (2 × 75-bp paired-end reads), except for strain 886340, which was sequenced using the Illumina MiSeq platform with a MiSeq reagent kit v2 (2 × 250-bp paired-end reads) (9). The trimming and assembly were performed using the CLC Genomics Workbench v11 (Qiagen, Germantown, MD, USA), and the quality of the genome assembly and the validity of the final genome were assessed using the quality control tool (QC report) under CLC Workbench. The draft genomes were annotated initially using Pathosystem Resources Integration Center (PATRIC) software v3.6.12, and the data were submitted to the NCBI for final annotation using the Prokaryotic Genome Annotation Pipeline (PGAP) under the accession numbers shown in Table 1 (10). Default parameters were used for all software unless otherwise specified. The average G+C content of these strains was approximately about 50.5% as estimated by the PATRIC database, which was used for annotation with default parameters. The genome sizes for these six strains ranged from 4,680,969 to 5,349,033 bp, the number of contigs ranged from 63 to 310, the N₅₀ values ranged from 61,625 to 200,856 bp, and the coverages were all 50× (Table 1).

Data availability.

All sequenced genome data have been deposited at DDBJ/ENA/GenBank under BioProject accession number PRJNA545531 and BioSample accession numbers SAMN16454162, SAMN16454163, SAMN16454164, SAMN16454165, SAMN16454166, and SAMN16454167.