Seven Salmonella enterica subsp. enterica isolates were identified as carrying the mcr-1 gene, by using a real-time fluorescence quantitative PCR method, from a total of 2,558 isolates which were cultured from various food origins in China between 2011 and 2016. Few complete genomes of Salmonella strains harboring the mcr-1 gene have been reported to date, so we report here the complete genome and plasmid sequences of all of these isolates to provide useful references for understanding the prevalence of foodborne Salmonella enterica subsp. enterica isolates carrying mcr-1.
ABSTRACT
Seven Salmonella enterica subsp. enterica isolates were identified as carrying the mcr-1 gene, by using a real-time fluorescence quantitative PCR method, from a total of 2,558 isolates which were cultured from various food origins in China between 2011 and 2016. Few complete genomes of Salmonella strains harboring the mcr-1 gene have been reported to date, so we report here the complete genome and plasmid sequences of all of these isolates to provide useful references for understanding the prevalence of foodborne Salmonella enterica subsp. enterica isolates carrying mcr-1.
ANNOUNCEMENT
The rise and dissemination of multidrug-resistant (MDR) Enterobacteriaceae, especially carbapenem-resistant Enterobacteriaceae (CRE), with mechanisms such as NDM-1, KPC, and OXA-48/181 in the last few decades have led to urgent challenges in the clinical treatment of MDR or extensively drug-resistant (XDR) pathogen diseases (1, 2). In this case, despite having a side effect of nephrotoxicity, colistin is still considered a last-resort antibiotic in the clinical treatment of serious infections caused by CRE (3). However, mobile colistin resistance (MCR), referring to a plasmid-mediated gene encoding a phosphoethanolamine transferase conferring resistance to colistin, was initially reported in 2015 in China and named mcr-1 (mobile colistin resistance 1) (4). This gene, originating from different bacterial species from human, animal, and environmental samples, has been reported in more than 30 countries across six continents, dating to at least the 1980s (5). Variants of mcr-1 and other mcr genes have been identified consecutively, with the mcr-8 gene reported in Klebsiella pneumoniae as the latest one (1). While the presence of mcr-1-mediated colistin resistance was predominantly reported among Escherichia coli, K. pneumoniae, and Enterobacter spp. in China, data for Salmonella isolates are lacking, particularly for isolates of food origins (6).
Therefore, 2,558 Salmonella isolates recovered from various kinds of food in China between 2011 and 2016 were screened for the mcr-1 gene by the real-time fluorescence quantitative PCR method in our laboratory, and seven isolates among them were identified as mcr-1 positive (7). We present here the complete genome and plasmid sequences of all seven foodborne Salmonella enterica subsp. enterica isolates. These genome sequences will be of great use in providing a genetic basis for Salmonella spp. harboring the mcr-1 gene, as references to aid in comparative genomics applications, as well as for epidemiological studies on outbreak detection and surveillance of Salmonella spp. in the future.
A single colony for each strain was grown overnight on brain heart infusion (BHI) broth at 37°C, and genomic DNA was extracted using the TIANamp bacterial DNA kit (catalog no. DP302; Tiangen Biotech, Beijing, China); this was followed by preparation of a 10-kb library from 5 μg of sheared and concentrated genomic DNA using a 10-kb template library preparation and sequencing procedure with the PacBio template prep kit. Whole-genome sequencing was performed using the single-molecule real-time Pacific Biosciences (SMRT PacBio) RS II platform (Tianjin Biochip Corporation, Tianjin, China). SMRT sequencing was conducted using the C4 sequencing chemistry and P6 polymerase with 1 SMRT cell. SMRT Analysis v2.3.0, available from PacBio, was used to perform demultiplexing, base calling, quality filtering of the raw read sequences, and de novo assembly according to the RS Hierarchical Genome Assembly Process (HGAP) workflow v3.0. Subsequently, Consed software v28.0 (http://www.phrap.org/consed/consed.html) was used to manually inspect and trim duplicate ends to generate single, complete, and closed sequences for each chromosome and plasmid. The genomes assembled from the PacBio data were then error corrected using Pilon software (v1.23) with Illumina MiSeq sequencing read data, of which a library was prepared with a NEBNext Ultra DNA library prep kit for Illumina (NEB catalog no. E7370), followed by sonication fragmentation (350-bp insert), and loaded onto the Illumina HiSeq platform with a paired-end (PE) 150-bp sequencing strategy (Novogene, Beijing, China) with the HiSeq X Ten reagent kit v2.5 (Illumina, San Diego, CA). The predicted serotypes and multilocus sequencing types (MLST) were identified using the Salmonella In Silico Typing Resource (SISTR; https://lfz.corefacility.ca/sistr-app/). Plasmid replicon types or incompatibility (Inc) groups were determined using the PlasmidFinder 2.0 platform (https://cge.cbs.dtu.dk/services/PlasmidFinder-2.0/).
Data availability.
The genome and plasmid sequence data of all seven Salmonella enterica subsp. enterica isolates have been deposited in NCBI GenBank under BioProject no. PRJNA498334. Automatic annotation was done using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP). The isolate information, sequencing metrics, and genomic data of the seven Salmonella enterica subsp. enterica strains are listed in Table 1.
TABLE 1.
Chromosome and plasmid sequence accession numbers and additional information for seven Salmonella enterica subsp. enterica strains harboring the mcr-1 gene
| Strain or plasmid name | Chromosome or plasmid |
Salmonella isolate information |
Sequencing metrics |
Genomic data |
||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Serotype | MLST | Yr | Food source | No. of reads | Mean read length (bp) | Coverage (×) | BioSample accession no. | GenBank accession no. | Size (bp) | G+C content (%) | No. of coding genes, pseudogenes, and RNA genes | Plasmid replicon type (Inc group)e | ||
| CFSA122 | Chromosome | Typhimurium | 34 | 2013 | Pork | 113,771 | 8,504 | 127.12 | SAMN10279393 | CP033226 | 4,990,577 | 52.1 | 4,903, 121, 125 | NA |
| pCFSA122-1a | Plasmid (complete) | CP033224 | 181,747 | 46.7 | IncHI2A, IncHI2 | |||||||||
| pCFSA122-2 | Plasmid (complete) | CP033225 | 6,758 | 46.2 | ColRNAI | |||||||||
| CFSA244 | Chromosome | Typhimuriumc | 34 | 2014 | Pork | 55,132 | 10,327 | 71.51 | SAMN10290237 | CP033255 | 4,957,526 | 52.1 | 4,883, 110, 122 | NA |
| pCFSA244-1 | Plasmid (complete) | CP033253 | 149,567 | 45.6 | IncHI2A, IncHI2 | |||||||||
| pCFSA244-2a | Plasmid (complete) | CP033254 | 60,381 | 42.3 | IncI2 | |||||||||
| CFSA12 | Chromosome | Typhimuriumc | 34 | 2014 | Pork | 91,574 | 9,412 | 101.28 | SAMN10290244 | CP033257 | 4,991,162 | 52.1 | 4,868, 110, 125 | NA |
| pCFSA12b | Plasmid (complete) | CP033256d | 147,918 | 45.0 | IncHI2A, IncHI2 | |||||||||
| CFSA1096 | Chromosome | London | 155 | 2015 | Pork | 69,175 | 10,269 | 89.86 | SAMN10291458 | CP033348 | 4,696,663 | 52.3 | 4,637, 145, 117 | NA |
| pCFSA1096a | Plasmid (complete) | CP033347 | 297,348 | 46.7 | IncHI2A, IncHI2 | |||||||||
| CFSA231 | Chromosome | Derby | 40 | 2016 | Pork | 65,092 | 8,680 | 68.13 | SAMN10291561 | CP033350 | 4,834,516 | 52.1 | 4,519, 134, 119 | NA |
| pCFSA231a | Plasmid (complete) | CP033349 | 33,309 | 41.9 | IncX4 | |||||||||
| CFSA629 | Chromosome | Typhimurium | 34 | 2016 | Egg | 54,855 | 9,083 | 57.39 | SAMN10291586 | CP033352 | 4,999,270 | 52.1 | 4,937, 117, 125 | NA |
| pCFSA629a | Plasmid (complete) | CP033351 | 210,674 | 45.2 | IncHI2A, IncHI2 | |||||||||
| CFSA664 | Chromosome | Indiana | 17 | 2011 | Chicken | 88,121 | 8,319 | 85.69 | SAMN10292850 | CP033356 | 4,733,813 | 52.1 | 4,760, 176, 120 | NA |
| pCFSA664-1 | Plasmid (complete) | CP033353 | 255,327 | 47.9 | IncHI2A, IncHI2, IncN, IncQ1 | |||||||||
| pCFSA664-2 | Plasmid (complete) | CP033354 | 41,696 | 45.4 | IncP-1-like trfA | |||||||||
| pCFSA664-3a | Plasmid (complete) | CP033355 | 61,841 | 42.4 | IncI2 | |||||||||
Contains the mcr-1 gene.
A spontaneous mcr-1 gene deletion was observed on pCFSA12 of Salmonella enterica subsp. enterica isolate CFSA12.
Predicted to be a potential monophasic variant of S. Typhimurium with a serotype antigenic formula of 4,[5],12:i:−.
The error-corrected sequence of CP033256 by Pilon with Illumina sequencing read data was identical to the previous version, so the accession number version did not change.
NA, plasmid replicon typing was not applicable for the chromosome. pCFSA664-2 was not predicted to have an Inc group using PlasmidFinder; however, the annotation contained an IncP-1-like trfA replication gene from an also-untypeable plasmid, pYDC107_41 (GenBank accession no. CP025711), which is 97.63% identical with 95% coverage to pCFSA664-2 (8).
ACKNOWLEDGMENTS
This work was financially supported by the National Key R&D Program of China (grants 2018YFC1603904 and 2016YFD0401102) and the China Food Safety Talent Competency Development Initiative: CFSA 523 Program.
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The genome and plasmid sequence data of all seven Salmonella enterica subsp. enterica isolates have been deposited in NCBI GenBank under BioProject no. PRJNA498334. Automatic annotation was done using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP). The isolate information, sequencing metrics, and genomic data of the seven Salmonella enterica subsp. enterica strains are listed in Table 1.
TABLE 1.
Chromosome and plasmid sequence accession numbers and additional information for seven Salmonella enterica subsp. enterica strains harboring the mcr-1 gene
| Strain or plasmid name | Chromosome or plasmid |
Salmonella isolate information |
Sequencing metrics |
Genomic data |
||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Serotype | MLST | Yr | Food source | No. of reads | Mean read length (bp) | Coverage (×) | BioSample accession no. | GenBank accession no. | Size (bp) | G+C content (%) | No. of coding genes, pseudogenes, and RNA genes | Plasmid replicon type (Inc group)e | ||
| CFSA122 | Chromosome | Typhimurium | 34 | 2013 | Pork | 113,771 | 8,504 | 127.12 | SAMN10279393 | CP033226 | 4,990,577 | 52.1 | 4,903, 121, 125 | NA |
| pCFSA122-1a | Plasmid (complete) | CP033224 | 181,747 | 46.7 | IncHI2A, IncHI2 | |||||||||
| pCFSA122-2 | Plasmid (complete) | CP033225 | 6,758 | 46.2 | ColRNAI | |||||||||
| CFSA244 | Chromosome | Typhimuriumc | 34 | 2014 | Pork | 55,132 | 10,327 | 71.51 | SAMN10290237 | CP033255 | 4,957,526 | 52.1 | 4,883, 110, 122 | NA |
| pCFSA244-1 | Plasmid (complete) | CP033253 | 149,567 | 45.6 | IncHI2A, IncHI2 | |||||||||
| pCFSA244-2a | Plasmid (complete) | CP033254 | 60,381 | 42.3 | IncI2 | |||||||||
| CFSA12 | Chromosome | Typhimuriumc | 34 | 2014 | Pork | 91,574 | 9,412 | 101.28 | SAMN10290244 | CP033257 | 4,991,162 | 52.1 | 4,868, 110, 125 | NA |
| pCFSA12b | Plasmid (complete) | CP033256d | 147,918 | 45.0 | IncHI2A, IncHI2 | |||||||||
| CFSA1096 | Chromosome | London | 155 | 2015 | Pork | 69,175 | 10,269 | 89.86 | SAMN10291458 | CP033348 | 4,696,663 | 52.3 | 4,637, 145, 117 | NA |
| pCFSA1096a | Plasmid (complete) | CP033347 | 297,348 | 46.7 | IncHI2A, IncHI2 | |||||||||
| CFSA231 | Chromosome | Derby | 40 | 2016 | Pork | 65,092 | 8,680 | 68.13 | SAMN10291561 | CP033350 | 4,834,516 | 52.1 | 4,519, 134, 119 | NA |
| pCFSA231a | Plasmid (complete) | CP033349 | 33,309 | 41.9 | IncX4 | |||||||||
| CFSA629 | Chromosome | Typhimurium | 34 | 2016 | Egg | 54,855 | 9,083 | 57.39 | SAMN10291586 | CP033352 | 4,999,270 | 52.1 | 4,937, 117, 125 | NA |
| pCFSA629a | Plasmid (complete) | CP033351 | 210,674 | 45.2 | IncHI2A, IncHI2 | |||||||||
| CFSA664 | Chromosome | Indiana | 17 | 2011 | Chicken | 88,121 | 8,319 | 85.69 | SAMN10292850 | CP033356 | 4,733,813 | 52.1 | 4,760, 176, 120 | NA |
| pCFSA664-1 | Plasmid (complete) | CP033353 | 255,327 | 47.9 | IncHI2A, IncHI2, IncN, IncQ1 | |||||||||
| pCFSA664-2 | Plasmid (complete) | CP033354 | 41,696 | 45.4 | IncP-1-like trfA | |||||||||
| pCFSA664-3a | Plasmid (complete) | CP033355 | 61,841 | 42.4 | IncI2 | |||||||||
Contains the mcr-1 gene.
A spontaneous mcr-1 gene deletion was observed on pCFSA12 of Salmonella enterica subsp. enterica isolate CFSA12.
Predicted to be a potential monophasic variant of S. Typhimurium with a serotype antigenic formula of 4,[5],12:i:−.
The error-corrected sequence of CP033256 by Pilon with Illumina sequencing read data was identical to the previous version, so the accession number version did not change.
NA, plasmid replicon typing was not applicable for the chromosome. pCFSA664-2 was not predicted to have an Inc group using PlasmidFinder; however, the annotation contained an IncP-1-like trfA replication gene from an also-untypeable plasmid, pYDC107_41 (GenBank accession no. CP025711), which is 97.63% identical with 95% coverage to pCFSA664-2 (8).