Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety
Shiga toxin-producing <i>Escherichia coli</i> (STEC) and <i>Listeria monocytogenes</i> are routinely responsible for severe foodborne illnesses in the United States. Current identification methods utilized by the U.S. Food Safety Inspection Service require at least four days...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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MDPI AG
2023-12-01
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| Series: | Foods |
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| Online Access: | https://www.mdpi.com/2304-8158/13/1/16 |
| _version_ | 1827384712417509376 |
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| author | Katrina L. Counihan Siddhartha Kanrar Shannon Tilman Andrew Gehring |
| author_facet | Katrina L. Counihan Siddhartha Kanrar Shannon Tilman Andrew Gehring |
| author_sort | Katrina L. Counihan |
| collection | DOAJ |
| description | Shiga toxin-producing <i>Escherichia coli</i> (STEC) and <i>Listeria monocytogenes</i> are routinely responsible for severe foodborne illnesses in the United States. Current identification methods utilized by the U.S. Food Safety Inspection Service require at least four days to identify STEC and six days for <i>L. monocytogenes</i>. Adoption of long-read, whole genome sequencing for food safety testing could significantly reduce the time needed for identification, but method development costs are high. Therefore, the goal of this project was to use NanoSim-H software to simulate Oxford Nanopore sequencing reads to assess the feasibility of sequencing-based foodborne pathogen detection and guide experimental design. Sequencing reads were simulated for STEC, <i>L. monocytogenes</i>, and a 1:1 combination of STEC and <i>Bos taurus</i> genomes using NanoSim-H. At least 2500 simulated reads were needed to identify the seven genes of interest targeted in STEC, and at least 500 reads were needed to detect the gene targeted in <i>L. monocytogenes</i>. Genome coverage of 30x was estimated at 21,521, and 11,802 reads for STEC and <i>L. monocytogenes</i>, respectively. Approximately 5–6% of reads simulated from both bacteria did not align with their respective reference genomes due to the introduction of errors. For the STEC and <i>B. taurus</i> 1:1 genome mixture, all genes of interest were detected with 1,000,000 reads, but less than 1x coverage was obtained. The results suggested sample enrichment would be necessary to detect foodborne pathogens with long-read sequencing, but this would still decrease the time needed from current methods. Additionally, simulation data will be useful for reducing the time and expense associated with laboratory experimentation. |
| first_indexed | 2024-03-08T15:07:23Z |
| format | Article |
| id | doaj.art-548a3c128d17492ba0ee89857382a175 |
| institution | Directory Open Access Journal |
| issn | 2304-8158 |
| language | English |
| last_indexed | 2024-03-08T15:07:23Z |
| publishDate | 2023-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Foods |
| spelling | doaj.art-548a3c128d17492ba0ee89857382a1752024-01-10T14:56:35ZengMDPI AGFoods2304-81582023-12-011311610.3390/foods13010016Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food SafetyKatrina L. Counihan0Siddhartha Kanrar1Shannon Tilman2Andrew Gehring3Eastern Regional Research Center, United States Department of Agriculture, Agricultural Research Service, Wyndmoor, PA 19038, USAEastern Regional Research Center, United States Department of Agriculture, Agricultural Research Service, Wyndmoor, PA 19038, USAEastern Regional Research Center, United States Department of Agriculture, Agricultural Research Service, Wyndmoor, PA 19038, USAEastern Regional Research Center, United States Department of Agriculture, Agricultural Research Service, Wyndmoor, PA 19038, USAShiga toxin-producing <i>Escherichia coli</i> (STEC) and <i>Listeria monocytogenes</i> are routinely responsible for severe foodborne illnesses in the United States. Current identification methods utilized by the U.S. Food Safety Inspection Service require at least four days to identify STEC and six days for <i>L. monocytogenes</i>. Adoption of long-read, whole genome sequencing for food safety testing could significantly reduce the time needed for identification, but method development costs are high. Therefore, the goal of this project was to use NanoSim-H software to simulate Oxford Nanopore sequencing reads to assess the feasibility of sequencing-based foodborne pathogen detection and guide experimental design. Sequencing reads were simulated for STEC, <i>L. monocytogenes</i>, and a 1:1 combination of STEC and <i>Bos taurus</i> genomes using NanoSim-H. At least 2500 simulated reads were needed to identify the seven genes of interest targeted in STEC, and at least 500 reads were needed to detect the gene targeted in <i>L. monocytogenes</i>. Genome coverage of 30x was estimated at 21,521, and 11,802 reads for STEC and <i>L. monocytogenes</i>, respectively. Approximately 5–6% of reads simulated from both bacteria did not align with their respective reference genomes due to the introduction of errors. For the STEC and <i>B. taurus</i> 1:1 genome mixture, all genes of interest were detected with 1,000,000 reads, but less than 1x coverage was obtained. The results suggested sample enrichment would be necessary to detect foodborne pathogens with long-read sequencing, but this would still decrease the time needed from current methods. Additionally, simulation data will be useful for reducing the time and expense associated with laboratory experimentation.https://www.mdpi.com/2304-8158/13/1/16Shiga toxin-producing <i>Escherichia coli</i> O157:H7<i>Listeria monocytogenes</i><i>Bos taurus</i>foodborne pathogensvirulence genes |
| spellingShingle | Katrina L. Counihan Siddhartha Kanrar Shannon Tilman Andrew Gehring Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety Foods Shiga toxin-producing <i>Escherichia coli</i> O157:H7 <i>Listeria monocytogenes</i> <i>Bos taurus</i> foodborne pathogens virulence genes |
| title | Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety |
| title_full | Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety |
| title_fullStr | Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety |
| title_full_unstemmed | Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety |
| title_short | Evaluation of Long-Read Sequencing Simulators to Assess Real-World Applications for Food Safety |
| title_sort | evaluation of long read sequencing simulators to assess real world applications for food safety |
| topic | Shiga toxin-producing <i>Escherichia coli</i> O157:H7 <i>Listeria monocytogenes</i> <i>Bos taurus</i> foodborne pathogens virulence genes |
| url | https://www.mdpi.com/2304-8158/13/1/16 |
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