Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR
Accurate detection of viable <i>Escherichia coli</i> O157:H7 in fresh produce wash water is critical for ensuring food safety and mitigating foodborne illnesses. This study evaluated an optimized on-filter propidium monoazide (PMA)-quantitative PCR (qPCR) method for detecting viable <...
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MDPI AG
2024-12-01
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| Series: | Microorganisms |
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| author | Zhao Chen |
| author_facet | Zhao Chen |
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| description | Accurate detection of viable <i>Escherichia coli</i> O157:H7 in fresh produce wash water is critical for ensuring food safety and mitigating foodborne illnesses. This study evaluated an optimized on-filter propidium monoazide (PMA)-quantitative PCR (qPCR) method for detecting viable <i>E. coli</i> O157:H7 in romaine lettuce wash water, involving PMA pretreatment on a filter to block DNA amplification from dead cells. The method consistently detected viable cells across chemical oxygen demand (COD) levels of 1000 and 200 mg O<sub>2</sub>/L, with no significant differences (<i>p</i> > 0.05), indicating its tolerance to organic matter interference. Optimization experiments identified 10 µM PMA with a 10 min exposure time as the most effective pretreatment, achieving efficient inhibition of DNA from dead cells while preserving viable cell integrity. The limit of detection (LOD) was 1.3 CFU/mL, confirming its suitability for detecting low bacterial loads. Performance evaluations revealed that PMA-qPCR was accurate at viable-to-dead cell ratios of 1:10 or higher but became less reliable when dead cells outnumbered viable cells by a factor of 10 or more. The study demonstrates the potential of on-filter PMA-qPCR for routine food safety monitoring protocols in the fresh produce industry, while highlighting the critical role of viable-to-dead cell ratios in ensuring accurate detection, particularly in challenging samples with high dead cell loads. |
| format | Article |
| id | doaj-art-60035c16808640ca940cc063317811ba |
| institution | Kabale University |
| issn | 2076-2607 |
| language | English |
| publishDate | 2024-12-01 |
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| series | Microorganisms |
| spelling | doaj-art-60035c16808640ca940cc063317811ba2025-01-24T13:42:22ZengMDPI AGMicroorganisms2076-26072024-12-011313410.3390/microorganisms13010034Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCRZhao Chen0Joint Institute for Food Safety and Applied Nutrition, University of Maryland, College Park, MD 20742, USAAccurate detection of viable <i>Escherichia coli</i> O157:H7 in fresh produce wash water is critical for ensuring food safety and mitigating foodborne illnesses. This study evaluated an optimized on-filter propidium monoazide (PMA)-quantitative PCR (qPCR) method for detecting viable <i>E. coli</i> O157:H7 in romaine lettuce wash water, involving PMA pretreatment on a filter to block DNA amplification from dead cells. The method consistently detected viable cells across chemical oxygen demand (COD) levels of 1000 and 200 mg O<sub>2</sub>/L, with no significant differences (<i>p</i> > 0.05), indicating its tolerance to organic matter interference. Optimization experiments identified 10 µM PMA with a 10 min exposure time as the most effective pretreatment, achieving efficient inhibition of DNA from dead cells while preserving viable cell integrity. The limit of detection (LOD) was 1.3 CFU/mL, confirming its suitability for detecting low bacterial loads. Performance evaluations revealed that PMA-qPCR was accurate at viable-to-dead cell ratios of 1:10 or higher but became less reliable when dead cells outnumbered viable cells by a factor of 10 or more. The study demonstrates the potential of on-filter PMA-qPCR for routine food safety monitoring protocols in the fresh produce industry, while highlighting the critical role of viable-to-dead cell ratios in ensuring accurate detection, particularly in challenging samples with high dead cell loads.https://www.mdpi.com/2076-2607/13/1/34<i>Escherichia coli</i> O157:H7produce wash waterquantitative PCRpropidium monoazideviability detection |
| spellingShingle | Zhao Chen Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR Microorganisms <i>Escherichia coli</i> O157:H7 produce wash water quantitative PCR propidium monoazide viability detection |
| title | Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR |
| title_full | Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR |
| title_fullStr | Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR |
| title_full_unstemmed | Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR |
| title_short | Enhanced Detection of Viable <i>Escherichia coli</i> O157:H7 in Romaine Lettuce Wash Water Using On-Filter Propidium Monoazide-Quantitative PCR |
| title_sort | enhanced detection of viable i escherichia coli i o157 h7 in romaine lettuce wash water using on filter propidium monoazide quantitative pcr |
| topic | <i>Escherichia coli</i> O157:H7 produce wash water quantitative PCR propidium monoazide viability detection |
| url | https://www.mdpi.com/2076-2607/13/1/34 |
| work_keys_str_mv | AT zhaochen enhanceddetectionofviableiescherichiacoliio157h7inromainelettucewashwaterusingonfilterpropidiummonoazidequantitativepcr |