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A Specific and Sensitive Aptamer-Based Digital PCR Chip for Salmonella typhimurium Detection.

ABSTRACT: Food poisoning and infectious diseases caused by Salmonella typhimurium (S. typhimurium) are serious public health concerns for human health and food safety. The diversity and complexity of food matrices pose great challenges for rapid and ultra-sensitive detection of S. typhimurium in food samples. A method capable of identification, detection, and quantification of S. typhimurium is essential for addressing these issues. In this study, aptamer-coated magnetic beads (Apt-MBs) are employed as capture bio-probes to specifically and selectively concentrate S. typhimurium in food samples. A self-priming chip-based digital PCR was then presented as another biosensor for on-site detection and quantification of S. typhimurium cells. The chip we developed was robust and did not require any external power for sample loading. The combination of Apt-MBs with an on-chip digital detection realized the integration into lab-on-a-chip-based biosensors for on-site monitoring of foodborne pathogens. It was possible to capture and detect S. typhimurium cells as low as 90 CFU/reaction with a capture efficiency of 94.5%. Additionally, the whole process only took about 2 h. This unique platform could also be used to monitor other target bacteria with high specificity and sensitivity by utilizing different aptamers. Furthermore, the platform has potential applications in point-of-care testing in the future.

SUBMITTER: Suo Y

PROVIDER: S-EPMC9313467 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The difficulty in detecting viable but non-culturable (VBNC) Salmonella by culture-dependent methods poses a risk to food safety. In our study, we applied a viability test to Salmonella following a lethal treatment and to flour samples inoculated with Salmonella to evaluate the effectiveness of viability polymerase chain reaction (PCR). Our findings revealed that the combination of both ddPCR and qPCR with those DNA-intercalating dyes could quantify viable cells at low concentrations when the plate counting method failed to detect them post-inactivation. Prolonged UV exposure did not induce cell membrane disruption, as confirmed with PMA-ddPCR, with insignificant differences in gene copies. However, samples exposed to DyeTox13 and DyeTox13 + EMA showed lower gene copy numbers, implying that enzymatic activity was decreased by UV exposure duration. In addition, temperature-dependent survival in flour revealed uniform decay rates and D values (time required for a 1 log reduction) of DNA in untreated samples across various temperatures. By contrast, different decay rates were observed with DNA-intercalating dyes (DyeTox13 and DyeTox13 + EMA), showing faster metabolic activity loss at higher temperatures in flour. The decay rates and D values, determined through plate counting and those DNA-intercalating dyes, indicated the potential presence of VBNC Salmonella. A strong correlation between DyeTox13 dyes and the plate counting method suggested DyeTox13 as a rapid alternative for detecting Salmonella in flour. The ddPCR with DNA-intercalating dyes could effectively evaluate Salmonella viability, facilitating more precise monitoring of VBNC in food.ImportanceSalmonella, a major foodborne pathogen, poses significant risks, particularly to vulnerable groups like infants, older people, and the immunocompromised. Accurate detection is vital for public health and food safety, given its potential to cause severe and life-threatening symptoms. Our study demonstrated digital polymerase chain reaction (ddPCR) with DNA-intercalating dyes for identifying the different physiological statuses of Salmonella. Also, the application of ddPCR with DNA-intercalating dyes offers quantification of viable cells post-disinfection as an alternative method in food. Utilizing ddPCR and DNA-intercalating dyes, we enhanced the detection of VBNC Salmonella, a form often undetectable by conventional methods. This innovative approach could significantly improve the precision and efficiency of detection for viable Salmonella. By providing deeper insights into its transmission potential, our method is a critical tool in preventing outbreaks and ensuring the safety of food products. This research contributes substantially to global efforts in controlling foodborne illnesses and safeguarding public health.

Dataset Information

A Specific and Sensitive Aptamer-Based Digital PCR Chip for Salmonella typhimurium Detection.

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