Full
Outline
To establish a recombinase polymerase amplification-lateral flow (RPA-LF) detection method for the detection of Listeria monocytogenes in dairy products, targeting the species-specific 16S rRNA gene and the hemolysin-encoding gene hly.
Primers and probes suitable for RPA-LF were designed based on the conserved fragments of the target sequence, and the optimal primer-probe combination was selected, the corresponding detection method was established, and its reaction conditions were optimized.
Under optimized conditions, the total reaction time of the RPA-LF assay was only 20 min, and the results could be observed visually by the naked eye. The method showed no cross-reactivity with 8 kinds of common foodborne pathogenic microorganisms, demonstrating good specificity. It could also distinguish Listeria monocytogenes from other Listeria species, indicating good anti-interference ability against bacteria from the same genus. The limit of detection was as low as 82 fg/μL, corresponding to approximately 1.8×102 CFU/mL, demonstrating high sensitivity. Furthermore,when used for routine sample detection, the method yielded results consistent with those obtained by traditional microbiological methods.
The RPA-LF method targeting the hemolysin gene of Listeria monocytogenes established in this study has several advantages, including low instrument dependency, simple operation, high sensitivity, good specificity and direct visual readout. It provides a new approach for the rapid on-site detection and screening diagnosis of this pathogen.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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