Sensing Staphylococcus Aureus In Fresh Meat Using Upconversion Fluorescence

Fresh meat is a very important part of people’s diet,rich in nutrients such as fat,protein,zinc,iron,vitamins,etc.However,due to the frequent occurrence of fresh meat safety accidents,the safety of fresh meat has been widely concerned by the society.Staphylococcus aureus as flesh-eating bacteria is one of the most common foodborne pathogens,and the limit of detection of Staphylococcus aureus is also clearly stipulated in the national standards of food safety.At present,although conventional detection methods can achieve high-sensitivity detection,they often have long detection cycles and complicated operation steps.Therefore,it is of great significance to develop a simple,rapid,specific and sensitive detection method for ensuring the safety and quality of fresh meat and promoting the green and healthy development of the fresh meat industry.In recent years,upconversion fluorescent nanotechnology has attracted widespread the attention of researchers due to its special luminescence mechanism and unique photochemical advantages,and has been widely applied in the field of food detection.Based on that,it is important to develop multiple detection modes and principles and gradually explore new detection methods to realize simple,rapid and highly sensitive detection of Staphylococcus aureus in fresh meat.The specific research details are as follows:1.Rapid detection of Staphylococcus aureus based on Fe³⁺quenching upconversion fluorescence.Based on the quenching effect of Fe³⁺on upconversion nanomaterials,a composite detection system of Fe³⁺-upconversion-aptamer-hydrogen peroxide was constructed to realize the rapid detection of Staphylococcus aureus in fresh meat.First,Fe²⁺was encapsulated by aptamers in the silica mesopores on the surface of upconversion nanoparticles.When Staphylococcus aureus was added in the system,the aptamers preferentially bound to Staphylococcus aureus and Fe²⁺was released from the mesopores,leading to the reduction of H₂O₂to Fe³⁺.Fe³⁺further formed a complex with the upconversion nanoparticles allowing the upconversion fluorescence quenching.Under optimal experimental conditions,the fluorescence intensity at 658 nm was linearly related to the logarithm of the concentration of Staphylococcus aureus with a limit of detection（LOD）25 CFU/m L in the range of 63-6.3×10⁶CFU/m L.At the same time,the developed method was applied to detect Staphylococcus aureus in the actual fresh meat samples with the recovery rate recorded in the of 91-105%,the relative standard deviation in the range of2.20-7.53%,and there was no significant difference between the results of the traditional plate counting method.The study indicates that the constructed detection system can be used for rapid quantitative detection of Staphylococcus aureus in fresh meat.2.Rapid detection of Staphylococcus aureus based on upconversion fluorescence and colorimetry.Based on magnetic separation,a dual-signal composite detection system of upconversion-horseradish peroxidase-aptamer-magnetic nanomaterials was constructed to realize the rapid detection of Staphylococcus aureus in fresh meat.The aptamer complementary chain and horseradish peroxidase both were modified on the surface of the upconversion nanomaterial,and the aptamer was labeled on the surface of the magnetic nanomaterial to form an immune complex through base complementation.After the addition of Staphylococcus aureus,the aptamer preferentially bound to Staphylococcus aureus,which resulted in the shedding of upconversion from the surface of the magnetic nanoparticles and restoring the fluorescence intensity and absorbance values in the supernatant.Under the optimal experimental conditions,a strong linear relationship was observed between the fluorescence intensity and absorbance with logarithmic Staphylococcus aureus concentration in the range of 56-5.6×10⁶CFU/m L with LOD of 22 CFU/m L for fluorescence and 20 CFU/m L for colorimetry.The spiked samples of different concentrations were examined,the recoveries were 92%-103%with the relative standard deviation of 3.88%-7.61%for the fluorescent method and 92%-104%with the relative standard deviation of 3.23%-6.53%for the colorimetric method.The study proves that the constructed detection system can be used for the quantitative detection of Staphylococcus aureus in fresh meat,and it has improved the detection sensitivity and stability compared with the previous chapter research.3.Rapid detection of Staphylococcus aureus based on upconversion fluorescence resonance energy transfer.Based on the fluorescence resonance energy transfer effect,an upconversion-aptamer-cobalt oxyhydroxide composite detection system was constructed.The aptamer was modified on the surface of the upconversion nanoparticles,and can self-assemble on the surface of the cobalt oxyhydroxide nanosheet through van der Waals force,resulting in the occurrence of fluorescence resonance energy transfer and the strong upconversion fluorescence can be effectively quenched.When Staphylococcus aureus was added,the aptamer preferentially bound to Staphylococcus aureus,so that the upconversion nanoparticles was separated from the surface of the cobalt oxyhydroxide nanosheet and the upconversion fluorescence signal was restored.Under the optimal experimental conditions,the linear range was 45-4.5×10⁶CFU/m Lwith LOD of 15 CFU/m L.Furthermore,the recovery rate was 92-104%and the relative standard deviation was between 3.23 and 6.52%in the spiked recovery tests of real samples.Compared with the detection in the previous two chapters,the system constructed not only simplifies the test preparation process and detection procedures,but also achieves a lower detection limit,indicating that the constructed detection system based on fluorescence resonance energy transfer can be used for simple,rapid and highly sensitive quantitative detection of Staphylococcus aureus in fresh meat.The research results provide a method for the rapid detection of food borne pathogens in fresh meat,which is of great significance to ensure the safety and quality of fresh meat in China.