A machine vision-assisted Argonaute-mediated fluorescence biosensor for the detection of viable Salmonella in food without convoluted DNA extraction and amplification procedures.

The precise identification viable pathogens hold paramount significance in the prevention of foodborne diseases outbreaks. In this study, we integrated machine vision and learning with single microsphere to develop a phage and Clostridium butyricum Argonaute (CbAgo)-mediated fluorescence biosensor for detecting viable Salmonella typhimurium (S. typhimurium) without convoluted DNA extraction and amplification procedures. Phage and lysis buffer was utilized to capture and lyse viable S. typhimurium, respectively. Subsequently, CbAgo can cleave the bacterial DNA to obtain target DNA that guides a newly targeted cleavage of fluorescent probes. After that, the resulting fluorescent signal accumulates on the streptavidin-modified single microsphere. The overall detection process is then analyzed and interpreted by machine vision and learning algorithms, achieving highly sensitive detection of S. typhimurium with a limit of detection at 40.5 CFU/mL and a linear range of 50-107 CFU/mL. Furthermore, the proposed biosensor demonstrates standard recovery rates and coefficients of variation at 93.22% - 106.02% and 1.47% - 12.75%, respectively. This biosensor exhibits exceptional sensitivity and selectivity, presenting a promising method for the rapid and effective detection of foodborne pathogens. ENVIRONMENTAL IMPLICATION: Bacterial pathogens exist widely in the environment and seriously threaten the safety of human life. In this study, we developed a phage and Clostridium butyricum Argonaute-mediated fluorescence biosensor for the detection of viable Salmonella typhimurium in environmental water and food samples. Compared with other Salmonella detection methods, this method does not need complex DNA extraction and amplification steps, which reduces the use of chemical reagents and experimental consumables in classic DNA extraction kit methods.

DNA Extraction- and Amplification-Free Nucleic Acid Biosensor for the Detection of Foodborne Pathogens Based on CRISPR/Cas12a and Argonaute Protein-Mediated Cascade Signal Amplification.

A novel method for detecting low levels of viable foodborne pathogens, specifically Salmonella typhimurium (S. typhimurium), has been developed. Traditional nucleic acid assay, such as polymerase chain reaction (PCR), often requires complex DNA extraction and amplification, making it challenging to differentiate between viable and nonviable pathogens. This assay employed a phage as the recognition element to precisely identify and lyse viable S. typhimurium that can undergo DNA extraction. It combined the efficient trans-cleavage activities of CRISPR/Cas12a with the specific cleavage advantages of Argonaute proteins, enabling ultrasensitive detection. This double-enzyme-mediated nucleic acid test can accurately distinguish viable and nonviable S. typhimurium with a detection limit of 23 CFU/mL without DNA amplification. The method was successfully applied to common food samples, producing results consistent with quantitative PCR tests. This work provides a promising platform for easily detecting viable foodborne pathogens with high sensitivity without the need for DNA extraction and amplification.

Intelligent active films of sodium alginate and konjac glucomannan mixed by Lycium ruthenicum anthocyanins and tea polyphenols for milk preservation and freshness monitoring.

To achieve real-time monitoring of food freshness, a pH-responsive film based on sodium alginate-konjac glucomannan loaded with Lycium ruthenicum anthocyanins (LRA) was prepared, with the addition of tea polyphenols (TP) to enhance the stability of LRA. The surface structure of the films was observed by AFM. The results of FTIR and molecular docking simulation showed that LRA and TP were bound to polysaccharide by hydrogen bonds. The mechanical properties, barrier properties, and antioxidant/antibacterial properties of the films were significantly improved and the films showed obvious color response to pH. Notably, the AFM images showed TP and LRA could lead to more severe damage to the bacterial structure. The results of molecular docking simulation suggested that TP and LRA could act on different components of the bacterial cell wall, indicating their synergistic mechanism in antimicrobial activity. Moreover, the stability of LRA was improved due to the interactions of TP and polysaccharides with LRA. The aggregates formed by TP and LRA were clearly observed by AFM. Finally, the film showed excellent preservation and freshness monitoring effect in milk. In conclusion, TP-LRA-SA-KGM intelligent film exhibited excellent performance and represented a promising novel food packaging material with potential applications.

Effect of potato peel on the determination of soluble solid content by visible near-infrared spectroscopy and model optimization.

The quantitative determination of the soluble solid content (SSC) of potatoes using NIR spectroscopy is useful for predicting the internal and external quality of potato products, especially fried products. In this study, the effect of peel on the partial least squares regression (PLSR) quantitative prediction of potato SSC was investigated by transmission and reflection. The results show that the variable sorting for normalization (VSN) pre-processing method improved model accuracy. Additive multiplicative scattering effects and intensity drift interference of the peels were reduced. The model accuracy reached a correlation coefficient of prediction (RP) of 0.85. The selection algorithm using variable combination population analysis and iterative retention of information variables (VCPA-IRIV) demonstrated that peel increases unnecessary information. When the effect of irrelevant variables was reduced, the results reached RP = 0.88 and the root mean square error of prediction (RMSEP) = 0.25 in the transmission mode was close to that of the full-wavelength peeled PLSR model (RP = 0.89 and RMSEP = 0.25). This indicates that the use of the combined algorithm (VSN-VCPA-IRIV) reduces the effect of the peel and enables samples with a peel to still be predicted accurately in the full-wavelength model. It also improves detection efficiency through the extraction of the necessary variables and optimizes the stability and accuracy of the model.

Characterization and release kinetics model of thymol from starch-based nanocomposite film into food simulator.

To improve the performance of potato starch films and solve the problems of high volatility and low stability of thymol (Thy), thymol was loaded into the channel of SBA-15 to prepare Thy-SBA-15, and the Thy-SBA-15/potato starch film was prepared. The results showed thymol was successfully loaded into the pores of SBA-15. The addition of Thy-SBA-15 enhanced the tensile strength of potato starch film (3.93 Mpa), reduced the water vapor permeability (1.56 × 10-12  g·d-1  m-1  Pa-1 , WVP) and moisture absorption (80.97%, MA), which enhanced the barrier properties of the films. Thy-SBA-15 had good compatibility with potato starch films. Notably, the thymol released from Thy-SBA-15/potato starch film was initially explosive, and then continuous, which showed this film could effectively slow down the release rate of thymol and prolong the fresh-keeping period of food. The Korsmeyer-Peppas model M t M ∞ = k t n (R2  > .96) had the best fit for the release curve of thymol. PRACTICAL APPLICATIONS: This work offers a new method for the preparation of potato starch sustained-release antibacterial film, and provides a theoretical basis and technical support for the development of intelligent packaging.

Characterization and Release Kinetics Study of Active Packaging Films Based on Modified Starch and Red Cabbage Anthocyanin Extract.

Active packaging films were prepared by adding red cabbage anthocyanin extract (RCAE) into acetylated distarch phosphate (ADSP). This paper investigated the influence of the interaction relationship between RCAE and the film matrix on the structure, barrier, antioxidant and release properties of active films. Sixteen principal compounds in RCAE were identified as anthocyanins based on mass spectroscopic analysis. Micromorphological observations indicated that the RCAE distribution uniformity in the films decreased as the RCAE content increased. When the concentration of RCAE was not higher than 20%, the moisture absorption and oxygen permeability of films decreased. The stability of RCAE in the films was enhanced by the electrostatic interaction between RCAE and ADSP with the formation of hydrogen bonds, which facilitated the sustainability of the antioxidant properties of films. The release kinetics of RCAE proved that the release rate of RCAE in active films was the fastest in distilled water, and Fickian's law was appropriate for portraying the release behavior. Moreover, the cytocompatibilty assay showed that the test films were biocompatible with a viability of >95% on HepG2 cells. Thus, this study has established the suitability of the films for applications in active and food packaging.

Pretreatment with high oxygen controlled atmosphere enhanced fresh-cut white mushroom (Agaricus bisporus) quality via activating wounding stress defenses.

BACKGROUND: High oxygen treatment has been proven to be effective in fresh-cut white mushroom preservation, however, the preservation effect and possible mechanisms in high oxygen controlled atmosphere pretreatment (HOCAP) on wounding stress are incompletely understood. RESULTS: In this study, based on the time chosen of HOCAP research, whole white mushrooms treated with 3 h HOCAP (80% O2 + 20% CO2 ) and the wounding resistant responses of their slices were mainly investigated through phenylpropane pathway, reactive oxygen species (ROS) scavenging system, and ascorbate-glutathione (AsA-GSH) cycle. Results showed that 3 h HOCAP can induce the production of hydrogen peroxide (H2 O2 ) and superoxide anion (O2 -• ) in the early stage, as well as the NADPH oxidase activity. Enzymes and endogenous antioxidants involved in ROS scavenging were enhanced by HOCAP during the whole storage. Besides, HOCAP maintained high level of phenylalanine ammonia-lyase (PAL) activity, enhanced the content of total phenolic and lignin, accelerated the AsA-GSH cycle. CONCLUSION: The results demonstrated that HOCAP induced defense responses by increasing the ROS in the early stage which stimulated the activities of ROS scavenging enzymes, along with the capability of increasing for wounding stress defense and resistance. This study provides a theoretical pretreatment technology for fresh-cut white mushroom preservation. © 2021 Society of Chemical Industry.

Characterization and release kinetics study of potato starch nanocomposite films containing mesoporous nano-silica incorporated with Thyme essential oil.

This study aimed to develop potato starch nanocomposite films containing mesoporous nano-silica (SBA-15, SBA-16 and MCM-41) incorporated with Thyme essential oil (TEO). TEO-SBA-15/potato starch films, TEO-SBA-16/potato starch films and TEO-MCM-41/potato starch films were prepared based on potato starch. The physical and mechanical properties of the nanocomposite films were also investigated. The results showed that the addition of mesoporous nano-silica incorporated with TEO improved the properties of potato starch nanocomposite films. Especially, the addition of TEO-MCM-41 markedly enhanced the tensile strength (4.33 MPa), and reduced the water vapor permeability (1.80 g·m-1·h-1·KPa-1) and moisture absorption (37.67%) of potato starch nanocomposite films. The results of scanning electron microscopy (SEM) analysis showed that TEO-MCM-41 hardly agglomerated in the potato starch nanocomposite films. Additionally, Fourier transform infrared (FTIR) spectroscopy and X-ray diffraction (XRD) confirmed that strong hydrogen bonds were formed between TEO-MCM-41 and potato starch. The release kinetics of TEO proved that incorporating TEO into the pores of mesoporous nano-silica could delay its release rate, and the Peleg model (t/(Mt - M0) = K1 + K2t) was suitable for describing the release behavior. The findings of this study suggested that TEO-MCM-41/potato starch films had a good application prospect in the field of slow-releasing and antimicrobial packaging materials.

Rat and human STINGs profile similarly towards anticancer/antiviral compounds.

Cyclic dinucleotides (CDNs) and antitumor/antiviral agents (DMXAA and CMA) trigger STING-dependent innate immunity activation. Accumulative evidences have showed that DMXAA and CMA selectively activate mouse, but not human STING signaling. The mechanism underlying this species selectivity remains poorly understood. In this report, we have shown that human and rat STINGs display more similar signaling profiles toward DMXAA and CMA than that of human and mouse STINGs, suggesting that rat is more suitable for preclinical testing of STING-targeted drugs. We have also determined the crystal structures of both apo rat STING and its complex with cyclic GMP-AMP with 2'5' and 3'5' phosphodiester linkage (2'3'-cGAMP), a human endogenous CDN. Structure-guided biochemical analysis also revealed the functional importance of the connecting loop (A140-N152) between membrane and cytosolic domains in STING activation. Taken together, these findings reveal that rat STING is more closely related to human STING in terms of substrate preference, serving as a foundation for the development of STING-targeted drugs.