Food safety is one of the biggest public issues occurring around the world. Microbiological, chemical, and physical hazards can lead to food safety issues, which may occur at all stages of the supply chain. In order to tackle food safety issues and safeguard consumer health, rapid, accurate, specific, and field-deployable detection methods meeting diverse requirements are one of the imperative measures for food safety assurance. CRISPR-Cas system, a newly emerging technology, has been successfully repurposed in biosensing and has demonstrated huge potential to establish conceptually novel detection methods with high sensitivity and specificity. This review focuses on CRISPR-Cas-based detection and its current status and huge potential specifically for food safety inspection. We firstly illustrate the pending problems in food safety and summarize the popular detection methods. We then describe the potential applications of CRISPR-Cas-based detection in food safety inspection. Finally, the challenges and futuristic opportunities are proposed and discussed. Generally speaking, the current food safety detection methods are still unsatisfactory in some ways such as being time-consuming, displaying unmet sensitivity and specificity standards, and there is a comparative paucity of multiplexed testing and POCT. Recent studies have shown that CRISPR-Cas-based biosensing is an innovative and fast-expanding technology, which could make up for the shortcomings of the existing methods or even replace them. To sum up, the implementation of CRISPR-Cas and the integration of CRISPR-Cas with other techniques is promising and desirable, which is expected to provide "customized" and "smart" detection methods for food safety inspection in the coming future.

CRISPR-Cas-based detection for food safety problems: Current status, challenges, and opportunities.

Infectious pathogens cause severe human illnesses and great deaths per year worldwide. Rapid, sensitive, and accurate detection of pathogens is of great importance for preventing infectious diseases caused by pathogens and optimizing medical healthcare systems. Inspired by a microbial defense system (i.e., CRISPR/ CRISPR-associated proteins (Cas) system, an adaptive immune system for protecting microorganisms from being attacked by invading species), a great many new biosensors have been successfully developed and widely applied in the detection of infectious viruses and pathogenic bacteria. Moreover, advanced nanotechnologies have also been integrated into these biosensors to improve their detection stability, sensitivity, and accuracy. In this review, the recent advance in CRISPR/Cas systems-based nano/biosensors and their applications in the detection of infectious viruses and pathogenic bacteria are comprehensively reviewed. First of all, the categories and working principles of CRISPR/Cas systems for establishing the nano/biosensors are simply introduced. Then, the design and construction of CRISPR/Cas systems-based nano/biosensors are comprehensively discussed. In the end, attentions are focused on the applications of CRISPR/Cas systems-based nano/biosensors in the detection of infectious viruses and pathogenic bacteria. Impressively, the remaining opportunities and challenges for the further design and development of CRISPR/Cas system-based nano/biosensors and their promising applications are proposed. 

CRISPR/Cas Systems‐Inspired Nano/Biosensors for Detecting Infectious Viruses and Pathogenic Bacteria

Innovative application of surface-enhanced Raman scattering (SERS) for rapid and nondestructive analyses has been gaining increasing attention for food safety and quality. SERS is based on inelastic scattering enhancement from molecules located near nanostructured metallic surfaces and has many advantages, including ultrasensitive detection and simple protocols. Current SERS-based quality analysis contains composition and structural information that can be used to establish an electronic file of the food samples for subsequent reference and traceability. SERS is a promising technique for the detection of chemical, biological, and harmful metal contaminants, as well as for food poisoning, and allergen identification using label-free or label-based methods, based on metals and semiconductors as substrates. Recognition elements, including immunosensors, aptasensors, or molecularly imprinted polymers, can be linked to SERS tags to specifically identify targeted contaminants and perform authenticity analysis. Herein, we highlight recent studies on SERS-based quality and safety analysis for different foods categories spanning the whole food chain, ‘from farm to table’ and processing, genetically modified food, and novel foods. Moreover, SERS detection is a potential tool that ensures food safety in an easy, rapid, reliable, and nondestructive manner during the COVID-19 pandemic.

https://www.mdpi.com/2304-8158/11/14/2097/pdf?version=1657865232

Innovative Application of SERS in Food Quality and Safety: A Brief Review of Recent Trends

Salmonella infection, particularly that caused by drug-resistant strain, has become a worldwide public health issue. Herein, we presented a CRISPR/Cas12a-signaling ARMS-PCR assay, termed cARMS, capable of sensitively detecting drug-resistant Salmonella enterica (S. enterica) involving single-nucleotide polymorphism (SNP). Owing to the dual-recognition processes, i.e., allele-specific primed polymerization and CRISPR/Cas12 binding, the cARMS assay yielded a high sensitivity for detecting SNP down to ∼0.5%. We used the cARMS assay to investigate the adaptation of SNP-involved drug-resistant S. enterica to salt stress. It was found that the mutants exhibited stronger adaptation to salt stress, indicating the potential risk of using high salt content as a sterilization strategy. The results verified the feasibility of the cARMS assay in controlling SNP-involved bacteria-associated biosafety.

Sensitive Detection of a Single-Nucleotide Polymorphism in Foodborne Pathogens Using CRISPR/Cas12a-Signaling ARMS-PCR.

High-efficiency enzyme-free catalyzed hairpin assembly-mediated homogeneous SERS and naked-eyes dual-mode assay for ultrasensitive and portable detection of mycotoxin.

An ultrasensitive surface-enhanced Raman scattering (SERS) biosensor driven by CRISPR/Cas12a was proposed for on-site nucleic acid detection. We tactfully modified single-strand DNA (ssDNA) with a target-responsive Prussian blue (PB) nanolabel to form a probe and fastened it in the microplate. Attributed to the specific base pairing and highly efficient trans-cleavage ability of the CRISPR/Cas12a effector, precise target DNA recognition and signal amplification can be achieved, respectively. In the presence of target DNA, trans-cleavage towards the probe was activated, leading to the release of a certain number of PB nanoparticles (NPs). Then, these free PB NPs would be removed. Under alkali treatment, the breakdown of the remaining PB NPs in the microplate was triggered, producing massive ferricyanide anions (Fe(CN)64-), which could exhibit a unique characteristic Raman peak that was located in the "biological Raman-silent region". By mixing the alkali-treated solution with the SERS substrate, Au@Ag core-shell NP, the concentration of the target DNA was finally exhibited as SERS signals with undisturbed background, which can be detected by a portable Raman spectrometer. Importantly, this strategy could display an ultralow detection limit of 224 aM for target DNA. Furthermore, by targeting cow milk as the adulterated ingredient in goat milk, the proposed biosensor was successfully applied to milk authenticity detection.

Ultrasensitive CRISPR/Cas12a-Driven SERS Biosensor for On-Site Nucleic Acid Detection and Its Application to Milk Authenticity Testing.

While one has yet to be built in China, a rice supply flow network would provide essential information for stabilizing and optimizing food distribution and ensuring food security. Moreover, it is unclear how socioeconomic and natural environmental factors affect the rice supply flow. In this study, 35,768 random samples from the rice sales market were collected to build the rice flow network of China. The rice supply patterns at a regional and provincial level were revealed by the rice flow network. At the regional level, Northeast, Central, and East China are the main rice source areas, supplying almost all of China. At the provincial level, economically developed megacities, such as Beijing and Shanghai, and provinces with low temperatures like Tibet and Qinghai, rely heavily on imported rice. Determinant analysis shows that the rice supply patterns are mainly attributable to the cultivated area, primary industry employees, the permanent population, and tertiary industry proportion. These factors explain 75%, 59%, 48%, and 31%, respectively, of the rice supply spatial patterns. Furthermore, the interactions of cultivated area and temperature, cultivated area and primary industry proportion, and cultivated area and precipitation can explain 94%, 91%, and 89%, respectively, of the rice supply patterns. These findings reveal China's rice supply patterns and their determinants in recent years, providing essential information for optimizing the layout of rice production and ensuring food security in the future scenarios of climate change and socioeconomic development.

Rice supply flows and their determinants in China

The development of efficient, economic, reliable, and accurate monitoring of hypochlorite (ClO-) in food matrices is in great demand for food safety assessment, particularly during its massive use against the COVID-19 epidemic. Here, we prepared an aggregation-induced emission (AIE) fluorophore tetraphenylethylene (TPE)-incorporated curcumin-based hybrid ratiometric fluorescence nanoprobe (Curcumin/TPE@HyNPs) through amphiphilic phospholipid polymer-powered nanoprecipitation, which exhibited a fast, highly sensitive, and selective response to the residual ClO- in real food matrices. Because of the inner filter effect (IFE) from curcumin toward TPE inside the nanoprobe, the bright fluorescence of TPE aggregation at ∼437 nm was effectively quenched, along with an enhanced fluorescence of curcumin at ∼478 nm. Once there was a ClO- residue in food matrices, ClO- triggered the oxidation of o-methoxyphenol inside curcumin and led to the almost complete absorption collapse, thereby terminating curcumin fluorescence at ∼478 nm and the IFE process. Accordingly, the fluorescence of TPE at ∼437 nm was recovered. In this case, a ratiometric fluorescent response of Curcumin/TPE@HyNPs toward the residual ClO- in food matrices (e.g., milk) was proposed with a low detection limit of 0.353 μM and a rapid response time of 140.0 s. Notably, the phospholipid polymer as the protection layer effectively reduced/evaded the nonspecific binding of signal reporters inside the nanoprobe, facilitating it to directly monitor the residual ClO- in real food matrices. This work provided a novel approach to utilize the unconventional AIE luminophors for constructing the efficient and reliable early warning mechanisms toward various food contaminants.

Aggregation-Induced Emission Fluorophore-Incorporated Curcumin-Based Ratiometric Nanoprobe for Hypochlorite Detection in Food Matrices.

To investigate the exposure of infants to phenylpyrazole and diamide insecticides during lactation, we collected 3467 breast milk samples of lactating women from 100 cities of 24 provinces in China and prepared 100 pooled samples together city-by-city. Among phenylpyrazole insecticides, fipronil and its metabolites (63-100%) were widely detected in breast milk, with total detection concentrations ranging from 178 to 2947 ng/L (median: 921 ng/L). Among diamide insecticides, chlorantraniliprole and flubendiamide were detected in breast milk, but their detection frequencies (20-85%) and concentration levels (nondetected to 89.9 ng/L) were far lower than those of total fipronils. The average estimated daily intake of infants exposed to total fipronils through breast milk is 209 ng/kg-bw/day by upper-bound scenario evaluation, which is higher than the acceptable daily intake (200 ng/kg-bw/day). This study indicates that infants have far higher exposure levels to fipronil than adults, while exposure levels to other types of phenylpyrazoles and diamide insecticides are low.

Occurrence of Phenylpyrazole and Diamide Insecticides in Lactating Women and Their Health Risks for Infants.

Sulfonamide antibiotics (SAs) are widely used in animal husbandry and aquaculture, and the excess residues of SAs in animal-derived foods will harm the health of consumers. In reality, various SAs were alternately used in animal husbandry and aquaculture, and thus, it is urgent need to develop simple and high-throughput methods for simultaneously detecting multiple SAs or groups of SAs in order to realize rapid screening of total SAs residues in animal-derived foods. We herein isolated a broad-specificity aptamer for SAs by using a multi-SAs systematic evolution of ligands by exponential enrichment (SELEX) strategy. The isolated broad-specificity aptamer has a higher binding affinity to five different SAs including sulfaquinoxaline (SQ), sulfamethoxypyridazine (SMPZ), sulfametoxydiazine (SMD), sulfachloropyridazine (SCP), and sulfapyridine (SPD) and, thus, can be used as a bioreceptor for developing various high-throughput methods for the simultaneous detection or rapid screening of above five SAs. Based on the isolated broad-specificity aptamer and Cy7 (diethylthiatricarbocyanine) displacement strategy, a colorimetric aptasensor was developed for the simultaneous detection of SQ, SMPZ, SMD, SCP, and SPD with a visual detection limit of 2.0-5.0 μM and a spectrometry detection limit of 0.2-0.5 μM. The colorimetric aptasensor was successfully used to detect SQ, SMPZ, SMD, SCP, and SPD in fish muscle with a recovery of 82%-92% and a RSD (n = 5) < 7%. The success of this study provided a promising bioreceptor for developing various high-throughput methods for on-site rapid screening of multiple SAs residues, as well as a simple method for the rapid and cost-effective screening of total SQ, SMPZ, SMD, SCP, and SPD in seafood.

Yongning Wu

Papers

Ultrasensitive CRISPR/Cas12a-Driven SERS Biosensor for On-Site Nucleic Acid Detection and Its Application to Milk Authenticity Testing.

Rice supply flows and their determinants in China

Aggregation-Induced Emission Fluorophore-Incorporated Curcumin-Based Ratiometric Nanoprobe for Hypochlorite Detection in Food Matrices.

Occurrence of Phenylpyrazole and Diamide Insecticides in Lactating Women and Their Health Risks for Infants.

Broad-Specificity Aptamer of Sulfonamides: Isolation and Its Application in Simultaneous Detection of Multiple Sulfonamides in Fish Sample.