Application of CRISPR/Cas System-based Biosensors in Detection of Foodborne Pathogens

Authors

Miao ZOU, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Jingyi ZENG, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Dandan ZENG, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Yuhan FU, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Xu ZHANG, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Bo ZHAO, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , China
Tianlu MO, School of Health Science and Engineering , University of Shanghai for Science and Technology , Shanghai 200093 , ChinaFollow

Corresponding Author(s)

莫天录（1988—），男，博士，副教授，博士研究生导师，主要从事微生物学和合成生物学研究。E-mail：motianlu@usst.edu.cn

Abstract

The CRISPR/Cas system can recognize foreign DNA,cut the target,and block or silence the expression of the foreign gene.Due to its precise targeting function,the CRISPR/Cas system has been developed as a highly efficient gene editing tool.With high specificity and operability,the CRISPR/Cas system has been combined with biosensors to improve the detection specificity and efficiency of the sensors.This review introduces the characteristics and applications of CRISPR/Cas system,analyzes the latest progress in the application of CRISPR/Cas system-based biosensors in the detection of foodborne pathogens,and summarizes and prospects the application of such biosensors.

Publication Date

5-15-2025

First Page

10

Last Page

21

DOI

10.12441/spyswjs.20240913001

Recommended Citation

ZOU, Miao; ZENG, Jingyi; ZENG, Dandan; FU, Yuhan; ZHANG, Xu; ZHAO, Bo; and MO, Tianlu (2025) "Application of CRISPR/Cas System-based Biosensors in Detection of Foodborne Pathogens," Journal of Food Science and Biotechnology: Vol. 44: Iss. 5, Article 2.
DOI: 10.12441/spyswjs.20240913001
Available at: https://spsw.spyswjs.cnjournals.com/journal/vol44/iss5/2

References

［1］ MATHER A E，GILMOUR M W，REID S W J，et al.Foodborne bacterial pathogens：genome-based approaches for enduring and emerging threats in a complex and changing world ［J］.Nature Reviews Microbiology，2024，22（9）：543-555.
［2］ 孙秀兰，付旭冉，鲍琦，等.新一代诊断技术：CRISPR 系统及其在分子诊断中的应用 ［J］.食品与生物技术学报，2022，41（7）：57-70.SUN X L，FU X R，BAO Q，et al.Next generation diagnostic technology：CRISPR system and its application in molecular diagnostics ［J］.Journal of Food Science and Biotechnology，2022，41（7）：57-70.（in Chinese ）
［3］ LI X，ZHANG X L，SHI X L，et al.Review in isothermal amplification technology in food microbiological detection ［J］.Food Science and Biotechnology，2022，31（12）：1501 -1511.
［4］ LI R，SHI J X，LIU B，et al.Inter-laboratory validation of visual loop-mediated isothermal amplification assays for GM contents screening ［J］.Food Chemistry，2019，274：659-663.
［5］ ZENG D D，JIAO J L，MO T L.Combination of nucleic acid amplification and CRISPR/Cas technology in pathogen detection［J］.Frontiers in Microbiology，2024，15：1355234.
［6］ SONG J Z，LIU C C，MAUK M G，et al.Two-stage isothermal enzymatic amplification for concurrent multiplex molecular detection ［J］.Clinical Chemistry，2017，63（3）：714-722.
［7］ VAN DONGEN J E，BERENDSEN J T W，STEENBERGEN R D M，et al.Point-of-care CRISPR/Cas nucleic acid detection：recent advances，challenges and opportunities ［J］.Biosensors and Bioelectronics，2020，166：112445.
［8］ 王芳妹，徐越，林丹，等.包装饮用水中铜绿假单胞菌实时荧光定量 PCR快速检测［J］.食品与机械，2023，39（6）：75-80.WANG F M，XU Y，LIN D，et al.Study on the rapid detection of Pseudomonas aeruginosa by real-time fluorescence quantitative PCR in packaged drinking water［J］.Food & Machinery，2023，39（6）：75-80.（in Chinese ）.
［9］ LI G D，WANG J J，TIAN L，et al.Rapid and ultrasensitive detection of Staphylococcus aureus by a one-pot system integrating Pyrococcus furiosus argonaute with loop-mediated isothermal amplification ［J］.Journal of Microbiology and Biotechnology，2025，35：e2504034.
［10］ LIU Y，XIA W Q，ZHAO W，et al.RT-RPA- PfAgo system：a rapid，sensitive，and specific multiplex detection method for rice-infecting viruses ［J］.Biosensors，2023，13（10）：941.
［11］ MAKAROVA K S，WOLF Y I，IRANZO J，et al.Evolutionary classification of CRISPR-Cas systems：a burst of class 2 and derived variants ［J］.Nature Reviews Microbiology，2020，18（2）：67-83.
［12］ MACHUCA B，BIANCHI J.Aplicação do CAR-T cell e CRISPR/Cas 9 no tratamento de mieloma múltiplo recidivado e/ou refratário ［J］.Hematology，Transfusion and Cell Therapy，2023，45（Suppl.4）：S410-S411.
［13］ LI S Y，CHENG Q X，LIU J K，et al.CRISPR-Cas 12a has both cis- and trans -cleavage activities on single-stranded DNA ［J］.Cell Research，2018，28：491-493.
［14］ GOOTENBERG J S，ABUDAYYEH O O，LEE J W，et al.Nucleic acid detection with CRISPR-Cas 13a/C2c2［J］.Science，2017，356（6336）：438-442.
［15］ YANG Y，ZHAI S S，ZHANG L，et al.A gold nanoparticle-enhanced dCas 9-mediated fluorescence resonance energy transfer for nucleic acid detection ［J］.Talanta，2025，282：126978.
［16］ NAWALY H，TSUJI Y，MATSUDA Y.Rapid and precise genome editing in a marine diatom，Thalassiosira pseudonana by Cas 9 nickase （D10A） ［J］.Algal Research，2020，47：101855.
［17］ YU D H，LIANG B，XU H P，et al.CRISPR/Cas 12a-based assay for the rapid and high-sensitivity detection of Streptococcus agalactiae colonization in pregnant women with premature rupture of membrane ［J］.Annals of Clinical Microbiology and Antimicrobials，2023，22（1）：8.
［18］ MAKAROVA K S，WOLF Y I，IRANZO J，et al.Evolutionary classification of CRISPR-Cas systems：a burst of class 2 and derived variants ［J］.Nature Reviews Microbiology，2020，18：67-83.
［19］ CHEN J S，MA E B，HARRINGTON L B，et al.CRISPR-Cas 12a target binding unleashes indiscriminate single-stranded DNase activity ［J］.Science，2018，360（6387）：436-439.
［20］ LI P R，WANG Z X，XU Z K，et al.An RPA-assisted CRISPR/Cas 12a assay combining fluorescence and lateral flow strips for the rapid detection of enterotoxigenic Bacillus cereus ［J］.Journal of Agricultural and Food Chemistry，2024，72（24）：14000 -14010.
［21］ GUO Y Q，LI C，GUO W，et al.Advanced electrochemical biosensing toward Staphylococcus aureus based on the RPA-CRISPR/Cas 12a system and conductive nanocomposite ［J］.Journal of Agricultural and Food Chemistry，2024，72（41）：22918 -22925.
［22］ TAO X Q，YUE L，TIAN T，et al.Sensitive and on-site detection of Staphylococcus aureus based on CRISPR/Cas 13a-assisted chemiluminescence resonance energy transfer ［J］.Analytical Chemistry，2024，96（22）：9270 -9277.
［23］ HARRINGTON L B，BURSTEIN D，CHEN J S，et al.Programmed DNA destruction by miniature CRISPR-Cas14 enzymes［J］.Science，2018，362（6416）：839-842.
［24］ LI F，YE Q H，CHEN M T，et al.An ultrasensitive CRISPR/Cas 12a based electrochemical biosensor for Listeria monocytogenes detection ［J］.Biosensors and Bioelectronics，2021，179：113073.
［25］ ZHEN D S，ZHANG S Q，YANG A F，et al.A supersensitive electrochemical sensor based on RCA amplification-assisted “silver chain ” -linked gold interdigital electrodes and CRISPR/Cas 9 for the detection of Staphylococcus aureus in food［J］.Food Chemistry，2024，440：138197.
［26］ ZHANG J L，ZHOU M，MAO B Y，et al.The construction of COFs functionalized CRISPR electrochemical sensor for ultrasensitive detection of bacteria by hyper-branched rolling circle amplification ［J］.Sensors and Actuators B：Chemical，2024，409：135610.
［27］ WANG J F，JIANG H，CHEN Y H，et al.CRISPR/Cas 9-mediated SERS/colorimetric dual-mode lateral flow platform combined with smartphone for rapid and sensitive detection of Staphylococcus aureus ［J］.Biosensors and Bioelectronics，2024，249：116046.
［28］ JIANG H，CHANG W，ZHU X F，et al.Development of a colorimetric and SERS dual-signal platform via dCas 9-mediated chain assembly of bifunctional Au@Pt nanozymes for ultrasensitive and robust Salmonella assay［J］.Analytical Chemistry，2024，96（31）：12684 -12691.
［29］ MA L，PENG L，YIN L J，et al.CRISPR-Cas 12a-powered dual-mode biosensor for ultrasensitive and cross-validating detection of pathogenic bacteria ［J］.ACS Sensors，2021，6（8）：2920 -2927.
［30］ CHEN X Y，WANG L，HE F，et al.Label-free colorimetric method for detection of Vibrio parahaemolyticus by trimming the G-quadruplex DNAzyme with CRISPR/Cas 12a［J］.Analytical Chemistry，2021，93（42）：14300 -14306.
［31］ MA L，WANG J J，LI Y R，et al.A ratiometric fluorescent biosensing platform for ultrasensitive detection of Salmonella typhimurium via CRISPR/Cas 12a and silver nanoclusters ［J］.Journal of Hazardous Materials，2023，443：130234.
［32］ CHEN R，ZHAO J P，HAN M J，et al.DNA extraction- and amplification-free nucleic acid biosensor for the detection of foodborne pathogens based on CRISPR/Cas 12a and argonaute protein-mediated cascade signal amplification［J］.Journal of Agricultural and Food Chemistry，2023，71（46）：18037 -18045.
［33］ WANG Y S，DU P F，SHAO Y C，et al.An innovative and efficient fluorescent detection technique for Salmonella in animal-derived foods using the CRISPR/Cas 12a-HCR system combined with PCR/RAA ［J］.Journal of Agricultural and Food Chemistry，2024，72（15）：8831 -8839.
［34］ JIA F，LI B Y，HE Y W，et al.An amplification-free CRISPR-SERS biosensor for specific，sensitive and rapid detection of Salmonella typhimurium in poultry ［J］.LWT-Food Science and Technology，2023，189：115476.
［35］ DUAN M L，LI B Y，HE Y W，et al.A CG@MXene nanocomposite-driven E-CRISPR biosensor for the rapid and sensitive detection of Salmonella typhimurium in food［J］.Talanta，2024，266（1）：125011.
［36］ WANG C Y，ZHANG Y T，LIU S S，et al.Allosteric probe-triggered isothermal amplification to activate CRISPR/Cas 12a for sensitive electrochemiluminescence detection of Salmonella ［J］.Food Chemistry，2023，425：136382.
［37］ ZHUANG J W，ZHAO Z Y，LIAN K，et al.SERS-based CRISPR/Cas assay on microfluidic paper analytical devices for supersensitive detection of pathogenic bacteria in foods［J］.Biosensors and Bioelectronics，2022，207：114167.
［38］ XING G W，SHANG Y T，WANG X R，et al.Multiplexed detection of foodborne pathogens using one-pot CRISPR/Cas 12a combined with recombinase aided amplification on a finger-actuated microfluidic biosensor［J］.Biosensors and Bioelectronics，2023，220：114885.
［39］ XIANG X R，XING G W，LIU Y T，et al.Immunomagnetic separation combined with RCA-CRISPR/Cas 12a for the detection of Salmonella typhimurium on a figure-actuated microfluidic biosensor［J］.Journal of Agricultural and Food Chemistry，2023，71（36）：13518 -13526.
［40］ WU H，CHEN Y J，YANG Q Q，et al.A reversible valve-assisted chip coupling with integrated sample treatment and CRISPR/Cas 12a for visual detection of Vibrio parahaemolyticus ［J］.Biosensors and Bioelectronics，2021，188：113352.
［41］ ZHOU B Q，YE Q H，LI F，et al.CRISPR/Cas 12a based fluorescence-enhanced lateral flow biosensor for detection of Staphylococcus aureus ［J］.Sensors and Actuators B：Chemical，2022，351：130906.
［42］ QIAN J J，HUANG D，NI D S，et al.A portable CRISPR Cas12a based lateral flow platform for sensitive detection of Staphylococcus aureus with double insurance ［J］.Food Control，2022，132：108485.
［43］ LEE S Y，OH S W.Lateral flow biosensor based on LAMP-CRISPR/Cas 12a for sensitive and visualized detection of Salmonella spp.［J］.Food Control，2023，145：109494.
［44］ SHANG Y T，XING G W，LIN J X，et al.Multiplex bacteria detection using one-pot CRISPR/Cas 13a-based droplet microfluidics ［J］.Biosensors and Bioelectronics，2024，243：115771.
［45］ XIANG X R，LI F，YE Q H，et al.High-throughput microfluidic strategy based on RAA-CRISPR/Cas 13a dual signal amplification for accurate identification of pathogenic Listeria［J］.Sensors and Actuators B：Chemical，2022，358：131517.
［46］ ZHANG T，ZHOU W H，LIN X Y，et al.Light-up RNA aptamer signaling-CRISPR-Cas 13a-based mix-and-read assays for profiling viable pathogenic bacteria ［J］.Biosensors and Bioelectronics，2021，176：112906.
［47］ ZHOU J，YIN L J，DONG Y N，et al.CRISPR-Cas 13a based bacterial detection platform：sensing pathogen Staphylococcus aureus in food samples ［J］.Analytica Chimica Acta，2020，1127：225-233.
［48］ 陈欢，王宇，黄钰，等.基于 CRISPR/Cas 系统的病原微生物检测体系构建研究进展 ［J］.微生物学报，2022，62（9）：3271 -3288.CHEN H，WANG Y，HUANG Y，et al.Research progress in construction of pathogenic microorganism detection system based on CRISPR/Cas system ［J］.Acta Microbiologica Sinica，2022，62（9）：3271 -3288.（in Chinese）
［49］ YANG Y J，LI X Y，WANG X B，et al.CRISPR-Cas-based colorimetric strategies for nucleic acids detection ［J］.TrAC Trends in Analytical Chemistry，2025，182：118058.
［50］ CREGO-VICENTE B，DEL OLMO M D，MURO A，et al.Multiplexing LAMP assays：a methodological review and diagnostic application ［J］.International Journal of Molecular Sciences，2024，25（12）：6374.
［51］ 曹宁，申炳阳，喻梓瑄，等.用于 3种食源性致病菌核酸提取的微流控芯片制备 ［J］.食品与生物技术学报，2022，41（1）：28-35.CAO N，SHEN B Y，YU Z X，et al.Microfluidic chip for nucleic acid extraction of three food-borne pathogenic bacteria［J］.Journal of Food Science and Biotechnology，2022，41（1）：28-35.（in Chinese ）
［52］ SHU T W，YIN X E，XIONG Q Y，et al.Lift-CM：an integrated lift-heater centrifugal microfluidic platform for point-of-care pathogen nucleic acid detection using isothermal amplification and CRISPR/Cas 12a［J］.Biosensors and Bioelectronics，2025，274：117178.
［53］ GHRERA A S，PANDEY C M，MALHOTRA B D.Multiwalled carbon nanotube modified microfluidic-based biosensor chip for nucleic acid detection ［J］.Sensors and Actuators B：Chemical，2018，266：329-336.
［54］ LI X T，WANG T Y，LIU X H，et al.Advances of engineered microfluidic biosensors via CRISPR/Cas in bacteria and virus monitoring ［J］.Chemical Engineering Journal，2024，491：152038.
［55］ BOEHRINGER H R，O’FARRELL B J.Lateral flow assays in infectious disease diagnosis ［J］.Clinical Chemistry，2022，68（1）：52-58.
［56］ ZENG M C，KE Y Q，ZHUANG Z Y，et al.Harnessing multiplex crRNA in the CRISPR/Cas 12a system enables an amplification-free DNA diagnostic platform for ASFV detection ［J］.Analytical Chemistry，2022，94（30）：10805 -10812.
［57］ WANG D，WANG X D，YE F D，et al.An integrated amplification-free digital CRISPR/Cas-assisted assay for single molecule detection of RNA ［J］.ACS Nano，2023，17（8）：7250 -7256.
［58］ FU R J，XIANYU Y L.Gold nanomaterials-implemented CRISPR-Cas systems for biosensing ［J］.Small，2023，19（21）：2300057.