Immunocapture Magnetic Beads Enhanced the LAMP-CRISPR/Cas12a Method for the Sensitive, Specific, and Visual Detection of Campylobacter jejuni.
Immunocapture Magnetic Beads Enhanced the LAMP-CRISPR/Cas12a Method for the Sensitive, Specific, and Visual Detection of Campylobacter jejuni.
复制标题
DOI：
10.3390/bios12030154
复制
发表时间：
2022-03-02
期刊：
影响因子：
--
通讯作者：
Wang H
中科院分区：
文献类型：
--
作者：
Li C;Chen X;Wen R;Ma P;Gu K;Li C;Zhou C;Lei C;Tang Y;Wang H
空肠弯曲杆菌（Campylobacter jejuni）是食源性传染病的重要病原之一，对食品安全和公共卫生构成了挑战。建立快速、准确、灵敏、简便的空肠梭菌检测方法，有助于早期诊断、早期干预和预防病原体传播。本研究建立了一种免疫捕获磁珠（ICB）增强环介导等温扩增（LAMP） CRISPR/Cas12a方法（ICB-LAMP-CRISPR/Cas12a），用于快速、目视检测空肠假丝酵母。采用ICB-LAMP- crispr /Cas12a方法，首先通过ICB捕获空肠C.，然后通过加热释放细菌基因组DNA并用于LAMP反应。LAMP反应后，将LAMP产物混合，用CRISPR/Cas12a裂解混合物检测。ICB-LAMP-CRISPR/Cas12a方法可在70 min内检测到至少8 CFU/mL的空肠假丝酵素。此外，该方法除ICB捕获外，还在封闭管中进行，从而消除了分离扩增前和扩增产物转移的需要，避免了气溶胶污染。ICB-LAMP-CRISPR/Cas12a方法通过对来自不同蛋鸡养殖场的31份大肠杆菌阳性粪便样本进行进一步验证。该方法是一种简便、快速、超灵敏、超特异、目视的无仪器诊断空肠梭菌的检测方法，在今后的工作中具有广泛的应用潜力。
Campylobacter jejuni is one of the most important causes of food-borne infectious disease, and poses challenges to food safety and public health. Establishing a rapid, accurate, sensitive, and simple detection method for C. jejuni enables early diagnosis, early intervention, and prevention of pathogen transmission. In this study, an immunocapture magnetic bead (ICB)-enhanced loop-mediated isothermal amplification (LAMP) CRISPR/Cas12a method (ICB-LAMP-CRISPR/Cas12a) was developed for the rapid and visual detection of C. jejuni. Using the ICB-LAMP-CRISPR/Cas12a method, C. jejuni was first captured by ICB, and the bacterial genomic DNA was then released by heating and used in the LAMP reaction. After the LAMP reaction, LAMP products were mixed and detected by the CRISPR/Cas12a cleavage mixture. This ICB-LAMP-CRISPR/Cas12a method could detect a minimum of 8 CFU/mL of C. jejuni within 70 min. Additionally, the method was performed in a closed tube in addition to ICB capture, which eliminates the need to separate preamplification and transfer of amplified products to avoid aerosol pollution. The ICB-LAMP-CRISPR/Cas12a method was further validated by testing 31 C. jejuni-positive fecal samples from different layer farms. This method is an all-in-one, simple, rapid, ultrasensitive, ultraspecific, visual detection method for instrument-free diagnosis of C. jejuni, and has wide application potential in future work.
登录
查看更多内容
DOI：
10.1073/pnas.2010196117
发表时间：
2020-10-13
影响因子：
11.1
作者：
Lee RA;Puig H;Nguyen PQ;Angenent-Mari NM;Donghia NM;McGee JP;Dvorin JD;Klapperich CM;Pollock NR;Collins JJ
通讯作者：
Collins JJ
影响因子：
6.7
作者：
Giallourou N;Medlock GL;Bolick DT;Medeiros PH;Ledwaba SE;Kolling GL;Tung K;Guerry P;Swann JR;Guerrant RL
通讯作者：
Guerrant RL
影响因子：
16.6
作者：
Ding X;Yin K;Li Z;Lalla RV;Ballesteros E;Sfeir MM;Liu C
通讯作者：
Liu C
影响因子：
4.6
作者：
Ewers EC;Anisowicz SK;Ferguson TM;Seronello SE;Barnhill JC;Lustik MB;Agee W 3rd;Washington MA;Nahid MA;Burnett MW;Bodhidatta L;Srijan A;Rukasiri S;Wassanarungroj P;Ruekit S;Nobthai P;Swierczewski BE;Lurchachaiwong W;Serichantalergs O;Ngauy V
通讯作者：
Ngauy V
从监视样品中快速检测到念珠菌的实时PCR分析的开发和验证。
影响因子：
9.4
作者：
Leach L;Zhu Y;Chaturvedi S
通讯作者：
Chaturvedi S