1.Development of multiplex loop-mediated isothermal amplification (mLAMP) for detection of Salmonella,Vibrio parahaemolyticus and Listeria monocytogenes
Ningwei LIU ; Dayang ZOU ; Derong DONG ; Zhan YANG ; Simei HUANG ; Xiaoming HE ; Da AO ; Wei LIU ; Liuyu HUANG
Military Medical Sciences 2016;40(9):767-772
Objective To establish a multiplex loop-mediated isothermal amplification(mLAMP)method for simultaneous detection of Salmonella,Vibrio parahaemolyticus (VPH)and Listeria monocytogenes (LM).Methods Three sets of mLAMP primers were designed to specifically target bcfD of Salmonella and tlh of VPH and iap of LM.The respective single LAMP assay of the three kinds of bacteria was developed,and the ratio of primer concentration was optimized to develop a multiplex LAMP system.The specificity and sensitivity of multiplex LAMP were observed.Results Turbidity monitoring results in real time suggests that the mLAMP was highly specific and amplification could be obtained within 45 min under isothermal conditions.The sensitivity of this mLAMP was found to be 300 fg/μl genomic DNAs for Salmonella and 4.2 pg/μl for VPH and 4.5 pg/μl for LM,which was consistent with conventional PCR.Conclusion The mLAMP described can potentially facilitate simultaneous detection of three kinds of bacteria in a large number of food samples, which could be used as a primary screening method and as a supplement to classical detection methods.
2.Rapid detection of influenza A/H1N1 virus by polymerase spiral reaction
Wen MA ; Derong DONG ; Dayang ZOU ; Ningwei LIU ; Xiaoming HE ; Da AO ; Zhan YANG ; Simo HUANG ; Yaqing XU ; Wei LIU ; Liuyu HUANG
Military Medical Sciences 2017;41(6):449-452
Objective To establish a polymerase spiral reaction (PSR) method for rapid detection of influenza A/H1N1 virus.Methods Six sets of primers were designed for influenza A/H1N1 virus HA gene, and the results were determined with real time kinetic turbidimetric assay and colorimetry method.Results and Conclusion The best primers were selected from six sets of primers, and the best temperature was determined as 65 degrees Celsius.Further experiments showed that the best primer had good specificity for detection of influenza A/H1N1 virus,without cross reactions with 14 other respiratory tract pathogenic nucleic acids.The sensitivity was up to 100 copies,and consistent with that of PCR.So a PSR method is established for rapid detection of the influenza A/H1N1 virus, which is simple, quick, highly specific and sensitive,and especially applicable to field and grass-roots units.
3. Expression of type 1 and type 2 cytokines from serum of coal miners and the evaluation in surveillance of coal workers' pneumoconiosis at earlier stage
Sanqiao YAO ; Ningwei YANG ; Feifei GUO ; Tianbang QIN ; Xiuping ZHU ; Zhigang DONG ; Zhichun LI ; Bijie JIANG ; Jingshun GAO ; Yongcheng YAO ; Guofu ZHANG ; Yang LIU ; Yao LU ; Haibin LI ; Jianfei SHUAI ; Yuping BAI ; Yulan JIN
Chinese Journal of Preventive Medicine 2018;52(11):1158-1163
Objectives:
To explore the expression regulation of type 1 and type 2 (Th1 and Th2) cytokines from serum of coal miners and the evaluation in surveillance of coal workers' pneumoconiosis, 630 coal miners were studied.
Methods:
A total of 90 male patients diagnosed as coal workers' pneumoconiosis (CWP) in a institute for occupational health and 19 male workers newly diagnosed as CWP patients was chosen as CWP group with simple random sampling method from a coal mine group from January 2013 to December in 2015. 180 male coal miners with abnormal but not diagnosed as CWP were selected as CWP suspected group with simple random sampling methods, meanwhile 180 male coal miners with normal chest X-ray photograph was as dust-exposed group by 1∶1 matched as age. And 161 healthy males accepted pre-employed examination were selected as control group, CWP suspected group, dust-exposed group and control group called as non-CWP group. According to screening test and diagnosis test, the basic information and occupational history of all subjects were collected, and cytokines including IL-1β, IL-8, IFN-γ, IL-6 and IL-10 of serum were detected. Receiver operator characteristic (ROC) curve was used to determine the optimal cutoff value of each cytokine. Area under curve (AUC), the validity and reliability were calculated and judged.
Results:
The average age of control group, dust-exposed group, CWP suspected group and CWP group were (27.4±5.0) , (43.4±10.7) , (48.2±6.2) , (64.7±7.0) years old, respectively. The median level of IL-1β, IL-8, IFN-γ and IL-6 in cases group (1 638.30, 2 099.49, 815.18,140.32 pg/ml) were higher than that of non-cases group (1 445.57, 1 402.26, 736.38, 95.73 pg/ml) (