Citation: ZOU Jing-Jing, LI Zhu-Jun, LIU Yun-Fan, OUYANG Xiu-Yun, WANG Yu, CAI Dong-Yang, HE Xiao-Wei, LIU Da-Yu. Integrated Microfluidic Droplet Digital Isothermal Amplification Enabled Rapid Detection of Urinary Tract Infection Bacteria[J]. Chinese Journal of Analytical Chemistry, ;2022, 50(8): 1158-1167,1187. doi: 10.19756/j.issn.0253-3820.221085 shu

Integrated Microfluidic Droplet Digital Isothermal Amplification Enabled Rapid Detection of Urinary Tract Infection Bacteria

1.
College of Food Science and Engineering, South China University of Technology, Guangzhou 510640, China;
2.
Department of Clinical Laboratory, The Second Affiliated Hospital of South China University of Technology, Guangzhou 510180, China;
3.
School of Mathematics and Computing Science, Guilin University of Electronic Technology, Guilin 541004, China

Corresponding author: HE Xiao-Wei, fexwhe@scut.edu.cn
Received Date: 21 February 2022
Revised Date: 10 May 2022

Fund Project: Supported by the China Postdoctoral Science Foundation (Nos.2020M682663, 2021M690780), the Natural Science Foundation of Guangdong Province,China (Nos.2020A1515010935, 2020A1515010754, 2020A1515110853, 2021A1515010182), the Science Foundation of Guangzhou First People's Hospital (No.PT81871726) and the Guangdong Provincial Key Laboratory of Point-of-care Testing (POCT) (No.2021B1212050016).

Digital nucleic acid analysis is a favorable tool for rapid molecular detection of pathogens due to its absolute quantification capability without the necessity of standard curves. However, current digital nucleic acid analysis platforms commonly perform the workflow with discrete facilities, which complicates the operation process and lengthens the turnaround time, therefore restricting its adoption in resource-limited settings. To address this issue, an integrated microfluidic droplet digital isothermal amplification system was developed in this work. This system integrated sequential droplets-based nucleic acid extraction unit, syringe-vacuum actuated droplet generation unit, and droplet digital loop-mediated isothermal amplification (LAMP) unit, achieving digital analysis of bacterial nucleic acid within 1.5 h in an integrated manner. The nucleic acid extraction unit could complete E.coli genomic DNA extraction with the efficiency of 93.68%±32.38%, the droplet generation unit could produce 20000 droplets in 4 min with the relative standard deviation (RSD) less than 10%. The LAMP reaction could be performed with a linear dynamic range of 4 orders of magnitude (2.36×10⁴-1.71×1 0⁷ CFU/mL). Compared to traditional culture method, the results obtained from analysis of E.coli in UTI clinical samples (n=13) showed that both the detection sensitivity and specificity of this digital nucleic acid analysis method were 100% (K_appa=1, p<0.01). With the advantages of accurate quantification and ease of operation, this integrated microfluidic droplet digital isothermal amplification system was expected to be a favorable tool for rapid point-of-care detection of pathogens in resource-limited settings.
- Microfluidic chip,
- Integration,
- Rapid point-of-care detection,
- Droplet digital nucleic acid testing,
- Urinary tract infection
1. [1]
  OZTURK R, MURT A. World J. Urol., 2020, 38(11):2669-2679.
2. [2]
  FLORES-MIRELES A L, WALKER J N, CAPARON M, HULTGREN S J. Nat. Rev. Microbiol., 2015, 13(5):269-284.
3. [3]
  CHAND K S, KAPOOR P. Homeopathy, 2020, 109(2):97-106.
4. [4]
  PRICE T K, DUNE T, HILT E E, THOMAS-WHITE K J, KLIETHERMES S, BRINCAT C, BRUBAKER L, WOLFE A J, MUELLER E R, SCHRECKENBERGER P C. J. Clin. Microbiol., 2016, 54(5):1216-1222.
5. [5]
  IPE D S, HORTON E, ULETT G C. Front. Cell. Infect. Microbiol., 2016, 6:14.
6. [6]
  CRAW P, BALACHANDRAN W. Lab Chip, 2012, 12(14):2469-2486.
7. [7]
  TRINH T N D, LEE N Y. Lab Chip, 2018, 18(16):2369-2377.
8. [8]
  KAYMAZ S V, ERGENC A F, AYTEKIN A O, LUCAS S J, ELITAS M. Biotechnol. Bioeng., 2022, 119(3):994-1003.
9. [9]
  XIANG X, SHANG Y, ZHANG J, DING Y, WU Q. TrAC, Trends Anal. Chem., 2022, 149:116568.
10. [10]
  BASU A S. SLAS Technol., 2017, 22(4):369-386.
11. [11]
  YEH E C, FU C C, HU L, THAKUR R, FENG J, LEE L P. Sci. Adv., 2017, 3(3):e1501645.
12. [12]
  RANE T D, CHEN L, ZEC H C, WANG T H. Lab Chip, 2015, 15(3):776-782.
13. [13]
14. [14]
  GENG Z, LI S, ZHU L, CHENG Z, JIN M, LIU B, GUO Y, LIU P. Anal. Chem., 2020, 92(10):7240-7248.
15. [15]
  YIN J, ZOU Z, HU Z, ZHANG S, ZHANG F, WANG B, LV S, MU Y. Lab Chip, 2020, 20(5):979-986.
16. [16]
  SHU B, LI Z, YANG X, XIAO F, LIN D, LEI X, XU B, LIU D. Chem. Commun., 2018, 54(18):2232-2235.
17. [17]
  SHU B, LIN L, WU B, HUANG E, WANG Y, LI Z, HE H, LEI X, XU B, LIU D. Biosens. Bioelectron., 2021, 181:113145.
18. [18]
19. [19]
  HILL J, BERIWAL S, CHANDRA I, PAUL V K, KAPIL A, SINGH T, WADOWSKY R M, SINGH V, GOYAL A, JAHNUKAINEN T, JOHNSON J R, TARR P I, VATS A. J. Clin. Microbiol., 2008, 46(8):2800-2804.
20. [20]
  HU F, LI J, ZHANG Z M, LI M, ZHAO S H, LI Z P, PENG N C. Anal. Chem., 2020, 92(2):2258-2265.
21. [21]
  XU B L, DU Y, LIN J Q, QI M Y, SHU B W, WEN X X, LIANG G T, CHEN B, LIU D Y. Anal. Chem., 2016, 88(23):11593-11600.
22. [22]
  REN K N, ZHOU J H, WU H K. Acc. Chem. Res., 2013, 46(11):2396-2406.
23. [23]
  YUAN H, CHAO Y, SHUM H C. Small, 2020, 16(9):e1904469.
24. [24]
25. [25]
  WITTE A K, MESTER P, FISTER S, WITTE M, SCHODER D, ROSSMANITH P. PLoS One, 2016, 11(12):e0168179.
26. [26]
  SEYRIG G, STEDTFELD R D, TOURLOUSSE D M, AHMAD F, TOWERY K, CUPPLES A M, TIEDJE J M, HASHSHAM S A. J. Microbiol. Methods, 2015, 119:223-227.
27. [27]
  HOLST-JENSEN A, CRESPO T, SIMPLÍCIO A, RICHL P, WELSCHE M, DOBNIK D, DREO T. Minimum Performance Parameters (MPPs) and Associated Acceptance Values (AAVs) for Nucleic Acid Amplification Methods (DECATHLON deliverable report D6.1):https://www.researchgate.net/publication/277943953, 2014:18.
28. [28]
  RICCHI M, BERTASIO C, BONIOTTI M B, VICARI N, RUSSO S, TILOLA M, BELLOTTI M A, BERTASI B. Front. Microbiol., 2017, 8:1174.
29. [29]
1. [1]
  Min Gu , Huiwen Xiong , Liling Liu , Jilie Kong , Xueen Fang . Rapid Quantitative Detection of Procalcitonin by Microfluidics: An Instrumental Analytical Chemistry Experiment. University Chemistry, 2024, 39(4): 87-93. doi: 10.3866/PKU.DXHX202310120
2. [2]
  Di Yang , Jiayi Wei , Hong Zhai , Xin Wang , Taiming Sun , Haole Song , Haiyan Wang . Rapid Detection of SARS-CoV-2 Using an Innovative “Magic Strip”. University Chemistry, 2024, 39(4): 373-381. doi: 10.3866/PKU.DXHX202312023
3. [3]
  Qilong Fang , Yiqi Li , Jiangyihui Sheng , Quan Yuan , Jie Tan . Magical Pesticide Residue Detection Test Strips: Aptamer-based Lateral Flow Test Strips for Organophosphorus Pesticide Detection. University Chemistry, 2024, 39(5): 80-89. doi: 10.3866/PKU.DXHX202310004
4. [4]
  Qin Tu , Anju Tao , Tongtong Ma , Jinyi Wang . Innovative Experimental Teaching of Escherichia coli Detection Based on Paper Chip. University Chemistry, 2024, 39(6): 271-277. doi: 10.3866/PKU.DXHX202309062
5. [5]
  Hongwei Ma , Fang Zhang , Hui Ai , Niu Zhang , Shaochun Peng , Hui Li . Integrated Crystallographic Teaching with X-ray,TEM and STM. University Chemistry, 2024, 39(3): 5-17. doi: 10.3866/PKU.DXHX202308107
6. [6]
  Xiaoyan Wang , Yan Qi , Lin Tang , Shuwen Wang , Huiling Wen , Hongtao Gao . Improvement of the Quality Construction of Basic Chemistry Experimental Teaching Center under the Background of Education Digitization. University Chemistry, 2024, 39(7): 40-48. doi: 10.12461/PKU.DXHX202404124
7. [7]
  Yan Su , Yuzhen Pan , Fuping Tian , Xiuyun Wang , Tieqi Xu , Yongce Zhang , Miao Cui , Wenfeng Jiang . Construction and Practice of the National Chemical Experimental Teaching Demonstration Center under the Background of Digital Education. University Chemistry, 2024, 39(7): 218-222. doi: 10.12461/PKU.DXHX202406001
8. [8]
  Simin Fang , Hong Wu , Wei Liu , Wei Wei , Hongyan Feng , Wan Li . Construction and Application of Teaching Resources for Inorganic and Analytical Chemistry Experimental Course in the Context of Digital Empowerment. University Chemistry, 2024, 39(10): 156-163. doi: 10.3866/PKU.DXHX202402053
9. [9]
  Bin Wang , Chuanli Qin , Naiying Fan , Zhibin Li , Zhibiao Zhu , Jiancong Liu , Shaoping Sun . Construction and Practice of Digital Platform and Teaching Resources of Chemistry Experimental Teaching Center: A Case Study of Provincial Demonstration Center for Experimental Chemistry Education (Heilongjiang University). University Chemistry, 2024, 39(7): 193-199. doi: 10.12461/PKU.DXHX202405116
10. [10]
  Hong LI , Xiaoying DING , Cihang LIU , Jinghan ZHANG , Yanying RAO . Detection of iron and copper ions based on gold nanorod etching colorimetry. Chinese Journal of Inorganic Chemistry, 2024, 40(5): 953-962. doi: 10.11862/CJIC.20230370
11. [11]
  Yang YANG , Pengcheng LI , Zhan SHU , Nengrong TU , Zonghua WANG . Plasmon-enhanced upconversion luminescence and application of molecular detection. Chinese Journal of Inorganic Chemistry, 2024, 40(5): 877-884. doi: 10.11862/CJIC.20230440
12. [12]
  Siyi ZHONG , Xiaowen LIN , Jiaxin LIU , Ruyi WANG , Tao LIANG , Zhengfeng DENG , Ao ZHONG , Cuiping HAN . Targeting imaging and detection of ovarian cancer cells based on fluorescent magnetic carbon dots. Chinese Journal of Inorganic Chemistry, 2024, 40(8): 1483-1490. doi: 10.11862/CJIC.20240093
13. [13]
  Xiaowei TANG , Shiquan XIAO , Jingwen SUN , Yu ZHU , Xiaoting CHEN , Haiyan ZHANG . A zinc complex for the detection of anthrax biomarker. Chinese Journal of Inorganic Chemistry, 2024, 40(10): 1850-1860. doi: 10.11862/CJIC.20240173
14. [14]
  Tongyu Zheng , Teng Li , Xiaoyu Han , Yupei Chai , Kexin Zhao , Quan Liu , Xiaohui Ji . A DIY pH Detection Agent Using Persimmon Extract for Acid-Base Discoloration Popularization Experiment. University Chemistry, 2024, 39(5): 27-36. doi: 10.3866/PKU.DXHX202309107
15. [15]
  Jinghan ZHANG , Guanying CHEN . Progress in the application of rare-earth-doped upconversion nanoprobes in biological detection. Chinese Journal of Inorganic Chemistry, 2024, 40(12): 2335-2355. doi: 10.11862/CJIC.20240249
16. [16]
  Jiakun BAI , Ting XU , Lu ZHANG , Jiang PENG , Yuqiang LI , Junhui JIA . A red-emitting fluorescent probe with a large Stokes shift for selective detection of hypochlorous acid. Chinese Journal of Inorganic Chemistry, 2024, 40(6): 1095-1104. doi: 10.11862/CJIC.20240002
17. [17]
  Hao BAI , Weizhi JI , Jinyan CHEN , Hongji LI , Mingji LI . Preparation of Cu₂O/Cu-vertical graphene microelectrode and detection of uric acid/electroencephalogram. Chinese Journal of Inorganic Chemistry, 2024, 40(7): 1309-1319. doi: 10.11862/CJIC.20240001
18. [18]
  Mengyao Shi , Kangle Su , Qingming Lu , Bin Zhang , Xiaowen Xu . Determination of Potassium Content in Tobacco Stem Ash by Flame Atomic Absorption Spectroscopy. University Chemistry, 2024, 39(10): 255-260. doi: 10.12461/PKU.DXHX202404105
19. [19]
  Kuaibing Wang , Honglin Zhang , Wenjie Lu , Weihua Zhang . Experimental Design and Practice for Recycling and Nickel Content Detection from Waste Nickel-Metal Hydride Batteries. University Chemistry, 2024, 39(11): 335-341. doi: 10.12461/PKU.DXHX202403084
20. [20]
  Junjie Zhang , Yue Wang , Qiuhan Wu , Ruquan Shen , Han Liu , Xinhua Duan . Preparation and Selective Separation of Lightweight Magnetic Molecularly Imprinted Polymers for Trace Tetracycline Detection in Milk. University Chemistry, 2024, 39(5): 251-257. doi: 10.3866/PKU.DXHX202311084

Get Citation

PDF

XML

Metrics

PDF Downloads(19)
Abstract views(638)
HTML views(121)

通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142