Ying Chen1, Linyan Zhang1, Ling Xu1, Xinjian Guo1, Huan Yang1, Linlin Zhuang2,3, Ying Li1, Zhenzhen Wang1, Bing Gu1. 1. School of Medical Technology, Xuzhou Medical University, Xuzhou 221004, China. 2. School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China. 3. Nanjing Nanoeast biotech Co., Ltd., Nanjing 210000, China.
Abstract
BACKGROUND: Bacillary dysentery caused by Shigella genus is a major cause of morbidity and mortality worldwide. In China, the popular strain was mainly Shigella flexneri (S. flexneri). Therefore, fluorescent microspheres (FMs)-based immunochromatographic test strip (ICTS), as a novel, reliable, sensitive and uncomplicated method, was evaluated to detect S. flexneri. METHODS: Sixty-three clinical samples of S. flexneri were collected in this paper. Polymerase chain reaction (PCR) combined with FMs-ICTS based on magnetic purification assay was developed for the quantitative detection of Shigella. And the genus-specific gene of ipaH and drug resistant gene of CTX-M-9 from Shigella were selected to investigate the potential of this new method. The sensitivity and specificity of this method were demonstrated by classical microbiological methods (API Coryne System), PCR assay based on agarose gel electrophoresis (PCR-GE) and the real-time fluorescent quantitative PCR (RTFQ-PCR) method. RESULTS: Under optimized conditions, the lower detection limits of PCR-ICTS, PCR-GE and RTFQ-PCR were 2.5×10-7, 2.5×10-5 and the 3.2×10-7 ng/µL, respectively. Experiments demonstrated the PCR-ICTS has a diagnostic agreement of 100% with conventional PCR and RTFQ-PCR on detection of clinical samples and could correctly recognize Shigella and non-Shigella from different microbial samples. After the purification of PCR products with Silicon coated magnetic nanoparticles (Si-MNPs), the false positive results were removed because of the strong screening ability of the purification process. Our results showed that FM-based ICTS was promising for measurable and sensitive detection of S. flexneri within 3 h. CONCLUSIONS: The results from immunochromatographic test were agreement with those from API Coryne system and RTFQ-PCR. Hence, this developed method might be useful for screening and monitoring clinical sample of S. flexneri, due to its speed, non-poisonous, simplicity and low-cost and helpful for promoting the prevention and control of communicable diseases caused by enteric pathogens such as S. flexneri. 2019 Annals of Translational Medicine. All rights reserved.
BACKGROUND: Bacillary dysentery caused by Shigella genus is a major cause of morbidity and mortality worldwide. In China, the popular strain was mainly Shigella flexneri (S. flexneri). Therefore, fluorescent microspheres (FMs)-based immunochromatographic test strip (ICTS), as a novel, reliable, sensitive and uncomplicated method, was evaluated to detect S. flexneri. METHODS: Sixty-three clinical samples of S. flexneri were collected in this paper. Polymerase chain reaction (PCR) combined with FMs-ICTS based on magnetic purification assay was developed for the quantitative detection of Shigella. And the genus-specific gene of ipaH and drug resistant gene of CTX-M-9 from Shigella were selected to investigate the potential of this new method. The sensitivity and specificity of this method were demonstrated by classical microbiological methods (API Coryne System), PCR assay based on agarose gel electrophoresis (PCR-GE) and the real-time fluorescent quantitative PCR (RTFQ-PCR) method. RESULTS: Under optimized conditions, the lower detection limits of PCR-ICTS, PCR-GE and RTFQ-PCR were 2.5×10-7, 2.5×10-5 and the 3.2×10-7 ng/µL, respectively. Experiments demonstrated the PCR-ICTS has a diagnostic agreement of 100% with conventional PCR and RTFQ-PCR on detection of clinical samples and could correctly recognize Shigella and non-Shigella from different microbial samples. After the purification of PCR products with Silicon coated magnetic nanoparticles (Si-MNPs), the false positive results were removed because of the strong screening ability of the purification process. Our results showed that FM-based ICTS was promising for measurable and sensitive detection of S. flexneri within 3 h. CONCLUSIONS: The results from immunochromatographic test were agreement with those from API Coryne system and RTFQ-PCR. Hence, this developed method might be useful for screening and monitoring clinical sample of S. flexneri, due to its speed, non-poisonous, simplicity and low-cost and helpful for promoting the prevention and control of communicable diseases caused by enteric pathogens such as S. flexneri. 2019 Annals of Translational Medicine. All rights reserved.
Entities:
Keywords:
Fluorescent microsphere (FM); Shigella flexneri (S. flexneri); detection; immunochromatographic test strip (ICTS); purification
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