Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction
To eliminate the influences of excipients and interference of dead bacterial DNA on the detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis capsules, a polymerase chain reaction (PCR) method with high sensitivity and...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2022-09-01
|
| Series: | Frontiers in Microbiology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2022.996794/full |
| _version_ | 1818480251859107840 |
|---|---|
| author | Xiaoling Zheng Yinhuan Wang WanZi Gong WanZi Gong Qianru Cai Jue Li Jiequn Wu |
| author_facet | Xiaoling Zheng Yinhuan Wang WanZi Gong WanZi Gong Qianru Cai Jue Li Jiequn Wu |
| author_sort | Xiaoling Zheng |
| collection | DOAJ |
| description | To eliminate the influences of excipients and interference of dead bacterial DNA on the detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis capsules, a polymerase chain reaction (PCR) method with high sensitivity and specificity was established. By combining bromide with propidium monoazide (PMA) -real-time quantitative PCR (qPCR) with microporous membrane filtration, excipients were removed, the filtrate was collected, and the bacteria were enriched using the centrifugal method. The optimal PMA working concentration, dark incubation time, and exposure time were determined. Specific E. coli, P. aeruginosa, S. paratyphoid B, and S. dysentery primers were selected to design different probes and a multiplex qPCR reaction system was established. The PMA-qPCR method was verified using different concentrations of dead and live bacteria. This method is efficient and accurate and can be widely applied to the detection of aforementioned pathogenic bacterial strains in live Bacillus licheniformis products. |
| first_indexed | 2024-12-10T11:20:56Z |
| format | Article |
| id | doaj.art-d063644a650f456681765d63d1a80e30 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-10T11:20:56Z |
| publishDate | 2022-09-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-d063644a650f456681765d63d1a80e302022-12-22T01:50:56ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2022-09-011310.3389/fmicb.2022.996794996794Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reactionXiaoling Zheng0Yinhuan Wang1WanZi Gong2WanZi Gong3Qianru Cai4Jue Li5Jiequn Wu6National Medical Products Administration (NMPA) Key Laboratory for Testing and Risk Warning of Pharmaceutical Microbiology, Key Laboratory of Drug Contacting Materials Quality Control of Zhejiang Provincial, Zhejiang Institute for Food and Drug Control, Hangzhou, ChinaNational Medical Products Administration (NMPA) Key Laboratory for Testing and Risk Warning of Pharmaceutical Microbiology, Key Laboratory of Drug Contacting Materials Quality Control of Zhejiang Provincial, Zhejiang Institute for Food and Drug Control, Hangzhou, ChinaNational Medical Products Administration (NMPA) Key Laboratory for Testing and Risk Warning of Pharmaceutical Microbiology, Key Laboratory of Drug Contacting Materials Quality Control of Zhejiang Provincial, Zhejiang Institute for Food and Drug Control, Hangzhou, ChinaSchool of Pharmacy, China Pharmaceutical University, Nanjing, ChinaCollaborative Innovation Center of Yangtze River Delta Region Green Pharmaceutical, Zhejiang University of Technology, Hangzhou, ChinaNational Medical Products Administration (NMPA) Key Laboratory for Testing and Risk Warning of Pharmaceutical Microbiology, Key Laboratory of Drug Contacting Materials Quality Control of Zhejiang Provincial, Zhejiang Institute for Food and Drug Control, Hangzhou, ChinaCollaborative Innovation Center of Yangtze River Delta Region Green Pharmaceutical, Zhejiang University of Technology, Hangzhou, ChinaTo eliminate the influences of excipients and interference of dead bacterial DNA on the detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis capsules, a polymerase chain reaction (PCR) method with high sensitivity and specificity was established. By combining bromide with propidium monoazide (PMA) -real-time quantitative PCR (qPCR) with microporous membrane filtration, excipients were removed, the filtrate was collected, and the bacteria were enriched using the centrifugal method. The optimal PMA working concentration, dark incubation time, and exposure time were determined. Specific E. coli, P. aeruginosa, S. paratyphoid B, and S. dysentery primers were selected to design different probes and a multiplex qPCR reaction system was established. The PMA-qPCR method was verified using different concentrations of dead and live bacteria. This method is efficient and accurate and can be widely applied to the detection of aforementioned pathogenic bacterial strains in live Bacillus licheniformis products.https://www.frontiersin.org/articles/10.3389/fmicb.2022.996794/fullpropidium monoazideqPCRprobioticsBacillus licheniformis live capsuleSalmonella paratyphoid BShigella dysentery |
| spellingShingle | Xiaoling Zheng Yinhuan Wang WanZi Gong WanZi Gong Qianru Cai Jue Li Jiequn Wu Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction Frontiers in Microbiology propidium monoazide qPCR probiotics Bacillus licheniformis live capsule Salmonella paratyphoid B Shigella dysentery |
| title | Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction |
| title_full | Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction |
| title_fullStr | Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction |
| title_full_unstemmed | Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction |
| title_short | Detection of Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphoid B, and Shigella dysentery in live Bacillus licheniformis products using propidium monoazide-real-time-quantitative polymerase chain reaction |
| title_sort | detection of escherichia coli pseudomonas aeruginosa salmonella paratyphoid b and shigella dysentery in live bacillus licheniformis products using propidium monoazide real time quantitative polymerase chain reaction |
| topic | propidium monoazide qPCR probiotics Bacillus licheniformis live capsule Salmonella paratyphoid B Shigella dysentery |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2022.996794/full |
| work_keys_str_mv | AT xiaolingzheng detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT yinhuanwang detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT wanzigong detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT wanzigong detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT qianrucai detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT jueli detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction AT jiequnwu detectionofescherichiacolipseudomonasaeruginosasalmonellaparatyphoidbandshigelladysenteryinlivebacilluslicheniformisproductsusingpropidiummonoaziderealtimequantitativepolymerasechainreaction |