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Multiple Cross Displacement Amplification-a more applicable technique in detecting Pseudomonas Aeruginosa of Ventilator-associated Pneumonia ( VAP)
Bin Chen
Xiamen Port Clinic of Xiamen Customs
Corresponding Author
ORCiD: https://orcid.org/0000-0002-3152-4021
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Early and rapid identification of Pseudomonas aeruginosa (P. aeruginosa) in patients with suspected ventilator-associated pneumonia (VAP) provides theoretical clinical advantages in therapeutic optimization strategies.
Methods: The P. aeruginosa-multiple cross displacement amplification (PA-MCDA) assay was conducted at an isothermal temperature during the amplification stage, and products were visually detected by color changes. The entire process was completed within 1 h. A total of 77 strains, including P. aeruginosa species and various other species of non-P. aeruginosa were used to evaluate PA-MCDA assays. Bronchoalveolar lavage fluid (BALF) of suspected VAP patients were examined by the MCDA assay.
Results: The MCDA assay exhibited a 100 percent analytical specificity in detecting PA from all 77 strains, and the limit of detection were as low as 100 fg DNA per reaction. A temperature of 65ºC was recommended as standard during the amplification stage. The agreement between PA-MCDA and bacteria culture was 91.18% (κ= 0.787; p =0.000) in identification of P. aeruginosa in BALF from suspected VAP. The PA-MCDA assay showed values of 92.31%, 90.78%, 77.41% and 97.18% for sensitivity, specificity, positive predictive value and negative predictive value, respectively. PA-MCDA had higher detective rate of P. aeruginosa than bacteria culture in patients with antipseudomonal therapy.
Conclusions: The instrument-free platform of the MCDA assay makes it a simple, rapid and applicable procedure for “on-site” diagnosis and point-of-care testing for the presence of P. aeruginosa without the need for specific bacterial culture.
Keywords
Pseudomonas aeruginosa, multiple cross displacement amplification, ventilated-associated pneumonia, bronchoalveolar lavage fluid, Limit of Detection
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CURRENT STATUS: Under Review
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Reviewer #2 agreed
On 16 May, 2020
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Review #1 received
Received 16 May, 2020
Editor assigned
On 13 May, 2020
Reviewers invited
Invitations sent on 13 May, 2020
Reviewer #1 agreed
On 13 May, 2020
Submission checks complete
On 12 May, 2020
Editor invited
On 12 May, 2020
No comments provided
Editorial decision: Major revision
On 13 Apr, 2020
Review #1 received
Received 12 Apr, 2020
Reviewer #2 agreed
On 09 Apr, 2020
Review #2 received
Received 09 Apr, 2020
Reviewers invited
Invitations sent on 08 Apr, 2020
Reviewer #1 agreed
On 08 Apr, 2020
Editor assigned
On 08 Apr, 2020
Submission checks complete
On 07 Apr, 2020
Editor invited
On 07 Apr, 2020
No comments provided
Editorial decision: Major revision
On 28 Feb, 2020
Reviewer #2 agreed
On 22 Feb, 2020
Review #2 received
Received 22 Feb, 2020
Review #1 received
Received 21 Feb, 2020
Reviewer #1 agreed
On 17 Feb, 2020
Reviewers invited
Invitations sent on 16 Feb, 2020
Editor assigned
On 10 Feb, 2020
Editor invited
On 09 Feb, 2020
Submission checks complete
On 06 Feb, 2020
First submitted
On 04 Feb, 2020
Metrics
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PDF Downloads: ...
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Subject Areas
Critical Care & Emergency Medicine
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This preprint is under consideration at Critical Care. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research
Multiple Cross Displacement Amplification-a more applicable technique in detecting Pseudomonas Aeruginosa of Ventilator-associated Pneumonia ( VAP)
Bin Chen
Xiamen Port Clinic of Xiamen Customs
Corresponding Author
ORCiD: https://orcid.org/0000-0002-3152-4021
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Review
Version 3
Posted 26 May, 2020
No community comments so far
Reviewer #2 agreed
On 16 May, 2020
Review #2 received
Received 16 May, 2020
Review #1 received
Received 16 May, 2020
Editor assigned
On 13 May, 2020
Reviewers invited
Invitations sent on 13 May, 2020
Reviewer #1 agreed
On 13 May, 2020
Submission checks complete
On 12 May, 2020
Editor invited
On 12 May, 2020
No comments provided
Editorial decision: Major revision
On 13 Apr, 2020
Review #1 received
Received 12 Apr, 2020
Reviewer #2 agreed
On 09 Apr, 2020
Review #2 received
Received 09 Apr, 2020
Reviewers invited
Invitations sent on 08 Apr, 2020
Reviewer #1 agreed
On 08 Apr, 2020
Editor assigned
On 08 Apr, 2020
Submission checks complete
On 07 Apr, 2020
Editor invited
On 07 Apr, 2020
No comments provided
Editorial decision: Major revision
On 28 Feb, 2020
Reviewer #2 agreed
On 22 Feb, 2020
Review #2 received
Received 22 Feb, 2020
Review #1 received
Received 21 Feb, 2020
Reviewer #1 agreed
On 17 Feb, 2020
Reviewers invited
Invitations sent on 16 Feb, 2020
Editor assigned
On 10 Feb, 2020
Editor invited
On 09 Feb, 2020
Submission checks complete
On 06 Feb, 2020
First submitted
On 04 Feb, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Critical Care & Emergency Medicine
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Early and rapid identification of Pseudomonas aeruginosa (P. aeruginosa) in patients with suspected ventilator-associated pneumonia (VAP) provides theoretical clinical advantages in therapeutic optimization strategies.
Methods: The P. aeruginosa-multiple cross displacement amplification (PA-MCDA) assay was conducted at an isothermal temperature during the amplification stage, and products were visually detected by color changes. The entire process was completed within 1 h. A total of 77 strains, including P. aeruginosa species and various other species of non-P. aeruginosa were used to evaluate PA-MCDA assays. Bronchoalveolar lavage fluid (BALF) of suspected VAP patients were examined by the MCDA assay.
Results: The MCDA assay exhibited a 100 percent analytical specificity in detecting PA from all 77 strains, and the limit of detection were as low as 100 fg DNA per reaction. A temperature of 65ºC was recommended as standard during the amplification stage. The agreement between PA-MCDA and bacteria culture was 91.18% (κ= 0.787; p =0.000) in identification of P. aeruginosa in BALF from suspected VAP. The PA-MCDA assay showed values of 92.31%, 90.78%, 77.41% and 97.18% for sensitivity, specificity, positive predictive value and negative predictive value, respectively. PA-MCDA had higher detective rate of P. aeruginosa than bacteria culture in patients with antipseudomonal therapy.
Conclusions: The instrument-free platform of the MCDA assay makes it a simple, rapid and applicable procedure for “on-site” diagnosis and point-of-care testing for the presence of P. aeruginosa without the need for specific bacterial culture.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5