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Article

A Propidium Monoazide (PMAxx)-Droplet Digital PCR (ddPCR) for the Detection of Viable Burkholderia cepacia Complex in Nuclease-Free Water and Antiseptics

1
Division of Microbiology, National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, AR 72079, USA
2
Department of Natural Sciences, Albany State University, Albany, GA 31705, USA
3
Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Beltsville, MD 20993, USA
4
Office of Pharmaceutical Quality, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD 20993, USA
*
Author to whom correspondence should be addressed.
Present address: Eagle Analytical Services, Houston, TX 77099, USA.
Academic Editor: Maurizio Ciani
Microorganisms 2022, 10(5), 943; https://doi.org/10.3390/microorganisms10050943
Received: 5 April 2022 / Revised: 26 April 2022 / Accepted: 28 April 2022 / Published: 30 April 2022
Pharmaceutical products contaminated with Burkholderia cepacia complex (BCC) strains constitute a serious health issue for susceptible individuals. New detection methods to distinguish DNA from viable cells are required to ensure pharmaceutical product quality and safety. In this study, we have assessed a droplet digital PCR (ddPCR) with a variant propidium monoazide (PMAxx) for selective detection of live/dead BCC cells in autoclaved nuclease-free water after 365 days, in 0.001% chlorhexidine gluconate (CHX), and in 0.005% benzalkonium chloride (BZK) solutions after 184 days. Using 10 μM PMAxx and 5 min light exposure, a proportion of dead BCC was quantified by ddPCR. The detection limit of culture-based method was 104 CFU/mL, equivalent to 9.7 pg/μL for B. cenocepacia J2315, while that of ddPCR was 9.7 fg/μL. The true positive rate from nuclease-free water and CHX using PMAxx-ddPCR assay was 60.0% and 38.3%, respectively, compared to 85.0% and 74.6% without PMAxx (p < 0.05), respectively. However, in BZK-treated cells, no difference in the detection rate was observed between the ddPCR assay on samples treated with PMAxx (67.1%) and without PMAxx (63.3%). This study shows that the PMAxx-ddPCR assay provides a better tool for selective detection of live BCC cells in non-sterile pharmaceutical products. View Full-Text
Keywords: Burkholderia cepacia complex; propidium monoazide (PMAxx); droplet digital polymerase chain reaction (ddPCR); nuclease-free water; antiseptics Burkholderia cepacia complex; propidium monoazide (PMAxx); droplet digital polymerase chain reaction (ddPCR); nuclease-free water; antiseptics
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MDPI and ACS Style

Daddy Gaoh, S.; Kweon, O.; Lee, Y.-J.; Hussong, D.; Marasa, B.; Ahn, Y. A Propidium Monoazide (PMAxx)-Droplet Digital PCR (ddPCR) for the Detection of Viable Burkholderia cepacia Complex in Nuclease-Free Water and Antiseptics. Microorganisms 2022, 10, 943. https://doi.org/10.3390/microorganisms10050943

AMA Style

Daddy Gaoh S, Kweon O, Lee Y-J, Hussong D, Marasa B, Ahn Y. A Propidium Monoazide (PMAxx)-Droplet Digital PCR (ddPCR) for the Detection of Viable Burkholderia cepacia Complex in Nuclease-Free Water and Antiseptics. Microorganisms. 2022; 10(5):943. https://doi.org/10.3390/microorganisms10050943

Chicago/Turabian Style

Daddy Gaoh, Soumana, Ohgew Kweon, Yong-Jin Lee, David Hussong, Bernard Marasa, and Youngbeom Ahn. 2022. "A Propidium Monoazide (PMAxx)-Droplet Digital PCR (ddPCR) for the Detection of Viable Burkholderia cepacia Complex in Nuclease-Free Water and Antiseptics" Microorganisms 10, no. 5: 943. https://doi.org/10.3390/microorganisms10050943

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