Multiplex Digital PCR-Based Development and Discussion of the Detection of Genetic Association Between and

Overview

Journal Infect Drug Resist

Publisher Dove Medical Press

Date 2024 May 28

PMID 38803520

Authors

Ciming Wu

Ming Ling

Qiong Chen

Hui Chai

Huan Chen

Affiliations

Soon will be listed here.

Abstract

Methicillin-resistant (MRSA) is a predominant nosocomial infection-causing bacteria. The aim of this study was to develop a novel single-bacteria multiplex digital PCR assays (SMD-PCR), which is capable of simultaneously detecting and discriminating Methicillin-sensitive (MSSA) and MRSA. This protocol employed TaqMan probes to detect and genes, with the latter being linked to methicillin resistance. A total of 72 samples from various specimen types were evaluated. The accuracy rates for the sputum samples, pus samples, swab samples, ear secretion samples, and catheter samples were 94.44%, 100%, 92%, 100%, and 100%, respectively. Our results showed that the clinical practicability of SMD-PCR has applicability to the rapid detection of MRSA without DNA extraction or bacterial culture, and can be utilized for the rapid detection of and the timely identification of MRSA in clinical samples, thereby providing an advanced platform for the rapid diagnosis of clinical MRSA infection.

Citing Articles

Advanced One-Pot RPA-CRISPR/Cas12a Reaction with Glycerol and Betaine for High-Sensitivity Diagnosis of -Carrying Strains in Clinical Samples.

Wang J, Wang D, Fan L, Ye X, Hu J, Wang X ACS Omega. 2025; 10(5):4599-4606.

PMID: 39959114 PMC: 11822479. DOI: 10.1021/acsomega.4c09078.

References

Pereira E, Schuenck R, Malvar K, Iorio N, Matos P, Olendzki A . Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus haemolyticus: methicillin-resistant isolates are detected directly in blood cultures by multiplex PCR. Microbiol Res. 2009; 165(3):243-9. DOI: 10.1016/j.micres.2009.03.003. View

Hiramatsu K, Hanaki H, Ino T, YABUTA K, Oguri T, Tenover F . Methicillin-resistant Staphylococcus aureus clinical strain with reduced vancomycin susceptibility. J Antimicrob Chemother. 1997; 40(1):135-6. DOI: 10.1093/jac/40.1.135. View

Palavecino E . Clinical, Epidemiologic, and Laboratory Aspects of Methicillin-Resistant Staphylococcus aureus Infections. Methods Mol Biol. 2019; 2069:1-28. DOI: 10.1007/978-1-4939-9849-4_1. View

Luo J, Li J, Yang H, Yu J, Wei H . Accurate Detection of Methicillin-Resistant Staphylococcus aureus in Mixtures by Use of Single-Bacterium Duplex Droplet Digital PCR. J Clin Microbiol. 2017; 55(10):2946-2955. PMC: 5625380. DOI: 10.1128/JCM.00716-17. View

Pannewick B, Baier C, Schwab F, Vonberg R . Infection control measures in nosocomial MRSA outbreaks-Results of a systematic analysis. PLoS One. 2021; 16(4):e0249837. PMC: 8026056. DOI: 10.1371/journal.pone.0249837. View

Brown J, Paladino J . Impact of rapid methicillin-resistant Staphylococcus aureus polymerase chain reaction testing on mortality and cost effectiveness in hospitalized patients with bacteraemia: a decision model. Pharmacoeconomics. 2010; 28(7):567-75. DOI: 10.2165/11533020-000000000-00000. View

Lee A, de Lencastre H, Garau J, Kluytmans J, Malhotra-Kumar S, Peschel A . Methicillin-resistant Staphylococcus aureus. Nat Rev Dis Primers. 2018; 4:18033. DOI: 10.1038/nrdp.2018.33. View

Trouillet-Assant S, Rasigade J, Lustig S, Lhoste Y, Valour F, Guerin C . Ward-specific rates of nasal cocolonization with methicillin-susceptible and -resistant Staphylococcus spp. and potential impact on molecular methicillin-resistant Staphylococcus aureus screening tests. J Clin Microbiol. 2013; 51(7):2418-20. PMC: 3697672. DOI: 10.1128/JCM.00491-13. View

Bridwell M, Bajaj S, Gress T, Hambuchen M, Clay T . Impact of MRSA polymerase chain reaction (PCR) wound swabs on antibiotic de-escalation in skin and soft tissue infections. Diagn Microbiol Infect Dis. 2022; 103(3):115722. DOI: 10.1016/j.diagmicrobio.2022.115722. View

10.

Kelley K, Cosman A, Belgrader P, Chapman B, Sullivan D . Detection of methicillin-resistant Staphylococcus aureus by a duplex droplet digital PCR assay. J Clin Microbiol. 2013; 51(7):2033-9. PMC: 3697713. DOI: 10.1128/JCM.00196-13. View

11.

Chen Y, Shi Y, Zhu W, You J, Yang J, Xie Y . Combining CRISPR-Cas12a-Based Technology and Metagenomics Next Generation Sequencing: A New Paradigm for Rapid and Full-Scale Detection of Microbes in Infectious Diabetic Foot Samples. Front Microbiol. 2021; 12:742040. PMC: 8529936. DOI: 10.3389/fmicb.2021.742040. View

12.

Kolman S, Arielly H, Paitan Y . Evaluation of single and double-locus real-time PCR assays for methicillin-resistant Staphylococcus aureus (MRSA) surveillance. BMC Res Notes. 2010; 3:110. PMC: 2877054. DOI: 10.1186/1756-0500-3-110. View

13.

de la Cruz Barron M, Kneis D, Elena A, Bagra K, Berendonk T, Klumper U . Quantification of the mobility potential of antibiotic resistance genes through multiplexed ddPCR linkage analysis. FEMS Microbiol Ecol. 2023; 99(4). PMC: 10054912. DOI: 10.1093/femsec/fiad031. View

14.

Bishr A, Abdelaziz S, Yahia I, Yassien M, Hassouna N, Aboshanab K . Association of Macrolide Resistance Genotypes and Synergistic Antibiotic Combinations for Combating Macrolide-Resistant MRSA Recovered from Hospitalized Patients. Biology (Basel). 2021; 10(7). PMC: 8301042. DOI: 10.3390/biology10070624. View

15.

Lipsky B, Berendt A, Cornia P, Pile J, Peters E, Armstrong D . 2012 Infectious Diseases Society of America clinical practice guideline for the diagnosis and treatment of diabetic foot infections. Clin Infect Dis. 2012; 54(12):e132-73. DOI: 10.1093/cid/cis346. View

16.

Palavecino E . Rapid Methods for Detection of MRSA in Clinical Specimens. Methods Mol Biol. 2019; 2069:29-45. DOI: 10.1007/978-1-4939-9849-4_2. View

17.

Nnadozie C, Odume O . Freshwater environments as reservoirs of antibiotic resistant bacteria and their role in the dissemination of antibiotic resistance genes. Environ Pollut. 2019; 254(Pt B):113067. DOI: 10.1016/j.envpol.2019.113067. View

18.

Zhao L, Huang X, Zhang T, Zhang X, Jiang M, Lu H . A point-of-care test device for MRSA rapid detection. J Pharm Biomed Anal. 2021; 209:114464. DOI: 10.1016/j.jpba.2021.114464. View

19.

Huggett J, Foy C, Benes V, Emslie K, Garson J, Haynes R . The digital MIQE guidelines: Minimum Information for Publication of Quantitative Digital PCR Experiments. Clin Chem. 2013; 59(6):892-902. DOI: 10.1373/clinchem.2013.206375. View

20.

Maree M, Thi Nguyen L, Ohniwa R, Higashide M, Msadek T, Morikawa K . Natural transformation allows transfer of SCCmec-mediated methicillin resistance in Staphylococcus aureus biofilms. Nat Commun. 2022; 13(1):2477. PMC: 9072672. DOI: 10.1038/s41467-022-29877-2. View