Rapid Detection Of Methicillin-resistant Staphylococcus Aureus Research Articles

Multiplex Digital PCR-Based Development and discussion of the Detection of Genetic Association Between Staphylococcus aureus and mecA.

Methicillin-resistant Staphylococcus aureus (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 Staphylococcus aureus (MSSA) and MRSA. This protocol employed TaqMan probes to detect SAOUHSC_00106 and mecA 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 Staphylococcus aureus and the timely identification of MRSA in clinical samples, thereby providing an advanced platform for the rapid diagnosis of clinical MRSA infection.

Screening for mecA and mecC Gene Carriage among Clinical Isolates of Methicillin Resistant Staphylococcus aureus at a Tertiary Care Hospital: A Cross-sectional Study

Introduction: Staphylococcus aureus (S. aureus) has emerged as one of the most important human pathogen, and has been a leading cause of hospital and community acquired infections. Methicillin-Resistant Staphylococcus aureus (MRSA) carrying the mecA gene is resistant to the majority of β-lactam antibiotics. In 2007, a new S. aureus strain harboring mecA gene homologue, mecC, was found in England which posed diagnostic problems. Accurate and rapid detection of MRSA is required for effective treatment. Aim: To screen for mecA and mecC genes among methicillin resistant isolates of S. aureus using conventional Polymerase Chain Reaction (PCR) and to associate their presence to KirbyBauer disc diffusion and automated Vitek 2 methods. Materials and Methods: This cross-sectional observational study was conducted in the Department of Microbiology, Victoria hospital, Bangalore Medical College and Research Institute, Bengaluru, Karnataka, India, from July to October 2019. A total of 60 duplicate S. aureus samples were obtained from various clinical samples during the study period. Isolates were subjected to antibiotic susceptibility by cefoxitin disc diffusion and automated Vitek-2. Isolates were screened for mecA and mecC gene carriage using conventional PCR. Descriptive statistics was used for the comparison of data and appropriate statistical charts were used to present the data. Results: Among 60 S. aureus isolates, 41 (68.33%) of them were considered MRSA by conventional Disc Diffusion Method (DDM) and 48 (80%) of them were considered MRSA by automated Vitek-2. By conventional PCR only 34 (56.67%) isolates carried the mecA gene and none of the clinical isolates possessed the mecC gene. Conclusion: An overall MRSA prevalence of 56.67% was observed by PCR in present study. The mecC gene was not detected in any of the S. aureus isolates. Present study indicates the presence of mecA and mecC negative phenotypically identified MRSA isolates. Rather than absolute dependence on the mecA gene as the defining standard in determining MRSA, alternative mechanisms of resistance-presence of mecC, mecB genes, hyperproduction of b-lactamase; can potentially be a knowledge trove for researchers to delve into.

Three-Dimensional Surface-Enhanced Raman Scattering Platform with Hotspots Built by a Nano-mower for Rapid Detection of MRSA.

Methicillin-resistant Staphylococcus aureus (MRSA) has become one of the greatest threats to human health due to its strong drug resistance, wide distribution range, and high infection rate. Rapid identification of MRSA strains is very important for accurate diagnosis and early treatment of MRSA infections. Here, we introduced an Exo III-assisted nanomotor mower to build 3D hotspots for rapid detection of MRSA by surface-enhanced Raman scattering (SERS). As the bacteria bound to the aptamer, two trigger chains were released from the double-stranded structure, and the nano-mowers were activated by opening a hairpin probe on gold nanoparticles (AuNPs). With the continued cleavage of Exo III and cyclic release of the trigger chain, multiple hairpin DNAs on the AuNPs were cleaved to increase the motor power. The resulting nano-mower continued slicing protective DNA from larger AuNPs, exposing the AuNPs. Without the protection of DNA, Mg2+ in the buffer induced spontaneous aggregation of the AuNPs, and a large number of hotspots were formed for SERS measurements. Under optimal conditions, MRSA can be detected within 40 min, and the concentration of MRSA showed a good linear relationship with the SERS intensity at 1342 cm-1, with a limit of detection as low as 1 CFU/mL and a wide linear range (100 to 107 CFU/mL). This strategy creates a rapid bacterial detection method that performs well on actual samples utilizing portable Raman spectroscopy instruments, with potential applications in food detection, water detection, clinical treatment, and diagnosis.

Development of a Real-Time Recombinase-Aided Amplification Method to Rapidly Detect Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant staphylococcus aureus (MRSA) is a major pathogen responsible for human hospital and community-onset diseases and severe invasive livestock infections. Rapid detection of MRSA is essential to control the spread of MRSA. Conventional identification methods and antibacterial susceptibility tests of MRSA are time-consuming. The commonly used qPCR assay also has the disadvantages of being complicated and expensive, restricting its application in resource-limited clinical laboratories. Here, a real-time fluorescent recombinase-assisted amplification (RAA) assay targeting the most conserved regions within the mecA gene of MRSA was developed and evaluated to detect MRSA. The detection limit of this assay was determined to be 10 copies/reaction of positive plasmids. The established RAA assay showed high specificity for MRSA detection without cross-reactivities with other clinically relevant bacteria. The diagnostic performance of real-time RAA was evaluated using 67 clinical S. aureus isolates from dairy farms, which were detected in parallel using the TaqMan probe qPCR assay. The results showed that 56 and 54 samples tested positive for MRSA by RAA and qPCR, respectively. The overall agreement between both assays was 97.01% (65/67), with a kappa value of 0.9517 (p < 0.001). Further linear regression analysis demonstrated that the detection results between the two assays were significantly correlated (R2 = 0.9012, p < 0.0001), indicating that this RAA assay possesses similar detection performance to the qPCR assay. In conclusion, our newly established RAA assay is a time-saving and convenient diagnostic tool suitable for MRSA detection and screening.

Rapid and Ultrasensitive Detection of Methicillin-Resistant Staphylococcus aureus Based on CRISPR-Cas12a Combined With Recombinase-Aided Amplification.

Staphylococcus aureus is one of the main pathogens causing hospital and community-acquired infections, in particular, infections caused by methicillin-resistant Staphylococcus aureus (MRSA) cause a higher mortality rate than those caused by methicillin-sensitive strains, which poses a serious global public health problem. Therefore, rapid and ultrasensitive detection of patients with clinical MRSA infection and timely control of infection are essential. Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) based on nucleic acid detection methods are well-known for its high specificity and sensitivity and programmability. Here, we successfully proposed a method based on CRISPR-Cas12a combined with recombinase-aided amplification (RAA) through fluorescent readout to achieve accurate identification and highly sensitive detection of MRSA in clinical samples. Results showed that the limit of detection (LoD) of the RAA-Cas12a method could reach 10 copies/μl at 60 min of reaction. Specificity tests showed that the method could distinguish MRSA from clinically common bacteria. The results of RAA-Cas12a were consistent with that of antimicrobial susceptibility tests (AST) and polymerase chain reaction (PCR) in 83 clinical samples. These results indicated that the detection method based on RAA-Cas12a has high sensitivity and specificity, and provides important value for rapid detection of MRSA.

A point-of-care test device for MRSA rapid detection

Staphylococcus aureus (SA) is one of the most common pathogenic bacteria, and methicillin-resistant SA (MRSA) is an equally common drug-resistant bacteria. MRSA detection is of great significance for clinical diagnosis, medication guidance, and prevention of antibiotic abuse. Traditional MRSA detection using the culture method is time-consuming, laborious, and difficult to conduct rapid on-site detection. In this research, we developed a device for rapid MRSA detection, which can detect the nuc gene in SA and mecA gene in MRSA simultaneously for 30–40 min. After simple sample processing, the mixture can be directly loaded onto the chip device. The detection results can be directly determined by a color change, with a limitation of approximately 102 copies. This isothermal amplification chip device can be widely applied in many fields, with simple operation and low contamination.

Rapid MRSA detection via tandem mass spectrometry of the intact 80\xa0kDa PBP2a resistance protein

Treatment of antibiotic-resistant infections is dependent on the detection of specific bacterial genes or proteins in clinical assays. Identification of methicillin-resistant Staphylococcus aureus (MRSA) is often accomplished through the detection of penicillin-binding protein 2a (PBP2a). With greater dependence on mass spectrometry (MS)-based bacterial identification, complementary efforts to detect resistance have been hindered by the complexity of those proteins responsible. Initial characterization of PBP2a indicates the presence of glycan modifications. To simplify detection, we demonstrate a proof-of-concept tandem MS approach involving the generation of N-terminal PBP2a peptide-like fragments and detection of unique product ions during top-down proteomic sample analyses. This approach was implemented for two PBP2a variants, PBP2amecA and PBP2amecC, and was accurate across a representative panel of MRSA strains with different genetic backgrounds. Additionally, PBP2amecA was successfully detected from clinical isolates using a five-minute liquid chromatographic separation and implementation of this MS detection strategy. Our results highlight the capability of direct MS-based resistance marker detection and potential advantages for implementing these approaches in clinical diagnostics.

Review on Recent Major Diagnostic Methods: The Diagnostic of Methicillin Resistant Staphylococcus Aurus

According to the World Health Organization (WHO) 20% of all documented S. aureus infections are attributable to methicillin resistant staphylococcus aureus (MRSA), although for some developing countries this value can exceed 80%. Thus the rapid and accurate detection of MRSA in low resource settings is becoming essential. Yet conventional microbial detection methods take from 1-5 days to identify MRSA. Recently, new types of automated laboratory methods as well as advances in nucleic acid testing, microfluidic technology, immunosensors, biosensors and point of care testing have reduced the time to detection to <1 hr. This review examines the current limitations and advances in methodologies employed in the rapid detection of MRSA. Keywords : S.aureus, MRSA, Pathogenesis, Diagnosis DOI: 10.7176/JBAH/11-2-03 Publication date: January 31 st 2021

Accurate Detection of Methicillin-Resistant Staphylococcus aureus in Mixtures by Use of Single-Bacterium Duplex Droplet Digital PCR.

Accurate and rapid identification of methicillin-resistant Staphylococcus aureus (MRSA) is needed to screen MRSA carriers and improve treatment. The current widely used duplex PCR methods are not able to differentiate MRSA from coexisting methicillin-susceptible S. aureus (MSSA) or other methicillin-resistant staphylococci. In this study, we aimed to develop a direct method for accurate and rapid detection of MRSA in clinical samples from open environments, such as nasal swabs. The new molecular assay is based on detecting the cooccurrence of nuc and mecA markers in a single bacterial cell by utilizing droplet digital PCR (ddPCR) with the chimeric lysin ClyH for cell lysis. The method consists of (i) dispersion of an intact single bacterium into nanoliter droplets, (ii) temperature-controlled release of genomic DNA (gDNA) by ClyH at 37°C, and (iii) amplification and detection of the markers (nuc and mecA) using standard TaqMan chemistries with ddPCR. Results were analyzed based on MRSA index ratios used for indicating the presence of the duplex-positive markers in droplets. The method was able to achieve an absolute limit of detection (LOD) of 2,900 CFU/ml for MRSA in nasal swabs spiked with excess amounts of Escherichia coli, MSSA, and other mecA-positive bacteria within 4 h. Initial testing of 104 nasal swabs showed that the method had 100% agreement with the standard culture method, while the normal duplex qPCR method had only about 87.5% agreement. The single-bacterium duplex ddPCR assay is rapid and powerful for more accurate detection of MRSA directly from clinical specimens.

Rapid MRSA PCR on respiratory specimens from ventilated patients with suspected pneumonia: a tool to facilitate antimicrobial stewardship.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of pneumonia in ventilated patients. Our objective was to evaluate the GeneXpert MRSA/SA SSTI Assay (Xpert MRSA/SA) (Cepheid, Sunnyvale, CA) for use in lower respiratory tract (LRT) specimens for rapid MRSA detection and to determine the potentially saved antibiotic-days if a culture-based identification method was replaced by this assay. Remnant LRT samples from ventilated patients submitted to the microbiology laboratory for routine culture were tested using conventional culture and Xpert MRSA/SA. One hundred of 310 LRT specimens met the inclusion criteria. Ten samples were positive for MRSA by Xpert MRSA/SA, while six were positive by routine culture methods. Xpert MRSA/SA correctly identified 5/6 positive and 89/94 negative MRSA specimens, for a sensitivity of 83.3%, specificity of 94.7%, positive predictive value of 45.6%, and negative predictive value of 98.9%. The assay also correctly detected 3/3 positive and 90/97 negative methicillin-susceptible S. aureus (MSSA) specimens, for a sensitivity of 100%, specificity of 92.8%, positive predictive value of 30%, and negative predictive value of 100%. A total of 748 vancomycin and 305 linezolid antibiotic-days were associated with the enrolled specimens. Vancomycin and linezolid utilization could decrease by 68.4% and 83%, respectively, if discontinued 1day after negative polymerase chain reaction (PCR) results. The Xpert MRSA/SA SSTI rapid MRSA PCR assay performed well in respiratory samples from ventilated patients with suspected pneumonia and has the potential to facilitate stewardship efforts such as reducing empiric vancomycin and linezolid therapy.

Evaluation of colorimetric nitrate reductase assay for rapid detection of methicillin resistance in clinical isolates of Staphylococcus aureus.

Background & objectives:Methicillin resistant Staphylococcus aureus (MRSA) remains a major cause of health care-associated infections. Rapid detection of MRSA facilitates the early initiation of appropriate treatment and infection control. Hence, the present study was undertaken to standardize and evaluate the performance of rapid colorimetric nitrate reductase assay (NRA) for determining methicillin resistance in S.aureus.Methods:A total of 160 clinical isolates of S. aureus, (80 each of methicillin susceptible and methicillin resistant) were included in the study. Minimum inhibitory concentration (MIC) was determined by NRA and reference broth micro dilution (BMD) methods. Results of NRA were compared with BMD and analyzed.Results:For MRSA, the MIC values ranged from 4 to ≥ 16 μg/ml and for MSSA, ≤ 0.5 to 2 μg/ml. Category and essential agreement for NRA as compared with BMD were found to be 99.4 and 89.7 per cent, respectively. No minor or major discrepancy was observed. A single resistant isolate showed very major discrepancy.Interpretation & conclusions:Colorimetric NRA being an inexpensive test requiring no special equipment can be employed as an alternative method for rapid detection of MRSA in resource limited settings.

Economic Features of Antibiotic Resistance: The Case of Methicillin-Resistant Staphylococcus aureus

This paper analyses and updates the economic information regarding methicillin-resistant Staphylococcus aureus (MRSA), including information that has been previously reviewed by other authors, and new information, for the purpose of facilitating health management and clinical decisions. The analysed articles reveal great disparity in the economic burden on MRSA patients; this is mainly due to the diversity of the designs of the studies, as well as the variability of the patients and the differences in health care systems. Regarding prophylactic strategies, the studies do not provide conclusive results that could unambiguously orientate health management. The studies addressing treatments noted that linezolid seems to be a cost-effective treatment for MRSA, mostly because it is associated with a shorter length of stay (LOS) in hospital. However, important variables such as antimicrobial susceptibility, infection type and resistance emergence should be included in these analyses before a conclusion is reached regarding which treatment is the best (most efficient). The reviewed studies found that rapid MRSA detection, using molecular techniques, is an efficient technique to control MRSA. As a general conclusion, the management of MRSA infections implicates important economic costs for hospitals, as they result in higher direct costs and longer LOS than those related to methicillin-susceptible S. aureus (MSSA) patients or MRSA-free patients; there is wide variability in those increased costs, depending on different variables. Moreover, the research reveals a lack of studies on other related topics, such as the economic implications of changes in MRSA epidemiology (community patients and lineages associated with farm animals).

The use of molecular methods for the detection and identification of methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogen in many hospitals and long-term care facilities as well as in the community. To limit the spread of MRSA, early detection and proper treatment are essential. Because conventional culture as gold standard is time consuming, new techniques such as PCR-based and hybridization assays have emerged for the rapid detection of MRSA. This review will focus on the currently available molecular-based assays and on their utility and performance for detection of S. aureus, of its virulence factors and of the markers for acquired resistance.

LightCycler? MRSA Advanced Test for Rapid MRSA Detection in Referral Patients Admitted to Intensive Care Units

OBJECTIVE: We studied the prevalence of Methicillin Resistant Staphylococcus Aureus (MRSA) nasal carrier in patients who were transferred and admitted to the intensive care unit (ICU). MATERIALS AND METHOD: By using LightCycler® MRSA advanced test for the direct detection of MRSA DNA in nasal colonization by polymerase chain reaction (PCR) and comparing to standard MRSA isolated from plates culture media. RESULTS: From December 2010 to May 2011, 100 patients were enrolled. They were referred to be admitted in ICUs of Bangkok Hospital Medical Center (BMC) (87), Samitivej Hospital Sukhumvit (12) and Bangkok Hospital Pattaya (1). Seventeen patients were excluded from the study. Of 83 patients, the light cycler MRSA test detected MRSA from an anterior nasal swab in 12 (14.5%) patients while concomitant plates culture grew MRSA in 10 (12.1%) (Kappa = 0.7913, 95% condence interval (CI) = 0.594-0.988). CONCLUSION: The LightCycler® MRSA advanced test is a diagnostic test for rapid MRSA detection. The test aids in the detection of hospital infection and in the control of MRSA infections by rapid detection, therefore identifying the appropiate and isolated patient whom has MRSA colonization particularly in high risk patients.

Do Active Surveillance and Contact Precautions Reduce MRSA Acquisition? A Prospective Interrupted Time Series

BackgroundConsensus for methicillin-resistant Staphylococcus aureus (MRSA) control has still not been reached. We hypothesised that use of rapid MRSA detection followed by contact precautions and single room isolation would reduce MRSA acquisition.MethodsThis study was a pre-planned prospective interrupted time series comparing rapid PCR detection and use of long sleeved gowns and gloves (contact precautions) plus single room isolation or cohorting of MRSA colonised patients with a control group. The study took place in a medical-surgical intensive care unit of a tertiary adult hospital between May 21st 2007 and September 21st 2009. The primary outcome was the rate of MRSA acquisition. A segmented regression analysis was performed to determine the trend in MRSA acquisition rates before and after the intervention.FindingsThe rate of MRSA acquisition was 18.5 per 1000 at risk patient days in the control phase and 7.9 per 1000 at-risk patient days in the intervention phase, with an adjusted hazard ratio 0.39 (95% CI 0.24 to 0.62). Segmented regression analysis showed a decline in MRSA acquisition of 7% per month in the intervention phase, (95%CI 1.9% to 12.8% reduction) which was a significant change in slope compared with the control phase. Secondary analysis found prior exposure to anaerobically active antibiotics and colonization pressure were associated with increased acquisition risk.ConclusionContact precautions with single room isolation or cohorting were associated with a 60% reduction in MRSA acquisition. While this study was a quasi-experimental design, many measures were taken to strengthen the study, such as accounting for differences in colonisation pressure, hand hygiene compliance and individual risk factors across the groups, and confining the study to one centre to reduce variation in transmission. Use of two research nurses may limit its generalisability to units in which this level of support is available.

Rapid detection of live methicillin-resistant Staphylococcus aureus by using an integrated microfluidic system capable of ethidium monoazide pre-treatment and molecular diagnosis

Methicillin-resistant Staphylococcus aureus (MRSA) is a bacterium resistant to all existing penicillin and lactam-based antimicrobial drugs and, therefore, has become one of the most prevalent antibiotic-resistant pathogens found in hospitals. The multi-drug resistant characteristics of MRSA make it challenging to clinically treat infected patients. Therefore, early diagnosis of MRSA has become a public-health priority worldwide. Conventionally, cell-culture based methodology and microscopic identification are commonly used for MRSA detection. However, they are relatively time-consuming and labor-intensive. Recently, molecular diagnosis based on nucleic acid amplification techniques, such as polymerase chain reaction (PCR), has been widely investigated for the rapid detection of MRSA. However, genomic DNA of both live and dead pathogens can be distinguished by conventional PCR. These results thus could not provide sufficient confirmation of an active infection for clinicians. In this study, live MRSA was rapidly detected by using a new integrated microfluidic system. The microfluidic system has been demonstrated to have 100% specificity to detect live MRSA with S. aureus and other pathogens commonly found in hospitals. The experimental results showed that the limit of detection for live MRSA from biosamples was approximately 10(2) CFU/μl. In addition, the entire diagnostic protocol, from sample pre-treatment to fluorescence observation, can be automatically completed within 2.5 h. Consequently, this microfluidic system may be a powerful tool for the rapid molecular diagnosis of live MRSA.

Multicenter Evaluation of the LightCycler MRSA Advanced Test, the Xpert MRSA Assay, and MRSASelect Directly Plated Culture with Simulated Workflow Comparison for the Detection of Methicillin-Resistant Staphylococcus aureus in Nasal Swabs

Rapid detection of nasal colonization with methicillin-resistant Staphylococcus aureus (MRSA) followed by appropriate infection control procedures reduces MRSA infection and transmission. We compared the performance and workflow of two Food and Drug Administration-approved nucleic acid amplification assays, the LightCycler MRSA Advanced Test and the Xpert MRSA test, with those of directly plated culture (MRSASelect) using 1202 nasal swabs collected at three U.S. sites. The sensitivity of the LightCycler test (95.2%; 95% CI, 89.1% to 98.4%) and Xpert assay (99%; 95% CI, 94.8% to 100%) did not differ compared with that of culture; the specificity of the two assays was identical (95.5%; 95% CI, 94.1% to 96.7%) compared with culture. However, sequencing performed on 71 samples with discordant results among the three methods confirmed the presence of MRSA in 40% of samples that were positive by both molecular methods but negative by culture. Workflow analysis from all sites including batch runs revealed average hands-on sample preparation times of 1.40, 2.35, and 1.44 minutes per sample for the LightCycler, Xpert, and MRSASelect methods, respectively. Discrete event simulation analysis of workflow efficiencies revealed that the LightCycler test used less hands-on time for the assay when greater than eight batched samples were run. The high sensitivity and specificity, low hands-on time, and efficiency gains using batching capabilities make the LightCycler test suitable for rapid batch screening of MRSA colonization.

MRSA screening: can one swab be used for both culture and rapid testing? An evaluation of chromogenic culture and subsequent Hain GenoQuick PCR amplification/detection

The use of a single swab for both MRSA culture and for rapid testing by PCR was evaluated, using the Hain GenoQuick (GQM) methicillin resistant Staphylococcus aureus (MRSA) assay for the rapid detection of MRSA, as a single swab would be the preferred option for routine diagnostic testing. GQM detected current prevalent Irish MRSA strains incorporating all known SSCmec types, including Panton–Valentine leukocidin-positive strains. Using the GQM method, all methicillin-resistant coagulase-negative staphylococci tested were confirmed to be negative, although three of seven gentamicin-resistant methicillin-sensitive Staphylococcus aureus strains tested were identified as MRSA. The theoretical ex-vivo limit of detection of the assay was 704 CFU per GQM assay reaction (1.7 × 104 CFU/mL) when MRSA suspensions were used for DNA extraction, or 1.4 × 103 CFU/swab (1.4 × 104 CFU/mL) using MRSA absorbed onto Copan screening swabs. Swab processing on chromogenic agar prior to PCR resulted in some inhibition of the PCR reaction, increasing the limit of detection of the assay by a factor of four. Based on 540 single swab screening specimens (nasal and groin) processed first for culture assay, then by GQM, the specificity and positive predictive value were both 100%, the negative predictive value was 92%, and the sensitivity was 57. Culture followed by PCR from a single specimen is not optimal for the rapid detection of MRSA. Further laboratory validation of the GQM assay is required to determine the true diagnostic sensitivity and value of this kit in routine microbiology laboratories, modifying the protocol for single specimens, or using two specimens.

Assessing the role of undetected colonization and isolation precautions in reducing Methicillin-Resistant Staphylococcus aureustransmission in intensive care units

BackgroundScreening and isolation are central components of hospital methicillin-resistant Staphylococcus aureus (MRSA) control policies. Their prevention of patient-to-patient spread depends on minimizing undetected and unisolated MRSA-positive patient days. Estimating these MRSA-positive patient days and the reduction in transmission due to isolation presents a major methodological challenge, but is essential for assessing both the value of existing control policies and the potential benefit of new rapid MRSA detection technologies. Recent methodological developments have made it possible to estimate these quantities using routine surveillance data.MethodsColonization data from admission and weekly nares cultures were collected from eight single-bed adult intensive care units (ICUs) over 17 months. Detected MRSA-positive patients were isolated using single rooms and barrier precautions. Data were analyzed using stochastic transmission models and model fitting was performed within a Bayesian framework using a Markov chain Monte Carlo algorithm, imputing unobserved MRSA carriage events.ResultsModels estimated the mean percent of colonized-patient-days attributed to undetected carriers as 14.1% (95% CI (11.7, 16.5)) averaged across ICUs. The percent of colonized-patient-days attributed to patients awaiting results averaged 7.8% (6.2, 9.2). Overall, the ratio of estimated transmission rates from unisolated MRSA-positive patients and those under barrier precautions was 1.34 (0.45, 3.97), but varied widely across ICUs.ConclusionsScreening consistently detected >80% of colonized-patient-days. Estimates of the effectiveness of barrier precautions showed considerable uncertainty, but in all units except burns/general surgery and one cardiac surgery ICU, the best estimates were consistent with reductions in transmission associated with barrier precautions.

Detection limits of a rapid MRSA detection assay based on multiplex real-time PCR

I wish to report the results of a study for the limits of detection of a rapid methicillin-resistant Staphylococcus aureus (MRSA) detection assay based on a multiplex realtime polymerase chain reaction (PCR) (BD GeneOhm MRSA assay, Fukushima, Japan), which was reported in the Journal of Infection and Chemotherapy, Volume 15 (No. 4), 2009 [1]. ATCC 33591 (MRSA) was recovered on Mueller Hinton agar by overnight incubation at 37 C. Colonies were picked from the agar, adjusted to Mcfarland 0.5, and diluted to 10–10 colony-forming units (CFU)/ml (corresponding to approximately 10–10 CFU/reaction) by the sample buffer (Tris-EDTA buffer) of the BD GeneOhm MRSA assay kit [1]. The BD GeneOhm MRSA assay was performed four times at about the same concentration to determine the limits of detection. Table 1 shows the result of the tests. All tests were positive with 10–10 and 10 CFU/reaction; however, only half of the tests were positive with at a reaction concentration of 10 CFU. Although the results of the study show that at least 10 CFU/reaction is necessary to detect MRSA stably by the BD GeneOhm MRSA assay, compilation of data and further study with clinical samples are needed. Reference