scholarly journals Development of SYBR Green Real time PCR for identification methicillin-resistance Staphylococcus aureus (MRSA)

2013 ◽  
Vol 7 (1) ◽  
pp. 67-71
Author(s):  
Noor Hashim Kareem ◽  
Majeed Arsheed Sabbah ◽  
Anas Noori Ibraheem ◽  
Al-Shayma’a Muhammad Said
Keyword(s):  
Staphylococcus Aureus ◽  
Real Time ◽  
Real Time Pcr ◽  
Methicillin Resistance ◽  
Sybr Green ◽  
Diffusion Method ◽  
Clinical Samples ◽  
Duplex Pcr ◽  
Minimum Concentration ◽  
Specific Identification

The increasing resistance of staphylococci to ß -lactam antibiotics has become a major clinical problem. Development of rapid and sensitive techniques for detection of MRSA is an important aim for public health. A duplex PCR were established for specific identification of methicillin-resistance Staphylococcus aureus (MRSA) in clinical samples. In this work a duplex SYBR Green real time PCR was developed for rapid identification of MRSA in local methicillin-resistance S. aureus isolates. Twenty methicillin-resistance S. aureus isolates, as determined by disc diffusion method, were subjected to DNA extraction and PCR amplification. Two genes were amplified successfully, mecA (533bp) and femA (314bp), as targets for methicillin-resistance and specific identification of S. aureus, respectively using conventional PCR. Sensitivity of the duplex PCR showed that the minimum concentration of DNA that gave positive results for the two genes was 30ng/µl. In order to develop rapid and sensitive test for identification of MRSA, serial dilutions of purified DNA were amplified gradually according to their concentrations using SYBR Green real time PCR. These results indicated that the SYBR Green real time PCR can be used for identification of methicillin-resistance S. aureus (MRSA) in clinical

scholarly journals Identification of the respiratory tract infection due to methicillin-resistant Staphylococcus aureus by TaqMan real-time PCR

2021 ◽  
Vol 9 (2) ◽  
pp. 86-92
Author(s):  
Sabah Saad Abdulsahib
Keyword(s):  
Staphylococcus Aureus ◽  
Respiratory Tract ◽  
Real Time ◽  
Real Time Pcr ◽  
Gene Sequence ◽  
Methicillin Resistance ◽  
Clinical Samples ◽  
Female Ratio ◽  
Methicillin Resistant ◽  
Tract Infections

Abstract The methicillin-resistant Staphylococcus aureus (MRSA) is a significant human pathogenic bacterium that is endemic within hospitals around the world. The identification and inspection of MRSA in clinical samples is quite helpful both in advising individual patients about the required care and in tracking these species. The goal of this study was to present a modern, faster, and more accurate diagnostic technique to operate on the real-time duplex PCR applicable to S. aureus/MRSA monitoring in Iraqi patients. For this reason, the S. aureus-specific nuc gene sequence and the mecA gene sequence were checked simultaneously. To estimate the assay efficiency, a set of six target strains, 34 non-target strains, and 296 clinical specimens were used. The findings obtained from the diagnosis of a total of 296 isolates based on phenotypic characteristics and biochemical tests showed that 146 (49.32%) were classified as individuals with respiratory tract infections of S. aureus with a total male to female ratio of 1.47, and 142 isolates demonstrated methicillin resistance. 142 MRSA isolates were investigated in the molecular analysis, all MRSA isolates had positive results for the nuc gene and 138 isolates were positive for the mecA gene. The current real-time PCR assay has 97% sensitivity, 100% specificity, and 98% accuracy. Running title: Identification of the MRSA by real time PCR

scholarly journals Susceptibility of Clinical Isolates of Staphylococcus aureus to Ceftaroline

2021 ◽  
Author(s):  
Harsha Sreedharan ◽  
KB Asha Pai
Keyword(s):  
Staphylococcus Aureus ◽  
Tertiary Care Hospital ◽  
Methicillin Resistance ◽  
Descriptive Analysis ◽  
Tertiary Care ◽  
The United States ◽  
Clinical Isolates ◽  
Diffusion Method ◽  
Clinical Samples ◽  
Care Hospital

Introduction: Methicillin-Resistant Staphylococcus aureus(MRSA) infection is a major global healthcare problem, the prevalence of which varies from 25-50% in India. It is known to cause Skin and Soft tissue Infections (SSI), endovascular infections, endocarditis, pneumonia, septic arthritis, osteomyelitis, and sepsis. Vancomycin is the drug of choice for treating severe MRSA infections. Ceftaroline, a fifth-generation cephalosporin has been approved by the United States Food and Drug Administration (US FDA) for treating acute bacterial SSI caused by susceptible micro-organisms including MRSA, Community acquired respiratory tract infection, MRSA bacteremia and endocarditis. Aim: To assess the susceptibility of clinical isolates of S. aureusto ceftaroline, in a Tertiary Care Hospital. Materials and Methods: This prospective study was conducted in the Department of Microbiology of a Tertiary Care Hospital over a period of two months from June 2019 to July 2019. S.aureus isolates from various clinical samples were screened for methicillin resistance by disc diffusion method using cefoxitin disc and ceftaroline susceptibility of these isolates was assessed by E-strip method. The isolates were classified as ceftaroline susceptible, Susceptibility Dose Dependent (SDD) and ceftaroline resistant respectively as per CLSI guidelines. A descriptive analysis of the data was done and the results were presented as frequencies and percentages. Results: All the S.aureus isolates were found to be susceptible to ceftaroline. Methicillin Sensitive Staphylococcus aureus(MSSA) isolates had lower Minimum Inhibitory Concentration (MIC) when compared to MRSA. The highest MIC among MRSA was 0.5 μg/mL. Conclusion: Ceftaroline can be considered as an effective alternative for treatment of infections caused by MRSA.

scholarly journals Molecular characterization of resistance mechanisms in Pseudomonas aeruginosa isolates resistant to carbapenems

2018 ◽  
Vol 11 (12) ◽  
pp. 935-943 ◽  
Author(s):  
Mona Shaaban ◽  
Ahmed Al-Qahtani ◽  
Mohammed Al-Ahdal ◽  
Rasha Barwa
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Real Time ◽  
Real Time Pcr ◽  
Treatment Options ◽  
Carbapenem Resistance ◽  
Pulse Field ◽  
Diffusion Method ◽  
Clinical Samples ◽  
Pcr Analysis ◽  
Carbapenem Resistant

Introduction: Emergence of carbapenem resistance in Pseudomonas aeruginosa increases the therapeutic dilemma. In this study, we investigated various mechanisms involved in the resistance of P. aeruginosa clinical isolates to carbapenems. Methodology: P. aeruginosa isolates were isolated from different clinical samples. The antimicrobial susceptibility was evaluated by disc diffusion method. Carbapenemases were detected among carbapenem resistant isolates. Expression level of mexB and oprD was determined by real-time PCR. Molecular relatedness among isolates was detected based on pulse-field gel electrophoresis (PFGE). Results: Ninety P. aeruginosa isolates were purified from clinical specimens. High levels of resistance to imipenem and meropenem were detected in 16 isolates. PCR analysis of carbapenemases indicated the prevalence of Verona integron-encoded metallo-beta-lactamase (VIM); four isolates produced only VIM enzymes (VIM-1 or VIM-2), while the remaining twelve co-produced both VIM-1 or VIM-2 and NDM enzymes. Additionally, real-time PCR analysis elucidated high expression levels of mexB in seven of the carbapenem resistant isolates and low expression of oprD in seven isolates. The identified carbapenem-resistant isolates were clustered into eleven PFGE profiles where clusters E1 and E2 involved isolates exhibiting multiple carbapenemase genes (blaNDM-1, blaVIM-1 and blaVIM-2). Conclusion: Various mechanisms underlying carbapenem resistance have been detected in our P. aeruginosa cohort of isolates. Emergence of P. aeruginosa as a reservoir of multiple carbapenemases is increasing over time limiting the treatment options to this serious infection. This increases the urgency for infection control practices to reduce the incidence of this infection.

scholarly journals Determination of the prevalence of Salmonella spp. and S. aureus in meat products by Real-Time PCR and testing their antibiotic susceptibility

2021 ◽  
Vol 77 (06) ◽  
pp. 6533-2021
Author(s):  
REYHAN IRKIN ◽  
BERKAY BOZKURT ◽  
GULENDAM TUMEN
Keyword(s):  
Public Health ◽  
Staphylococcus Aureus ◽  
Risk Factor ◽  
Real Time ◽  
Real Time Pcr ◽  
Meat Products ◽  
Penicillin G ◽  
Diffusion Method ◽  
Salmonella Spp ◽  
And Staphylococcus Aureus

From a public health point of view meat products contain high pathogenic risk factors. This is because they can be contaminated with Salmonella spp. and Staphylococcus aureus microorganisms, and, more importantly, antibiotic resistance has been reported in these microorganisms at an increasing frequency. To examine the presence of Salmonella spp. and Staphylococcus aureus in samples of raw and semi-cooked (chicken doner, meat doner, chicken, beef, and lamb products) meat from markets of Izmir and Balikesir, Turkey were analysed. The presence of microorganisms in the samples was determined by Real-Time PCR method using Salmonella spp. and S. aureus specific primers. Following Real-Time PCR, microorganisms were isolated by selective culture methods and biochemical tests from the positive meat samples and tested for their antibiotic susceptibility using the Kirby-Bauer Disk Diffusion Method. The antibiotic disc diffusion method showed that S. aureus was resistant to penicillin G, oxytetracycline, sulfamethoxazole, tetracycline, erythromycin, and ampicillin, whereas Salmonella spp. was resistant to penicillin G, sulfamethoxazole, erythromycin, and ampicillin. As these products are consumed frequently, their contamination with S. aureus (≥ 5 × 103 cfu/g) and Salmonella spp. can be a risk factor for food poisoning. The contamination of meat products’ with S. aureus and Salmonella spp. can be a risk factor for public health and the antibiotics to be preferred in illness treatment are of critical importance.

scholarly journals Detection of inducible clindamycin resistance in Staphylococcus aureus from various samples in a tertiary care hospital

2019 ◽  
Vol 7 (12) ◽  
pp. 4696
Author(s):  
Rohit Kumar ◽  
Jagarti . ◽  
Mrinmoy Sarma ◽  
Gautam Shalini
Keyword(s):  
Staphylococcus Aureus ◽  
Methicillin Resistance ◽  
Tertiary Care ◽  
Antibiotic Sensitivity ◽  
Clinical Isolates ◽  
Diffusion Method ◽  
Clinical Samples ◽  
Care Hospital ◽  
Sensitivity Testing ◽  
Inducible Clindamycin Resistance

Background: The increasing frequency of MRSA infections and rapidly changing patterns in antimicrobial resistance, led to renewed interest in the usage of Macrolides-Lincosamide-Streptogramin B (MLSB) antibiotics to treat Staphylococcus aureus infection. Clindamycin is an important drug used in the treatment of MRSA and MSSA infection. The aim of this study was to determine inducible and constitutive clindamycin resistance among clinical isolates of Staphylococcus aureus by D-test.Methods: During a period of 6 months from July 2018 to December 2018, a total of 100 Staphylococcus aureus isolated from different clinical samples were subjected to routine antibiotic sensitivity testing by Kirby Bauer’s disc diffusion method. Methicillin-resistance was determined by using the cefoxitin (30 µg) disc. Incidence of MLSBc and MLSBi in Staphylococcus aureus isolates by D-test as per CLSI guidelines.Results: Out of 100 isolates of Staphylococcus aureus obtained from 350 clinical samples, 70(70%) were found to be MRSA and 30(30%) were MSSA. Among 100 Staphylococcus aureus isolates, 40% isolates showed MLSBi resistance, 28% isolates showed MLSBc resistance, 6% isolates showed MS phenotype and 26% isolates showed Sensitive phenotype. MLSBc and MLSBi were found to be higher in MRSA as compared to MSSA (21%, 27% and 7%, 10% respectively). All clinical isolates showed 100% sensitivity to Vancomycin and Linezolid in routine antibiotic susceptibility testing.Conclusions: Continuous surveillance of the MLSB resistance is important and required before the prescription of clindamycin to treat MRSA infections.

Molecular Investigation of Staphylococcal Cassette Chromosome mec (SCCmec) Elements Isolated from Intensive Care Unit

2020 ◽  
Author(s):  
Fahimeh Nourbakhsh ◽  
Vajiheh Nourbakhsh ◽  
Samaneh Borooni ◽  
Elaheh Tajbakhsh ◽  
Dana Daneshmand
Keyword(s):  
Intensive Care Unit ◽  
Staphylococcus Aureus ◽  
Antibiotic Resistance ◽  
Intensive Care ◽  
Methicillin Resistance ◽  
Diffusion Method ◽  
Clinical Samples ◽  
High Morbidity ◽  
Molecular Investigation ◽  
Mrsa Strains

Background and Aims: Based on the results, Staphylococcus aureus is one of the serious infectious agents found in community and hospitals with remarkable potential for high morbidity and mortality around the globe. The present study was carried out for molecular investigation of methicillinresistant Staphylococcus aureus strains and Staphylococcal Chromosomal Cassette mec (SCCmec) phenotypes isolated from the intensive care unit in Hazrat Fatemeh Zahra hospital of Isfahan. Materials and Methods: A total of 76 clinical wound samples were collected from Hazrat Fatemeh Zahra Hospital in Isfahan and evaluated by polymerase chain reaction (PCR) methods. The Methicillin resistance Staphylococcus aureus (MRSA) screening was performed by genotypic and phenotypic methods; also antibiotic resistance pattern was determined by using the disk diffusion method and related genes by PCR. Results: Totally, 53 (69.7%) out of 76 clinical samples were positive for MRSA. Of the 76 MRSA strains, 39 (63.51%) were PVL positive (51.3%). The most commonly infected samples were collected from wounds (40.8%). The most commonly detected antibiotic resistance genes were mecA (89.61%), tetK (88.23%), tetM (49.15%) and msrA (46.93%). Resultantly, it was shown that MRSA has the highest level of resistance against methicillin (98%), penicillin (97.24%), tetracycline (89.64%). It was also revealed that the most commonly detected SCCmec types in the MRSA strains are types II (14.53%) and III (16.82%). Conclusions: In summary, this paper argues that the orderly surveillance of hospital-associated infections and initial management and supervision of the antibiotic resistance patterns are required to control the prevalence of MRSA.

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