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1.
Mikrobiyol Bul ; 58(3): 284-292, 2024 Jul.
Artículo en Turco | MEDLINE | ID: mdl-39046210

RESUMEN

Viral load monitoring is important in identifying patients at risk of developing cytomegalovirus (CMV) related complications after transplantation and for this purpose, quantitative real-time polymerase chain reaction (Rt-qPCR) tests are most commonly used. The main problem in CMV DNA Rt-qPCR tests that make quantitative measurements is that there are significant differences in measurements performed with different kits in different laboratories. Comparability of viral load measurements between laboratories has increased with the introduction of quantitative PCR tests calibrated with the CMV International Quantitation Standard (IQS) developed by the World Health Organization (WHO). However, quantitative agreement between measurements made with different kits has still not been fully achieved. In this study, it was aimed to investigate the quantitative compatibility between measurements made with Cobas 6800 (Roche Diagnostics, Mannheim, Germany) and NeuMoDx (Qiagen, Ann Arbor, USA) CMV DNA Rt-qPCR tests, which are fully automated new generation systems calibrated with the WHO CMV IQS. The results of 214 plasma samples, which were studied simultaneously with Cobas 6800 CMV Rt-qPCR and NeuMoDx CMV Rt-qPCR tests were analyzed. In the tests, the extraction, amplification and detection stages were carried out fully automatically. CMV DNA was detected in 144 (67.28%) samples in both tests and was not detected in 53 (24.76%) samples. Incompatible results were obtained in a total of 17 (7.94%) samples. Good agreement was found between the qualitative results of both tests (kappa= 0.80, p< 0.001). When the quantitative results (n= 129) obtained in the dynamic measurement range of both tests were examined, the median viral load values measured by Cobas 6800 CMV Rt-qPCR and NeuMoDx CMV Rt-qPCR tests were 513 IU/mL (range= 35-37000) and 741 IU/mL (range= 68-48978), respectively. According to the correlation analysis, a very strong correlation was found between the results of both tests (r= 0.94, p< 0.001). According to Bland-Altman analysis; the average difference between the results of the NeuMoDx CMV Rt-qPCR test and the Cobas 6800 CMV Rt-qPCR test was found to be -0.14 log10 [standard deviation (SD)= 0.23] IU/mL and it was determined that the Cobas 6800 CMV Rt-qPCR test had lower measurements than the NeuMoDx CMV Rt-qPCR test. In 120 of 129 samples (93%) whose results were within the dynamic measurement range of both tests, the measurement difference was within ± 0.5 log10 IU/mL and in 9 (7%), it was detected as more than ± 0.5 log10 (median 0.54 log10 IU/ml; range= 0.51-0.81). No measurement difference of more than ± 1.0 log10 was detected in any sample. In this study, quantitative agreement was found in the measurements made in plasma samples with the fully automated Cobas 6800 CMV Rt-qPCR and NeuMoDx CMV Rt-qPCR tests calibrated with the CMV IQS. To the best of our knowledge, a study comparing viral load measurements made with Cobas 6800 and NeuMoDx fully automated systems in the detection of CMV DNA has not yet been conducted, and this is the first study on this subject.


Asunto(s)
Infecciones por Citomegalovirus , Citomegalovirus , ADN Viral , Reacción en Cadena en Tiempo Real de la Polimerasa , Carga Viral , Humanos , Infecciones por Citomegalovirus/diagnóstico , Infecciones por Citomegalovirus/virología , Citomegalovirus/genética , Citomegalovirus/aislamiento & purificación , Carga Viral/métodos , Carga Viral/normas , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Reacción en Cadena en Tiempo Real de la Polimerasa/normas , ADN Viral/análisis , ADN Viral/sangre , Juego de Reactivos para Diagnóstico/normas
2.
Microb Pathog ; 149: 104397, 2020 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-32707315

RESUMEN

BACKGROUND: High viral loads are observed in Torque Teno Virus (TTV) infection after hematopoietic stem cell transplantation (HSCT). We aimed to analyze the kinetics of plasma TTV-DNA load in pediatric patients who received immunosuppressive therapy and developed infection complications in the first 100 days after HSCT. METHODS: As a patient group; 113 plasma samples taken from 33 pediatric HSCT recipients at a time interval after transplantation and as a control group; 38 plasma samples from 38 children without known chronic disease were included in the study. Viral nucleic acid isolation was performed by using the NucliSENS easyMAG (bioMerieux, France) system. A laboratory designed quantitative polymerase chain reaction process was performed on 7300 Real-Time PCR system (Applied Biosystems, CA, USA) with the amplification mixture containing primer and probe sequences for the UTR gene region. RESULTS: TTV-DNA was detected in all patient's samples and the median viral load was calculated as 7.67 Log10 copies/mL (range: 2.84-9.59). In the control group, the TTV-DNA median viral load was calculated as 5.51 Log10 copies/mL (range: 2.50-7.04), except for one negative sample. A significant difference was observed between the control group and the patient group in terms of TTV viral load levels. In nine patients, a median 2.15 Log10 copies/mL viral load increase was observed at 31-60 days post-transplant compared to the pre-transplant period. CONCLUSION: TTV-DNA levels should be closely monitored to understand the immune status of the first 100 days after transplantation and the effects of treatment regimens on patients with HSCT.


Asunto(s)
Infecciones por Virus ADN , Trasplante de Células Madre Hematopoyéticas , Torque teno virus , Niño , ADN Viral/genética , Humanos , Torque teno virus/genética , Carga Viral
3.
Clin Lab ; 60(7): 1213-5, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25134392

RESUMEN

BACKGROUND: Because of the emergence and spread of extended-spectrum-beta-lactamase (ESBL)-producing strains which are resistant to many antibiotics, reliable detection of ESBL is very important for infection control. Several chromogenic media have been proposed for the detection of ESBL producers in addition to the conventional phenotypic and genotyping methods. The aim of the present study was to evaluate the performance of Brilliance ESBL agar (Oxoid; Thermo Fisher Scientific, UK), a selective chromogenic agar for the detection of ESBL-producing Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) strains. METHODS: A total of 237 strains (143 ESBL producers (76 isolates of E. coli and 67 isolates of K. pneumoniae) and 94 non-ESBL producers (44 isolates of E. coli and 50 isolates of K. pneumoniae)) isolated from various clinical specimens were included in the study. Isolates were identified by conventional methods, Phoenix system (Becton Dickinson, USA), and mass spectrometry. ESBL confirmation was performed by phenotypical tests. A 10 microL aliquot of each isolate's 0.5 McFarland suspension was streaked onto Brilliance ESBL agar. All plates were incubated at 37 degrees C for 24 hours and then were interpreted for growth and colony color according to the manufacturer's recommendations. Identification and ESBL test results were used to calculate the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the medium evaluated at 24 hours. RESULTS: The sensitivity, specificity, PPV, and NPV of the medium were 97.9%, 100%, 100%, and 96.9%, respectively, when considering only species specific colored colonies of the isolates. CONCLUSIONS: Brilliance ESBL agar could provide a practical alternative to the traditional methods for the identification of ESBL producers.


Asunto(s)
Escherichia coli/enzimología , Klebsiella pneumoniae/enzimología , beta-Lactamasas/biosíntesis , Medios de Cultivo
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