[Comparison of Cytomegalovirus Polymerase Chain Reaction Test Results Obtained with Fully Automated Systems: Roche Cobas 6800 and Qiagen NeuMoDx Experience]. / Tam Otomatize Sistemlerde Çalisilan Sitomegalovirüs Polimeraz Zincir Reaksiyonu Test Sonuçlarinin Karsilastirilmasi: Roche Cobas 6800 ile Qiagen NeuMoDx Deneyimi.

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.

Evaluation of the Effect of Sampling Strategy on Test Results in the COVID-19 Pandemic.

BACKGROUND: The oronasopharyngeal (ONP) sampling phase is critical in the diagnosis of infection during the coronavirus disease-2019 (COVID-19) pandemic. In this study, the aim is to investigate the effect of the standardized operational team in the sampling unit on test results and repetitions. METHODS: Patients who applied to the Adult Pandemic Polyclinic between August 2020 and October 2021 and whose ONP samples were taken for Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) Polymerase Chain Reaction (PCR) test were included in the study. For August 2020, the Ear, Nose and Throat (ENT) specialists were appointed to work in the sample collection unit. For the following months, physicians from other clinics as well as ENT specialists were appointed under the same conditions. The ENT and other specialties were compared in terms of PCR positivity and test repetition. RESULTS: Out of a total of 129,808 patients, 21,602 (16.6%) of them were sampled by ENT physicians, and 108,206 (83.4%) by others. The first three months with the highest number of ONP samples were August 2021 (n: 20,317; 15.7%), July 2021 (n: 11,767; 9.1%), and November 2020 (n: 11,511; 8.9%). The highest positive PCR results were in August 2021 followed by August 2021 (37.3%), December 2020 (32.8%), and November 2020 (32.5%). In September 2020, more positive results were obtained from the samples taken by ENT specialists (p = 0.001). The repetition frequency of ONP samples taken by ENT and other specialists was calculated as 221 (1%) and 878 (0.8%), respectively (p > 0.05), and the month with the highest re-test rate was November 2020. CONCLUSIONS: It is seen that the training of ENT specialists given to non-ENT specialists on sampling technique is effective and the process is managed with optimal benefit. This study can be evaluated as the first step data in comparing all these organizations and emphasizing the importance of the cooperation created by the institutions during the pandemic process.

[Comparison of Two Different Quantitative Tests for Anti-Spike Measurement After Two Doses of Inactivated SARS-CoV-2 Vaccine in Healthcare Professionals]. / Saglik Çalisanlarinda Uygulanan Iki Doz Inaktif SARS-CoV-2 Asisinin Ardindan Anti-Spike Ölçümünde Iki Farkli Kantitatif Testin Karsilastirilmasi.

Serological tests developed to detect antibodies against severe acute respiratory syndrome Coronavirus disease-2 (SARS-CoV-2) antigens are used to demonstrate immunity level, vaccine efficacy and duration of protection. However, the evaluation of these tests and the interpretation of the results are still under investigation. In this study, it was aimed to compare the anti-spike measurement values in healthcare workers who have not experienced Coronavirus-2019 (COVID-19) after vaccination with two doses of inactivated SARS-CoV-2 vaccine with two commercial quantitative antibody detection tests which were used to give results in the same unit and work with different methods. The results obtained with Elecys Anti-SARS-CoV-2 S (Roche Diagnostics, Mannheim, Germany) and Aeskulisa Anti-SARS-CoV-2 S1 (Aesku diagnostics, Wendelsheim, Germany) kits of the serum samples of 90 healthcare workers were evaluated qualitatively and quantitatively in line with the recommendations of both manufacturers. Quantitative values given as units/mL (U/mL) were also evaluated by converting the binding antibody unit (BAU)/mL with the conversion factor obtained as a result of the studies carried out by the manufacturers with the World Health Organization anti-SARS-CoV-2 international standard. Positive results were obtained in 86 (95.6%) samples and negative results in 4 (4.4%) samples with the Elecsys kit; 55 (61.1%) positive, 20 (22.2%) negative and 15 (16.7%) intermediate results were obtained with the Aeskulisa kit. In common with both kits, negative results were obtained in four samples, and positive results were obtained in 55 samples. While the percent agreement observed with both kits was found to be 78.6 in 75 samples, excluding 15 samples in the intermediate with Aeskulisa kit, Cohen's kappa value was calculated as 0.26, indicating a close to moderate agreement, statistically. A high correlation was observed in the correlation analysis of the measurements of both kits (r= 0.611). When the scattering of the differences against the mean of the quantitative measurements made with both kits was examined by Bland-Altman analysis; higher measurements were determined with the Elecsys kit than with the Aeskulisa kit and the mean difference was found to be 58.68 ± 52.62. As a result of the studies carried out by the manufacturers, it was observed that the average difference decreased to 41.09 ± 50.22 when the U/mL values were converted to BAU/mL with the conversion factor. In the whole sampling, the measurements determined with the Elecsys kit were found to be 4.58 ± 3.44 times higher on average compared to the Aeskulisa kit while the measurements were found to be 2.35 ± 1.77 when the BAU/mL values were recalculated. The measurement values found with the Elecsys kit used in our study were found to be higher than that of Aeskulisa kit and qualitatively more positive results were obtained with Elecsys kit than with the Aeskulisa kit. In this study, Elecsys kit measurements were on average 4.06 times higher than Aeskulisa kit in samples (n= 55) with results compatible with both kits. The increase in these rates to 4.30 and 7.72 values, respectively, for 15 samples reported as intermediate and 16 as negative with the Aeskulisa kit, suggested that the success of quantitation decreased with the Aeskulisa kit at low antibody values. Since the harmonization of anti-SARS-CoV-2 tests has not yet been achieved, individual immune monitoring should be performed using the same method and even the results converted to BAU/mL should be specific to the antigen subunit.

[Evaluation of the Rapid Antigen Detection Kit with the Polymerase Chain Reaction for Detection of SARS-CoV-2 in Respiratory Samples]. / Solunum Yolu Örneklerinde SARS-CoV-2 Tespiti için Hizli Antijen Tespit Kitinin Polimeraz Zincir Reaksiyonu ile Birlikte Degerlendirilmesi.

The disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) was named as coronavirus disease-2019 (COVID-19) by the World Health Organization (WHO) in February 11, 2020. The rapid diagnosis of COVID-19 patients is essential to reduce the disease spread. The reverse-transcription polymerase chain reaction (RT-PCR) is the gold standard test to diagnose SARS-CoV-2 acute infection. The rapid antigen test which can detect the presence of viral protein antigens in respiratory tract samples is being investigated as an alternative option, especially in cases where RT-PCR is not available or the test capacity is exceeded, due to its faster results, ease of application, low cost and lack of special equipment and personnel. In this study, it was aimed to evaluate the performance of a commercial rapid antigen test using nasopharyngeal samples of COVID-19 patients confirmed with RT-PCR. From the first day of the research, the first 80 consecutively SARS-CoV-2 RT-PCR positive and 40 RT-PCR negative respiratory samples sent to the Medical Microbiology Laboratory for routine SARS-CoV-2 RT-PCR testing were included in the study. RT-PCR tests of the samples were performed in routine studies with the BioSpeedy SARS-CoV-2 RT-PCR kit (Bioeksen, Turkey). Rapid antigen tests were performed with the Wesail COVID-19 antigen test kit (Guangdong, China) simultaneously with RT-PCR tests. Amongst the 80 positive RT-PCR samples, 56 were detected by the rapid antigen test. All the samples detected as positive with the rapid antigen tests were also positive with RT-PCR. There was a moderate agreement between the qualitative results of both tests (Kappa= 0.609, p<0.001). According to the PCR test, the sensitivity, specificity, positive predictive value (PPV), negative predictive value, and accuracy of the rapid antigen test were; 70%, 100%, 100%, 62.5%, and 80% (96/120), respectively. The sensitivities of the rapid antigen test were calculated as 92.6% in 54 samples with a cycle threshold (Ct) value of <17, 88.7% in 62 samples with a Ct value of <20, 77.8% in 72 samples with a Ct value of <22, and 74.7% in 75 samples with a Ct value of <25. According to our study data; the rapid antigen test was found less sensitive than the RT-PCR test. Negative results obtained with rapid antigen testing cannot exclude SARS-CoV-2 infection and must be confirmed by RT-PCR. In addition, according to the ROC analysis of rapid antigen test positivity obtained according to RT-PCR Ct values, the clinical performance of the rapid antigen test is good in samples with Ct values <20. The rapid antigen test should be evaluated as a reliable screening test in patients with high viral load. To the best of our knowledge, there is no other study in the literature performed with the Wesail COVID-19 rapid antigen test kit (Guangdong, China) used in our study. The fact that PPV was found to be 100% even at a low prevalence period of the pandemic will enable positive patients to be screened quickly and effectively with rapid antigen tests in the first step during the high prevalence period of the pandemic. In the light of these data and our results, it can be predicted that using the rapid antigen test as a screening test in the first step and confirming only negative patients with RT-PCR will contribute to the effective management of the pandemic process in terms of both time and cost. As a result of the study, the rapid antigen test with low sensitivity but high PPD can be included as a facilitating test in the first step of the diagnostic algorithm in terms of rapid identification of the patients with high viral load, initiation of treatment and providing filiation.

Evaluation of Aspergillus Lateral-Flow Test in Serum Samples of Pediatric Patients.

BACKGROUND: Diagnosis of invasive aspergillosis (IA) in patients with hematologic malignancies and under the risk of IA may be uncertain or may delay because of nonspecific clinical presentation of the patients and difficult application techniques of conventional methods. Early diagnosis can provide initial antifungal therapy and prevent high mortality. In this study, we investigated the performance of an Aspergillus lateral-flow device (LFD) test (OLM Diagnostics, Newcastle upon Tyne, United Kingdom) for the diagnosis of IA in pediatric febrile neutropenic patients with hematologic malignancies. METHODS: Three hundred and fourty seven serum samples of 26 febrile neutropenic episodes of 21 patients at risk for IA were tested. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy of the Aspergillus LFD test at episode level and at serum level were calculated. RESULTS: According to the reference diagnostic criteria of IA, one proven and 13 probable IA episodes were defined. Twelve episodes (46.1%) did not meet the criteria for IA. The sensitivity, specificity, PPV, NPV, accuracy of the Aspergillus LFD test at episode level and at serum level were 14.3%, 100%, 100%, 50%, 53.8% and 12.1%, 100%, 100%, 50.8%, 53.9%, respectively. CONCLUSIONS: Aspergillus LFD test is an easy-to-use assay with short hands-on time; however, further study of the clinical utility in children and especially in serum samples are needed. It is a highly specific test for IA on bronchoalveolar lavage (BAL) samples but is not useful as a screening test for serum samples unless combined with galactomannan (GM) antigen test because of its potentially suboptimal sensitivity.

[Distribution of Hepatitis C Virus (HCV) Genotypes Among Intravenous Drug and Non-Drug User Patients]. / Damar Içi Madde Bagimliligi Olan ve Madde Bagimlisi Olmayan Hastalar Arasinda Hepatit C Virus (HCV) Genotiplerinin Dagilimi.

Genotype distribution of hepatitis C virus (HCV) can vary over the years between different patient groups and regions. The prevalence of intravenous drug users (IVDU) is known to increase in our country, yet there are a limited number of studies investigating the distribution of HCV genotypes in this group. These data are essential for monitorization of the changes in HCV epidemiology. The present study aimed to evaluate the five-year results of HCV genotyping among patients infected with HCV related to IVDU and unrelated to drug use. Plasma samples of 720 patients (HCV antibody, HCV RNA positive), which were sent to our laboratory for HCV genotyping between January 2014-March 2019 were analyzed. HCV RNA extraction from plasma samples was performed in the automated-extraction system of EZ1 advanced (Qiagen, Germany) using the EZ1 virus mini kit v2.0 (Qiagen, Germany). Amplicons were obtained by amplifying the 5'NCR and core gene region in the Rotorgene 6000 real-time PCR (Qiagen, Germany) device with the HCV RNA real-time quantitative 2.0 (NLM, Italy) kit. For the genotyping, a commercial line probe assay (LIPA) based on in vitro reverse hybridization GEN-C2.0 kit (NLM, Italy) which can distinguish 1, 2, 3, 4, 6 genotypes and 1a, 1b, 2a/c, 2b, 3a, 3b, 3c, 3k, 4a, 4b, 4c/d, 4e, 4f, 4h, 5a, 6a/b, 6g, 6f/q, 6m, 7a subtypes of HCV, based on variations in the 5'-NCR and core regions was used. HCV genotype distribution of 266 IVDU (93.2%: male; median age: 25 ± 6.82) and 454 non-drug users (51.3%: male; median age: 56.5 ± 16.06) were examined. In order of frequency in the group with IVDU; genotype 1a, 3a, 1b, 4c/d, 2b, 4, 3 were observed and genotype 1, 2a/c and mixed genotype (1+3a) were detected in one patient. In the group without IVDU, in order of frequency; genotype 1b, 1a, 3a, 1, 2a/c, 4 were observed and genotype 2b, 4c/d, 5a, 6a/b, 6 and mixed genotype (3+4) were detected in one patient. Genotypes 1a and 3a were significantly higher in the IVDU group (p< 0.00001, p< 0.00001), while 1b was significantly higher in patients without IVDU (p< 0.00001). Genotypes 1a and 3a were more common in young men (p< 0.00001, p= 0.000163), while 1b was higher in middleaged women (p< 0.00001). The incidence of genotypes 1b (p= 0.021) and 3a (p= 0.012) was higher in foreign nationals than the Turkish patients. When the HCV genotype distribution was examined by years, it was observed that the percentages of genotype 1b and 1a were decreasing, while the percentage of genotype 3a was increasing. As a result, in this study, HCV genotype distribution among IVDU was observed to be different from the general population without IVDU. It was found that genotypes 1a and 3a were more common in the IVDU group. As in the other regions of our country, genotype 1b was found most frequently in the general population. Genotype 3a increases significantly compared to years. In our study, the determination of genotypes existing in different parts of the world may be due to the foreign nationals living in our city and our region is a tourism center. It is also necessary to investigate whether there is an increase in IVDU over the years.

Investigation of Torque Teno Virus (TTV) DNA as an immunological and virological marker in pediatric hematopoietic stem cell transplantation (HSCT) patients.

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.

Evaluation of Commercially Available Real-Time Polymerase Chain Reaction Assays for the Diagnosis of Invasive Aspergillosis in Patients with Haematological Malignancies.

Early diagnosis of invasive aspergillosis (IA) is a challenge. Non-specific clinical and radiologic findings, as well as difficulties in conventional diagnostic method application, may delay correct diagnosis. Nowadays, nucleic acid-based assays have reduced the need for conventional antigen detection and culture-based methods and provided new opportunities for patient care. Aspergillus PCR is now included in the latest European Cancer Research and Treatment Organization/Mycosis Study Group definition updates. We evaluated the performance of commercial real-time polymerase chain reaction (PCR) MycAssay Aspergillus PCR and Artus Aspergillus RG PCR assays and compared the results with galactomannan enzyme immunoassay. During 41 febrile neutropenic episodes, 168 serum samples were collected from 32 patients with haematological malignancies. IA diagnosis was established according to the revised guidelines of the European Organization for Research and Treatment of Cancer/Mycoses Study Group. Twenty-one probable episodes were identified. There were no proven IA cases in the study. In 20 episodes, patients did not fulfil the established criteria for the IA diagnosis. Artus Aspergillus RG PCR assay had a sensitivity of 47.6% and specificity of 100%, while those of MycAssay Aspergillus PCR were 61.9% and 100%, respectively. Two different PCR assays were used in this study. Although there are many studies that evaluated MycAssay Aspergillus PCR, data regarding Artus Aspergillus RG PCR assay are scarce. We found moderate sensitivity and high specificity in the diagnosis of IA in patients with haematological malignancy in both PCR methods. Our results demonstrated that commercial PCR assays can be applied for the early diagnosis and pre-emptive treatment of IA.

[Comparison of Two Commercial Quantitative Cytomegalovirus (CMV) Polymerase Chain Reaction Tests Calibrated by World Health Organization International CMV Standard]. / Dünya Saglik Örgütü Uluslararasi Sitomegalovirüs (CMV) Standardi ile Kalibre Edilmis Iki Ticari Kantitatif CMV Polimeraz Zincir Reaksiyonu Testinin Karsilastirilmasi.

Cytomegalovirus (CMV) viral load quantitation is important in diagnosis, follow-up, and monitoring of antiviral therapy in transplanted patients. In this study, it was aimed to compare the results of the two commercial World Health Organization (WHO) International CMV standard calibrated polymerase chain reaction tests, CMV Cobas Ampliprep/Cobas Taqman (CMV-CAP/CTM) (Roche, Germany) and Artus CMV QIASymphony-Rotorgene (CMV-QS-RGQ) (Qiagen, Germany). Both tests were performed simultaneously on 244 plasma samples. The results were measured in copies/ml and converted to IU/ml by multiplying with 0.91 for CMV-CAP/CTM and 1.64 for CMV-QS-RGQ, as specified by the manufacturers. CMV DNA was detected in 174 (71.3%) and was not detected in 52 (21.3%) of the samples and eighteen (7.4%) samples had discordant results by both of the tests. In 16 out of 18 samples with discordance, the viral load was below the dynamic measuring ranges of both tests. In one sample, CMV DNA could not be detected by CMV-CAP/CTM but detected by CMV-QS-RGQ with 497 copies/ml, and 334 copies/ml CMV DNA was detected by CMV-CAP/CTM in another sample where it could not be detected by CMV-QS-RGQ. A high degree of agreement was found between the qualitative results of the both tests (kappa= 0.80, p< 0.001). For quantitative results in the dynamic measuring range of both assays (n= 129), the median viral load values measured by CMV-CAP/CTM and CMV-QS-RGQ were 1140 copies/ml (range: 151-254000) and 1826 copies/ml (range: 189-551521). When the results were converted to IU/ ml, the median viral load values measured by CMV-CAP/CTM and CMV-QS-RGQ were 1037 IU/ml (range: 137-231140) and 2993 IU/ml (range: 310-904133), respectively. There was a very strong correlation (r= 0.94, p< 0.001; r= 0.94, p< 0.001, respectively) between the log10 values of the quantitative results in the dynamic measuring ranges (n= 129) as copies/ml and IU/ml of both tests. CMV-QS-RGQ values corresponding to 150, 1000, 3000 copies/ml in CMV-CAP/CTM were as 94.5, 1571, 323.5 copies/ml and CMV-QS-RGQ values corresponding to 137, 910, 2730 IU/ml in CMV CAP/CTM were as 154, 2557.6, 6965.9 IU/ml, respectively. A variation of 0.45 log10 was determined between these values. In a total of 131 samples; 129 of them with the result of both tests in the dynamic measuring range and two of them which CMV DNA was not detected in one of the tests; it was found that 112 (85.5%) results for copy/ ml, 73 (56%) results for IU/ml were within the measurement difference of ± 0.5 log10 and 19 (14.5%) results for copy/ml and 58 (44%) results for IU/ml were greater than ± 0.5 log10. Bland-Altman analysis showed that CMV-CAP/CTM test made lower measurements than CMV-QS-RGQ and the average difference for copy/ml and IU/ml results were 0.22 log10 copies/ml and 0.47 log10 IU/ml. In conclusion; when the results were converted to IU/ml, the number of samples with an acceptable measurement difference between the two test results (≤ 0.5 log10) decreased and the number of samples with a measurement difference > 0.5 log10 increased and the difference was found as statistically significant (p< 0.001). Calibrating the Roche CMV CAP/CTM and Artus CMV-QS-RGQ tests with the WHO international CMV standard did not increase comparability between quantitative results in plasma samples, on the contrary, it was found that when the results were converted to IU/ml, a measurement difference indicating biologically significant viral replication was detected between the two test results.

[Evaluation of the Two Different Real Time Polymerase Chain Reaction Methods Used for BK Virus (BKV) Quantification and BKV Genotype Assignment]. / BK Virüs (BKV) Kantitasyonunda Kullanilan Iki Farkli Gerçek Zamanli Polimeraz Zincir Reaksiyonu Yönteminin Karsilastirmasi ve BKV Genotip Tayini.

BK virus (BKV) viral load quantification has a distinct role in the clinical control of BKV nephropathy and organ rejection among renal transplant recipients. In this study, it was aimed to compare BKV DNA measurement values performed with two different real-time polymerase chain reaction (PCR) methods and to determine BKV genotypes in renal transplant recipients. Totally, 150 clinical samples tested previously in two different laboratories (Lab-1 and Lab-2) from adult and pediatric renal transplantation patients were included in the study. Fifty plasma samples of 50 different patients from Lab-1, 50 plasma and 50 urine samples of 58 different patients from Lab-2 were included in the study. Viral nucleic acid extraction was performed with automatized systems in Lab-1 and Lab-2 (EZ1, Qiagen, Germany and MagNA Pure 96, Roche Diagnostics, Germany; respectively;). Real-time PCR procedure was carried out in Lab-1 with an amplification mixture of primer, probe sequences targeting VP-1 gene region using RotorGene (Qiagen, Germany) and in Lab-2 with an amplification mixture of primer, probe sequences targeting VP-2 gene region using ABI Prism 7500 (Applied Biosystems, USA). BKV genotyping was performed with multiplex PCR using primer, probe sequences for BKV genotypes I-IV. In both of the laboratories, 82 (54.6%) of the samples were found as positive, 37(24.6%) samples were found as negative and a moderate agreement was found between qualitative results of two real-time PCR methods (ƙ= 0.56, p<0.001). Median viral load values were 4.1 x 104 copies/ml (321-6 x 109) in Lab-1 and 3.3 x 105 copies/ml (224-8.3 x 1010) in Lab-2 for positive samples. According to the lineer regression analysis of quantitative results, moderate (R2= 0.52, p<0.001) and high (R2= 0.88, p<0.001) correlation was found for plasma (n= 52) and urine (n= 30) samples, respectively. Bland-Altman analysis yielded a mean difference of -0.58 log10 for all samples. For plasma samples mean difference was -0.29 log10, while it was -1.1 log10 for urine samples. In all samples, Lab-1 measurements were lower than Lab-2 measurements. A mean difference of -1.1 log10 indicated that the measurement values of Lab-2 were more higher than Lab-1 measurments with an average of 1.1 log10. Supporting this result, 71.9% of the samples had a measurement difference more than 0.5 log 10 and 29.2% of the samples had a measurement difference more than 1 log10. Only 28.1% of the samples were measured within clinically acceptable log difference range (less than 0.5 log10). BKV genotyping was performed only for 74 different patient samples with sufficient copy numbers and genotype I (81.7%), IV (15.5%), II (1.4%), I+IV (1.4%) were detected. When the results were compared; 66.6% (n= 12) of the genotype IV samples had more than 1 log10 and 83.3% of them had more than 0.5 log10 viral load measurement difference. Correlation and linear regression analyzes were insufficient for the comparison ofthe results of the two different tests. It will be appropriate for each center to monitor patients with the same test until the international BKV standard developed by the World Health Organization is optimized. The clinical correlation of the tests is limited to the currently used test. The result of incorrect BKV quantification affects the clinical decision. Measurements less than the actual value will lead to the development of BKV nephropathy, and higher measurements will lead to unnecessary allograft biopsy and unnecessary reduction of immunosuppression.

Assessment of women who applied for the uterine transplant project as potential candidates for uterus transplantation.

AIM: To review the medical charts of women who applied for the uterine transplant project from June 2008 to June 2011 in our hospital retrospectively (18-40 years). METHODS: The data for 144 women were retrieved, and information was collected on the etiology of uterine factor infertility(UFI); ovarian reserve tests; and accompanying anatomic, infectious, genetic and endocrinological problems. RESULTS: There were 119 patients with primary amenorrhea and uterovaginal agenesis and 25 patients with a history of hysterectomy. The complete Müllerian agenesis patients formed the largest group of the UFI patients with better anti-Müllerian hormone levels and antral follicle count. Anatomical anomalies such as a solitary pelvic kidney may accompany Mayer-Rokitansky-Kuster-Hauser syndrome (MRKH) and impede surgery. The mean ages in MRKH, hysterectomy and complete androgen insensitivity syndrome (CAIS) cases were 24.7, 35.0 and 34.4 years, respectively. The karyotype analysis showed 46XX (MRKH) in 109 patients and 46XY (CAIS) in 10 of the primary amenorrhea patients. CONCLUSION: Hysterectomy may deteriorate ovarian blood flow and decrease ovarian reserve. Fertility preservation may be considered in young woman undergoing hysterectomy.

Evaluation of a chromogenic medium for detection of extended-spectrum-beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae strains.

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.

Bacterial and fungal secondary infections occurring in COVID-19 patients followed in intensive care: a retrospective study.

INTRODUCTION: We aimed to investigate the effects of secondary bacterial and fungal infections on patient outcomes in patients followed up in the intensive care unit (ICU) due to coronavirus disease 2019 (COVID-19). METHODOLOGY: We retrospectively analyzed reverse transcriptase polymerase chain reaction (RT-PCR) positive COVID-19 patients followed in the ICU of our hospital between March 2020 and June 2021, using the hospital information system. Demographic data, pathogens causing a secondary infection, onset time of secondary infection, and patient outcomes were recorded. RESULTS: A total of 251 RT-PCR positive patients who met the inclusion criteria were evaluated. The mean length of stay (LOS) in the ICU was 13.3 ± 9.6 days. During this period, 165 (65.7%) patients died. When blood, urine, respiratory tract, and catheter cultures were examined, the number of patients with growth in at least one culture was 129 (51.4%). There was growth in a total of 227 cultures. The highest culture positivity rate was observed in respiratory tract samples (n = 94, 41.4%). Gram-negative bacterial pathogens (n = 130, 58.4%) predominated. Candida spp. was more frequent in urine cultures. The median day of the occurrence of secondary infection was 10 (range: 6-15). Patients who developed secondary infection had a longer LOS and higher mortality rate than patients who did not (p < 0.001). CONCLUSIONS: Gram-negative secondary infections, predominantly in respiratory tract cultures, occurred in COVID-19 patients followed in the ICU. As a result, the LOS was prolonged and mortality rates increased.

Comparison of markers of ovarian reserve between patients with complete müllerian agenesis and age-matched fertile and infertile controls.

OBJECTIVE: To investigate ovarian reserve in complete müllerian agenesis (CMA) patients and to compare the ovarian reserve of CMA patients with that of age-matched fertile and infertile controls. DESIGN: Prospective cohort study. SETTING: University gynecology outpatient clinic. PATIENT(S): Fifty-eight typical CMA (type A) patients, 8 atypical CMA (type B) patients, 39 fertile patients, and 38 infertile patients were compared for ovarian reserve. INTERVENTION(S): Ovarian reserve was evaluated via antimüllerian hormone (AMH) levels and antral follicle counts (AFCs). MAIN OUTCOME MEASURE(S): Investigation of ovarian reserve in CMA patients and a comparison of the ovarian reserve of the CMA patients with that of age-matched fertile and infertile controls. RESULT(S): Fifty-eight type A and eight type B CMA patients and 39 fertile and 38 infertile control patients were assessed for ovarian reserve. The mean (±SD) ages of the type A and type B CMA patients and the fertile and infertile groups were 25.8 ± 5.3, 33.3 ± 5.9, 32.6 ± 4.8, and 33.9 ± 3.3 years, respectively. After age standardization of the groups, AMH levels and AFCs were found to be lower in the atypical CMA group. The differences in AMH levels and AFC were found to be highly significant. CONCLUSION(S): The present study showed that atypical CMA patients had decreased ovarian reserve compared with age-matched fertile and infertile controls.

Evaluation of vancomycin resistance 3 multiplexed PCR assay for detection of vancomycin-resistant enterococci from rectal swabs.

BACKGROUND: Active screening for vancomycin-resistant enterococci (VRE) using rectal specimens is recommended to limit the spread of antimicrobial resistance within certain high-risk populations. We evaluated the diagnostic performance of Vancomycin Resistance 3 Multiplexed Tandem PCR assay (AusDiagnostics, Australia), a rapid multiplex real-time PCR assay that detects vanA and/or vanB. METHODS: Two-hundred-and-eleven rectal swabs from Hematology and Oncology unit were submitted for VRE surveillance via direct detection of vanA and/or vanB by culture and by using Vancomycin Resistance 3 Multiplexed Tandem PCR assay. Enterococci were identified to the species level by using standard biochemical tests and BD Phoenix Automated Microbiology System (BD Diagnostic Systems, USA). Vancomycin susceptibility of enterococci was determined using Etest (BioMerieux, France). RESULTS: Compared to the culture method, Vancomycin Resistance 3 Multiplexed Tandem PCR assay had a sensitivity of 84.0%, specificity of 98.8%, positive predictive value (PPV) of 91.3%, and negative predictive value (NPV) of 97.6%. The assay failed to detect 18 (8.5%) specimens because of the presence of PCR inhibitors; of the remaining 193 specimens, 25 (12.9%) were positive, 23 for vanA, and 2 for vanB. Although both sensitivity and specificity for vanA VRE was 100% compared to the culture method, all vanB-positive specimens tested negative by VRE culture. CONCLUSIONS: Vancomycin Resistance 3 Multiplexed Tandem PCR assay is a rapid and laborsaving option for VRE surveillance for direct use on rectal swabs. However, the high rate of PCR failure owing to the inhibitors in the specimens and the low specificity for vanB should be considered when interpreting the results.