Performance evaluation of the SMG HHV-6 Q Real-Time PCR Kit for quantitative detection and differentiation of human herpesvirus 6A and 6B.

The aim of this study was to compare the performance of the newly developed SMG HHV-6 Q Real-Time PCR Kit (SMG assay) with the RealStar HHV-6 PCR Kit (RealStar assay). The analytical sensitivity and specificity, linearity, and precision of the SMG assay were evaluated. The clinical performance of the SMG assay was assessed and compared with that of the RealStar assay using 207 clinical specimens (HHV-6A positive, n = 51; HHV-6B positive, n = 64; HHV-6A/B negative, n = 92). The limit of detection of the SMG assay was 2.92 log10 copies/mL for HHV-6A DNA and 2.88 log10 copies/mL for HHV-6B DNA. The linear range was determined to be 3.40-9.00 log10 copies/mL for both viruses. Intra- and inter-assay variability were below 5% at concentrations ranging from 4 to 9 log10 copies/mL. No cross-reactivity was observed with the 25 microorganisms included in the specificity panel. The clinical sensitivity and specificity of the SMG and RealStar assays compared to in-house polymerase chain reaction and sequencing were as follows: SMG assay, 98.0% and 100% for HHV-6A DNA, respectively, and 96.9% and 100% for HHV-6B DNA, respectively; RealStar assay, 98.0% and 100% for HHV-6A DNA, respectively, and 90.6% and 100% for HHV-6B DNA, respectively. The correlation coefficients between viral loads measured by the two assays were 0.948 and 0.975, with mean differences of 0.62 and 0.32 log10 copies/mL for HHV-6A and HHV-6B DNA, respectively. These results demonstrate that the SMG assay is a sensitive and reliable tool for the quantitative detection and differentiation of HHV-6A and HHV-6B DNA.IMPORTANCEQuantitative real-time PCR (qPCR) that can distinguish between HHV-6A and HHV-6B DNA is recommended for diagnosis of active infection. The SMG HHV-6 Q Real-Time PCR Kit (SMG assay) is a newly developed qPCR assay that can differentiate between HHV-6A and HHV-6B DNA; however, little is known about its performance. In this study, we assessed the performance of the SMG assay and compared it with that of a commercially available qPCR assay, the RealStar HHV-6 PCR Kit (RealStar assay). The SMG assay demonstrated excellent analytical sensitivity and specificity, precision, and linearity. Furthermore, the viral loads measured by the SMG assay were highly correlated with those measured by the RealStar assay. Our results suggest that the SMG assay is a useful diagnostic tool for quantitative detection and differentiation of HHV-6A and HHV-6B DNA.

Optimization of extraction-free protocols for SARS-CoV-2 detection using a commercial rRT-PCR assay.

In the ongoing global fight against coronavirus disease 2019 (COVID-19), the sample preparation process for real-time reverse transcription polymerase chain reaction (rRT-PCR) faces challenges due to time-consuming steps, labor-intensive procedures, contamination risks, resource demands, and environmental implications. However, optimized strategies for sample preparation have been poorly investigated, and the combination of RNase inhibitors and Proteinase K has been rarely considered. Hence, we investigated combinations of several extraction-free protocols incorporating heat treatment, sample dilution, and Proteinase K and RNase inhibitors, and validated the effectiveness using 120 SARS-CoV-2 positive and 62 negative clinical samples. Combining sample dilution and heat treatment with Proteinase K and RNase inhibitors addition exhibited the highest sensitivity (84.26%) with a mean increase in cycle threshold (Ct) value of + 3.8. Meanwhile, combined sample dilution and heat treatment exhibited a sensitivity of 79.63%, accounting for a 38% increase compared to heat treatment alone. Our findings highlight that the incorporation of Proteinase K and RNase inhibitors with sample dilution and heat treatment contributed only marginally to the improvement without yielding statistically significant differences. Sample dilution significantly impacts SARS-CoV-2 detection, and sample conditions play a crucial role in the efficiency of extraction-free methods. Our findings may provide insights for streamlining diagnostic testing, enhancing its accessibility, cost-effectiveness, and sustainability.

Comparison of the STANDARD F and SD BIOLINE stool antigen tests for diagnosis of Helicobacter pylori infection.

We compared the performance of the STANDARD F and SD BIOLINE stool antigen tests in 335 patients. The performance of STANDARD F (sensitivity: 95.6%; specificity: 94%) was highly comparable to that of SD BIOLINE (sensitivity: 92.6%; specificity: 93.5%), suggesting that STANDARD F is useful for the detection of Helicobacter pylori infection.

Comparison between the STANDARD™ F S. Pneumoniae Ag FIA and BinaxNOW S. Pneumoniae Antigen Card for Detection of Streptococcus Pneumoniae Urinary Antigen.

We compared the performance of STANDARD F S. pneumoniae Ag FIA with that of BinaxNOW S. pneumoniae Antigen Card using 206 urine samples. The performance of STANDARD F was highly comparable to that of BinaxNOW. STANDARD F assay could be a valuable tool for diagnosis of invasive pneumococcal disease.

Performance Evaluation of the PowerChek SARS-CoV-2, Influenza A & B Multiplex Real-Time PCR Kit in Comparison with the BioFire Respiratory Panel.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and influenza viruses may pose enormous challenges to our healthcare system. We evaluated the performance of the PowerChek SARS-CoV-2, Influenza A & B Multiplex Real-time PCR Kit (PowerChek; Kogene Biotech, Seoul, Korea) in comparison with the BioFire Respiratory Panels 2 and 2.1 (RP2 and RP2.1; bioMérieux, Marcy l'Étoile, France), using 147 nasopharyngeal swabs. The limit of detection (LOD) of the PowerChek assay was determined using SARS-CoV-2, influenza A, and B RNA standards. The LOD values of the PowerChek assay for SARS-CoV-2 and influenza A and B were 1.12, 1.24, and 0.61 copies/µL, respectively. The positive and negative percent agreements of the PowerChek assay compared with RP2 and RP2.1 were 97.5% (39/40) and 100% (107/107) for SARS-CoV-2; 100% (39/39) and 100% (108/108) for influenza A; and 100% (35/35) and 100% (112/112) for influenza B, respectively. The performance of the PowerChek assay was comparable to that of RP2 and RP2.1 for detecting SARS-CoV-2 and influenza A and B, suggesting its use in diagnosing SARS-CoV-2 and influenza infections.

Whole-Genome Sequencing Analysis of a stx-Negative Escherichia coli O63:H6 Isolate Associated with Hemolytic Uremic Syndrome.

Shiga toxin-encoding genes (stx) of enterohemorrhagic Escherichia coli (EHEC) can be lost during infection or in vitro cultivation, and in clinical practice, it is difficult to distinguish EHEC that have lost stx (EHEC-LST) from enteropathogenic E. coli (EPEC), as both are stx-negative and eae-positive. In this study, we performed whole-genome sequencing (WGS) of a stx-negative, eae-positive E. coli O63:H6 isolate from a child with hemolytic uremic syndrome and compared its genome with those of nine E. coli O63:H6 strains in public databases. Virulence gene profiles were analyzed and core-genome multilocus sequence typing (cgMLST) was conducted. The virulence gene profile of our isolate was consistent with EHEC, except for the absence of stx, and the isolate clustered with seven EHEC strains but was distant from two EPEC strains in cgMLST. In genome alignment, our isolate exhibited a high nucleotide identity with EHEC strain 377323_2f but displayed a gap corresponding to the stx-harboring prophage sequence. Overall, our isolate was genetically closely related to EHEC strains, consistent with this being an EHEC-LST strain. As EHEC-LST may be misdiagnosed as EPEC in routine laboratories, comparative genomic analysis using WGS can be useful to determine whether stx-negative and eae-positive isolates are EHEC-LST or EPEC.

Comparison of the PowerChek SARS-CoV-2, Influenza A&B, RSV Multiplex Real-time PCR Kit and BioFire Respiratory Panel 2.1 for simultaneous detection of SARS-CoV-2, influenza A and B, and respiratory syncytial virus.

The potential co-circulation of SARS-CoV-2, influenza, and respiratory syncytial virus (RSV) could pose an unprecedented challenge to healthcare systems worldwide. Here, we compared the performance of the PowerChek SARS-CoV-2, Influenza A&B, RSV Multiplex Real-time PCR Kit (PowerChek) for simultaneous detection of SARS-CoV-2, influenza A and B, and respiratory syncytial virus with that of BioFire Respiratory Panel 2.1 (RP2.1) using 175 nasopharyngeal swab (NPS) specimens. Positive percent agreement and negative percent agreement of the PowerChek assay compared to RP2.1 were as follows: 100 % (40/40) and 100 % (135/135) for SARS-CoV-2; 100 % (39/39) and 100 % (136/136) for influenza A; 100 % (35/35) and 100 % (140/140) for influenza B; and 93.1 % (27/29) and 100 % (146/146) for RSV, respectively. The limit of detection (LOD) was accessed using RNA standards for each virus, and the LOD values of the PowerChek assay for SARS-CoV-2, influenza A and B, and RSV were 0.36, 1.24, 0.09, and 0.63 copies/µL, respectively. Our results demonstrate that the PowerChek assay is sensitive and accurate for detection of SARS-CoV-2, influenza A and B, and RSV, suggesting that this assay can be a valuable diagnostic tool when SARS-CoV-2, influenza, and RSV are co-circulating.

Prevalence of granular corneal dystrophy type 2-related TGFBI p.R124H variant in a South Korean population.

Purpose: Granular corneal dystrophy type 2 (GCD2) is an autosomal dominant disorder and is associated with the arginine to histidine substitution at codon 124 (p.R124H) of the TGFBI gene. Although TGFBI p.R124H is known to be the most common corneal dystrophy-related pathogenic variant, there are few data on the frequency of this variant in the South Korean population. Methods: In total, 2,060 anonymous DNA samples from a public umbilical cord blood bank were tested for the TFGBI p.R124H variant using real-time PCR. Results: Six of the 2,060 samples [0.29%; 95% confidence interval (CI), 0.12-0.67%] were heterozygous for the TGFBI p.R124H variant. The prevalence of the GCD2-related TGFBI p.R124H variant in this population was estimated to be 291.3 per 100,000 [95% confidence interval (CI), 118.5-667.0]. Conclusions: To our knowledge, this is the largest study that has estimated the prevalence of the GCD2-related TGFBI p.R124H variant in South Korea.

Genetic Analysis of Korean Adult Patients with Nontuberculous Mycobacteria Suspected of Primary Ciliary Dyskinesia Using Whole Exome Sequencing.

PURPOSE: Nontuberculous mycobacteria (NTM) is ubiquitous in the environment, but NTM lung disease (NTM-LD) is uncommon. Since exposure to NTM is inevitable, patients who develop NTM-LD are likely to have specific susceptibility factors, such as primary ciliary dyskinesia (PCD). PCD is a genetically heterogeneous disorder of motile cilia and is characterized by chronic respiratory tract infection, organ laterality defect, and infertility. In this study, we performed whole exome sequencing (WES) and investigated the genetic characteristics of adult NTM patients with suspected PCD. MATERIALS AND METHODS: WES was performed in 13 NTM-LD patients who were suspected of having PCD by clinical symptoms and/or ultrastructural ciliary defect observed by transmission electron microscopy. A total of 45 PCD-causing genes, 23 PCD-candidate genes, and 990 ciliome genes were analyzed. RESULTS: Four patients were found to have biallelic loss-of-function (LoF) variants in the following PCD-causing genes: CCDC114, DNAH5, HYDIN, and NME5. In four other patients, only one LoF variant was identified, while the remaining five patients did not have any LoF variants. CONCLUSION: At least 30.8% of NTM-LD patients who were suspected of having PCD had biallelic LoF variants, and an additional 30.8% of patients had one LoF variant. Therefore, PCD should be considered in patients with NTM-LD with symptoms or signs suspicious of PCD.

Performance evaluation of Automated Fluorescent Immunoassay System ROTA and NORO for detection of rotavirus and norovirus: A comparative study of assay performance with RIDASCREEN® Rotavirus and Norovirus.

BACKGROUND: The Automated Fluorescent Immunoassay System ROTA (AFIAS-Rota) and NORO (AFIAS-Noro) assays (Boditech Med Inc.) are newly developed diagnostic tests for rotavirus and norovirus infections. METHODS: Performance of AFIAS-Rota/Noro assays was evaluated in comparison with RIDASCREEN® Rotavirus and Norovirus ELISA kits (R-Biopharm) using clinical stool samples submitted from November 2018 to January 2019. Multiplex real-time reverse transcription-polymerase chain reaction was used as reference method. RESULTS: A total of 256 clinical specimens were analyzed. AFIAS-Rota and RIDASCREEN Rotavirus had almost perfect agreement (Kappa value = 0.95), and substantial agreement was observed between AFIAS-Noro and RIDASCREEN Norovirus (Kappa value = 0.80). For detection of rotavirus, AFIAS and RIDASCREEN assays showed satisfactory diagnostic sensitivity (100% and 97.8%, respectively) and specificity (99.5% and 99.1%). For detection of norovirus, the RIDASCREEN assay showed significantly higher sensitivity than the AFIAS-Noro (86.0% and 66.0%, respectively; P = .002). Analytic specificity of AFIAS-Rota/Noro assays showed no cross-reactivity against any other bacteria (14 strains) or viruses (2 strains). Hands-on time (6 minutes) and turnaround time (26 minutes) required to perform AFIAS assays were much shorter than those required for RIDASCREEN assays (20 and 150 minutes, respectively). CONCLUSION: The AFIAS-Rota/Noro assays showed overall excellent agreement with the RIDASCREEN assays. Although the AFIAS-Noro assay exhibited lower sensitivity than the RIDASCREEN Norovirus assay for detection of norovirus, the AFIAS-Rota/Noro assays could be useful as a rapid initial screening test in clinical laboratories due to its convenience and rapid turnaround time.

Evaluation of the ASTA MicroIDSys matrix-assisted laser desorption ionization time-of-flight mass spectrometry system for identification of mycobacteria directly from positive MGIT liquid cultures.

OBJECTIVES: We evaluated the performance of the MicroIDSys Elite system, a newly developed matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry system for identification of mycobacteria directly from positive MGIT liquid cultures. METHODS: Analytical specificity was evaluated with 63 reference strains grown in mycobacteria growth indicator tube media. Prospective performance evaluation was conducted with primary liquid cultures of sputum samples for identification of mycobacteria, and results were compared to multigenerational sequencing as the reference method. Liquid media subcultures were also analyzed. RESULTS: The accuracy for the 63 reference strains was 98.4% (62/63). A total of 167 paired mycobacterial primary cultures and subcultures in liquid media, comprised of seven Mycobacterium tuberculosis isolates, 109 slowly growing nontuberculous mycobacterial isolates, and 51 rapidly growing nontuberculous mycobacterial isolates, was identified by the MicroIDSys Elite system. Using primary liquid cultures, the MicroIDSys Elite system correctly identified 143 (85.6%) isolates; 21 (12.6%) resulted in "no identification"; and three (1.8%) isolates were misidentified. Using liquid media subcultures with this system, 159 (95.2%) isolates were correctly identified; seven (4.2%) resulted in "no identification"; and one (0.6%) isolate was misidentified. CONCLUSION: The MicroIDSys Elite system is a useful routine diagnostic tool for identification of mycobacterial species from liquid culture.

Evaluation of the BioFire FilmArray Pneumonia Panel for rapid detection of respiratory bacterial pathogens and antibiotic resistance genes in sputum and endotracheal aspirate specimens.

OBJECTIVES: The performance of the investigational-use-only version of the BioFire FilmArray Pneumonia Panel (FA-Pneumo), a high-order nested multiplex PCR, was evaluated for the detection of typical respiratory bacterial pathogens and antibiotic resistance genes in sputa and endotracheal aspirate (ETA) specimens. METHODS: Thirty-one sputa and 69 ETA specimens were analyzed. The diagnostic performance of FA-Pneumo was assessed using routine microbiological methods as the reference standard. RESULTS: Overall sensitivity and specificity for organism detection using FA-Pneumo were 98.5% and 76.5%, respectively. The sensitivity for each pathogen was 100%, except for Klebsiella aerogenes, and the range of specificity was 83.3-99.0%. FA-Pneumo detected antimicrobial resistance genes in 17 out of 18 specimens (94.4%) that were resistant by antimicrobial susceptibility testing. FA-Pneumo additionally detected 25 resistance genes in 22 specimens, and sequencing for the presence of resistance genes confirmed the majority of these results (20/25, 80%). Semi-quantitative analysis of bacterial nucleic acid amounts by FA-Pneumo revealed that 88.2% of the identified bacteria (67/76) with ≥106 copies/ml also gave culture-positive results with significant amounts of bacteria. CONCLUSIONS: FA-Pneumo is a rapid test with high sensitivity for the detection of bacteria and antimicrobial resistance genes in sputum and ETA specimens and could aid in determining antibiotic therapy.

Carrier Frequency of Spinal Muscular Atrophy in a Large-scale Korean Population.

Spinal muscular atrophy (SMA) is an autosomal recessive disease characterized by progressive proximal muscle weakness and atrophy. Given the recent introduction of gene therapies, knowledge of the SMA carrier frequency in various populations has become important for developing screening programs for this disease. In total, 1,581 anonymous DNA samples from an umbilical cord blood bank were tested for SMN1 and SMN2 gene copies using a multiplex ligation-dependent probe amplification assay. Twenty-nine of the 1,581 newborns [1.83%; 95% confidence interval (CI), 1.25-2.66%] were SMA carriers with one copy of SMN1, and no homozygous SMN1 deletion was detected. The carrier frequency in this population was estimated to be 1,834 per 100,000 (95% CI, 1,254-2,659) or 1 in 55 (95% CI, 1/79-1/38). Our data indicate that SMA carriers are not uncommon in the Korean population and may serve as a reference for designing a population screening program in Korea.

Comparison between DiaPlexQ™ STI6 and GeneFinder™ STD I/STD II multiplex Real-time PCR Kits in the detection of six sexually transmitted disease pathogens.

BACKGROUND: The DiaPlexQ™ STI6 Detection Kit (DiaPlexQ; Solgent Co., Ltd., Daejeon, South Korea) is a multiplex real-time PCR assay for the detection of the following sexually transmitted disease (STD) pathogens: Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma hominis, Trichomonas vaginalis, Ureaplasma urealyticum, and Mycoplasma genitalium. We compared the performance of the DiaPlexQ assay with the GeneFinder™ STD I (CT/NG/UU) and STD II (MG/MH/TV) Multiplex Real-time PCR Kits (GeneFinder; Infopia Co., Ltd., Anyang, South Korea). METHODS: We evaluated the performance of the DiaPlexQ assay in comparison to that of GeneFinder using 1106 clinical specimens (542 genital swabs and 564 urine samples). The analytical performance of the DiaPlexQ assay, including the limit of detection (LOD) and analytical specificity, was evaluated using reference strains. RESULTS: The positive percent agreement, negative percent agreement, and kappa value between the two assays were 96.6%-99.4%, 98.2%-99.8%, and 0.93%-0.99%, respectively. No cross-reactivity was observed in a collection of 41 different microorganisms and the LOD of the DiaPlexQ assay ranged from 1 to 10 copies/reaction for each microorganism. CONCLUSION: The DiaPlexQ assay showed comparable performance to that of the GeneFinder assay so that it can be used for the screening and diagnosis of non-viral curable STD pathogens.

Bronchiectasis and Recurrent Respiratory Infections with a De Novo STAT1 Gain-of-Function Variant: First Case in Korea.

Bronchiectasis is a chronic disease characterized by airway infection and inflammation, leading to permanent dilation of the bronchi. Evaluation of underlying etiology is important in managing young bronchiectasis patients with recurrent infections caused by unusual pathogens. The signal transducer and activator of transcription 1 (STAT1) protein plays a key role in STAT signaling and immune system regulation. Heterozygotes for gain-of-function (GOF) alleles of the STAT1 gene usually display autosomal dominant chronic mucocutaneous candidiasis (CMC) and a wide range of clinical features, such as bronchiectasis. Here, we report on a patient with CMC and bronchiectasis with various types of infections who carried a pathogenic variant of the STAT1 gene. The 24-year-old female presented with recurrent respiratory bacterial and nontuberculous mycobacterial infections complicated by severe bronchiectasis and CMC. Whole-exome sequencing revealed a c.800C>T (p.Ala267Val) heterozygous mutation in the STAT1 gene. Further analysis by Sanger sequencing of STAT1 from the patient and her parents revealed the patient had a de novo occurrence of the variant. This is the first report of a Korean patient with a GOF pathogenic variant in STAT1. Physicians should be aware of the existence of this variant as a genetic factor associated with CMC and bronchiectasis complicated by recurrent infection.

Performance Evaluation of the PowerChek MERS (upE & ORF1a) Real-Time PCR Kit for the Detection of Middle East Respiratory Syndrome Coronavirus RNA.

BACKGROUND: Molecular detection of Middle East respiratory syndrome coronavirus (MERS-CoV) using real-time reverse transcription (rRT)-PCR assays is the method of choice for diagnosis of MERS. We evaluated the performance of the PowerChek MERS (upE & ORF1a) real-time PCR Kit (PowerChek MERS assay; Kogene Biotech, Korea) a one-step rRT-PCR assay for the qualitative detection of MERS-CoV. METHODS: We evaluated PowerChek MERS assay performance in comparison with nested RT-PCR and sequencing of the RNA-dependent RNA polymerase (RdRp) and N genes. To evaluate diagnostic sensitivity and specificity, 100 clinical specimens (50 positive and 50 negative for MERS-CoV) were simultaneously tested by using the PowerChek MERS and sequencing assays. Assay performance, including limit of detection and precision, was evaluated in vitro by using MERS-CoV RNA transcripts. Analytical specificity was evaluated with a diverse collection of 16 respiratory virus-positive clinical specimens and 14 respiratory bacterial isolates. RESULTS: The 95% limits of detection of the PowerChek MERS assay for the upE and the open rading frame (ORF)1a were 16.2 copies/µL and 8.2 copies/µL, respectively. No cross-reactivity was observed. The diagnostic sensitivity and specificity of the PowerChek MERS assay were both 100% (95% confidence interval, 91.1-100%). CONCLUSIONS: The PowerChek MERS assay is a straightforward and accurate assay for detecting MERS-CoV RNA. The assay will be a useful tool for the rapid diagnosis of MERS and could prove especially important for MERS outbreak control.

Performance Evaluation of Allplex Respiratory Panels 1, 2, and 3 for Detection of Respiratory Viruses and Influenza A Virus Subtypes.

The Allplex respiratory panels 1, 2, and 3 (Allplex) comprise a one-step real-time reverse transcription-PCR assay for the detection of respiratory viruses (RVs) and influenza A subtypes based on multiple detection temperature (MuDT) technology. The performance of the Allplex assay was compared with those of the AdvanSure RV real-time PCR kit (AdvanSure) and the PowerChek pandemic H1N1/H3N2/H5N1 real-time PCR kit (PowerChek) using 417 clinical respiratory specimens. In comparison with the AdvanSure assay for RV detection by each virus, the ranges of positive percent agreement, negative percent agreement, and kappa values with the Allplex assay were 82.8 to 100%, 95.5 to 100%, and 0.85 to 1.00, respectively. For influenza A virus (INF A) subtyping, the kappa values between the Allplex and PowerChek assays were 0.67 and 1.00 for the INF A H1N1-pdm09 and H3 subtypes, respectively. Uniplex PCR and sequencing for samples with discrepant results demonstrated that the majority of results were concordant with those from the Allplex assay. When testing 24 samples, the turnaround and hands-on time required to perform the Allplex assay were 4 h 15 min and 15 min, respectively. In conclusion, the Allplex assay produced results comparable to those from the AdvanSure and PowerChek assays.

Performance of the Real-Q EBV Quantification Kit for Epstein-Barr Virus DNA Quantification in Whole Blood.

There has been increasing interest in standardized and quantitative Epstein-Barr virus (EBV) DNA testing for the management of EBV disease. We evaluated the performance of the Real-Q EBV Quantification Kit (BioSewoom, Korea) in whole blood (WB). Nucleic acid extraction and real-time PCR were performed by using the MagNA Pure 96 (Roche Diagnostics, Germany) and 7500 Fast real-time PCR system (Applied Biosystems, USA), respectively. Assay sensitivity, linearity, and conversion factor were determined by using the World Health Organization international standard diluted in EBV-negative WB. We used 81 WB clinical specimens to compare performance of the Real-Q EBV Quantification Kit and artus EBV RG PCR Kit (Qiagen, Germany). The limit of detection (LOD) and limit of quantification (LOQ) for the Real-Q kit were 453 and 750 IU/mL, respectively. The conversion factor from EBV genomic copies to IU was 0.62. The linear range of the assay was from 750 to 106 IU/mL. Viral load values measured with the Real-Q assay were on average 0.54 log10 copies/mL higher than those measured with the artus assay. The Real-Q assay offered good analytical performance for EBV DNA quantification in WB.

Performance Evaluation of the Real-Q Cytomegalovirus (CMV) Quantification Kit Using Two Real-Time PCR Systems for Quantifying CMV DNA in Whole Blood.

Standardized cytomegalovirus (CMV) DNA quantification is important for managing CMV disease. We evaluated the performance of the Real-Q CMV Quantification Kit (Real-Q assay; BioSewoom, Korea) using whole blood (WB), with nucleic acid extraction using MagNA Pure 96 (Roche Diagnostics, Germany). Real-time PCR was performed on two platforms: the 7500 Fast real-time PCR (7500 Fast; Applied Biosystems, USA) and CFX96 real-time PCR detection (CFX96; Bio-Rad, USA) systems. The WHO international standard, diluted with CMV-negative WB, was used to validate the analytical performance. We used 90 WB clinical samples for comparison with the artus CMV RG PCR kit (artus assay; Qiagen, Germany). Limits of detections (LODs) in 7500 Fast and CFX96 were 367 and 479 IU/mL, respectively. The assay was linear from the LOD to 106 IU/mL (R² ≥0.9886). The conversion factors from copies to IU in 7500 Fast and CFX96 were 0.95 and 1.06, respectively. Compared with the artus assay, for values <1,000 copies/mL, 100% of the samples had a variation <0.7 log10 copies/mL; >1,000 copies/mL, 73.3% and 80.6% of samples in 7500 Fast and CFX96, respectively, had <0.5 log10 copies/mL. The Real-Q assay is useful for quantifying CMV in WB with the two real-time PCR platforms.