Genomic insights into inherited bone marrow failure syndromes in a Korean population.

Inherited bone marrow failure syndromes (IBMFS) pose significant diagnostic challenges due to overlapping symptoms and variable expressivity, despite evolving genomic insights. The study aimed to elucidate the genomic landscape among 130 Korean patients with IBMFS. We conducted targeted next-generation sequencing (NGS) and clinical exome sequencing (CES) across the cohort, complemented by whole genome sequencing (WGS) and chromosomal microarray (CMA) in 12 and 47 cases, respectively, with negative initial results. Notably, 50% (n = 65) of our cohort achieved a genomic diagnosis. Among these, 35 patients exhibited mutations associated with classic IBMFSs (n = 33) and the recently defined IBMFS, aplastic anaemia, mental retardation and dwarfism syndrome (AmeDS, n = 2). Classic IBMFSs were predominantly detected via targeted NGS (85%, n = 28) and CES (88%, n = 29), whereas AMeDS was exclusively identified through CES. Both CMA and WGS aided in identifying copy number variations (n = 2) and mutations in previously unexplored regions (n = 2). Additionally, 30 patients were diagnosed with other congenital diseases, encompassing 13 distinct entities including inherited thrombocytopenia (n = 12), myeloid neoplasms with germline predisposition (n = 8), congenital immune disorders (n = 7) and miscellaneous genomic conditions (n = 3). CES was particularly effective in revealing these diverse diagnoses. Our findings underscore the significance of comprehensive genomic analysis in IBMFS, highlighting the need for ongoing exploration in this complex field.

Frequency histograms of three high-sensitivity cardiac troponin assays in a reference population.

BACKGROUND: Cardiac troponin (cTn) values above the 99th percentile upper reference limit (URL) indicate myocardial injury. We established 99th percentile URLs for three high-sensitivity cTn (hs-cTn) assays (Beckman Coulter Access hs-cTnI, Abbott STAT hs-cTnI, and Roche Elecsys hs-cTnT) using a healthy population in Korea. METHODS: Each cTn value was measured by three assays and analyzed by dividing by gender and age. RESULTS: The frequency histograms of log-transformed cTn values for Beckman and Abbott assays exhibited a bell-shaped distribution. The 99th percentile URLs were 9.8, 17.4, and 17.3 ng/L in the total population; 10.9/9.0, 18.9/17.0, and 18.9/17.7 ng/L in the male/female population (p < 0.001 for all three assays); and 11.2/7.2, 19.9/14.5, and 22.7/9.3 ng/L in the older/younger population (p < 0.001 for all three assays) for Beckman, Abbott, and Roche assays, respectively. CONCLUSION: Among the three assays, bell-shaped distributions were observed in a frequency histogram of log-transformed cTn values for healthy population in Beckman and Abbott assays. Also, our findings show that the 99th percentile URLs for cTn levels vary not only by gender but age.

Laboratory characteristics of IgG4-related disease: A retrospective study from a single tertiary medical center.

Immunoglobulin G4-related disease (IgG4-RD) is an immune-mediated fibroinflammatory condition with unique histopathological features that can affect most organs, making diagnosis challenging. This study characterized detailed laboratory characteristics of IgG4-RD. Baseline clinical and laboratory features of 33 patients with IgG4-RD were reviewed, including serum IgG4 concentrations, serum free light chains (sFLCs), IgGĸ- and IgGλ-heavy/light chains (HLCs), capillary serum protein electrophoresis (SPE), and immunofixation electrophoresis (IFE) of IgG4 subclass. The cohort of 33 patients showed male predominance (94%), with 8 (24%) exhibiting multiple organ involvement. Most patients (88%) had an elevated IgG4 concentration, and 67% had elevated erythrocyte sedimentation rate and IgE levels. Median IgG4 concentration at baseline was significantly higher in patients with >2 organs involved than those with ≤2. Furthermore, erythrocyte sedimentation rate was significantly correlated with serum IgG4 concentrations at baseline. SPE results demonstrated polyclonal gammopathy in most patients. Half of the patients had an increased κ/λ sFLC ratio, 42% had an increased IgGκ/IgGλ HLC ratio. Most patients exhibited hypergammaglobulinemia in the anodal end of the ɤ region on SPE. This study describes detailed laboratory features of IgG4-RD. Although none of these tests are considered diagnostically sufficient by itself, the provided laboratory characteristics can increase awareness of this disorder and help distinguish it from other IgG4-RD mimics.

Diagnostic Characteristics of Urinary Red Blood Cell Distribution Incorporated in UF-5000 for Differentiation of Glomerular and Non-Glomerular Hematuria.

BACKGROUND: Automated urine sediment analysis has been developed to address the limitations of microscopic examination of dysmorphic red blood cells (RBCs). We evaluated the urinary RBC distribution (URD) parameter of a recently launched automated urinary flow cytometry analyzer, UF-5000 (Sysmex, Kobe, Japan), to differentiate glomerular hematuria (GH) from non-GH (NGH). METHODS: Samples submitted for urine sediment analysis from patients with hematuria (>20 RBCs/µL) were divided into derivation (N=156; 101 GH, 55 NGH) and validation cohorts (N=107; 60 GH, 47 NGH). The clinical diagnosis of GH or NGH was established based on clinical data review. Differences in UF-5000 parameters (URD, small RBC, lysed RBC, RBC-P70FSC, RBC-SF-FSC-W, mean forward-scattered light, and mean side-scattered light) between GH and NGH, and areas under the ROC curves (AUC) were analyzed in the derivation cohort. The derived ideal cut-off value was evaluated in the validation cohort. We applied the Kitasato criteria to compare the diagnostic performance. RESULTS: URD (%), differed significantly between GH and NGH (P<0.001) in the two cohorts. The AUC of URD was 0.814 and 0.806 in the derivation and validation cohorts, respectively. Using a cut-off of >20.1%, the sensitivity was 99.0%/89.4% and the specificity was 50.9%/63.3% in the derivation/validation cohort. When the Kitasato criteria were applied, the sensitivity and specificity were 80.2% and 52.7%, respectively. CONCLUSIONS: URD is a rapid, objective, and quantitative measure that can be used to differentiate GH and NGH.

Assessment of M-protein quantification using capillary electrophoresis and immunosubtraction-based integration in clinical samples with low M-protein concentrations.

OBJECTIVES: Quantification of monoclonal protein (M-protein) by serum protein electrophoresis (SPE) is indispensable for diagnosing and monitoring monoclonal gammopathies. However, quantification of small and beta migrating M-proteins is challenging because of overlapping non-immunoglobulin and/or polyclonal immunoglobulin protein fractions. We compared a new integration method based on immunosubtraction (IS-CE) using capillary zone electrophoresis (CZE) against the routine method, which includes a combination of perpendicular drop (0.4%), corrected perpendicular drop (1%) and tangent skimming (98.5%). DESIGN & METHODS: The proposed method of M-protein quantification involves calculating the difference in area under the curve between the SPE and a class-specific IS-CE trace. We analyzed the difference in estimated M-protein concentrations obtained with the new method and routine integration methods using 913 samples. For IgA M-proteins at < 10 g/L, the estimated M-protein concentrations were compared with the total IgA concentration. RESULTS: The median M-protein concentration of 913 consecutive samples was 6.2 g/L (IQR 2.1-14.3 g/L). The median and median % difference between the two integration methods was 0.68 g/L (IQR 0.01-1.55 g/L) and 10.9% (IQR 0.18-38.7%), showing a larger estimated M-protein concentration with the new method. More than 25% difference was observed in 38% of the samples and was associated with lower total protein concentration, lower M-protein concentration, IgA and IgM heavy chain isotypes, and beta- or beta-gamma migration. When 161 samples with IgA M-protein < 10 g/L were compared against total IgA concentration, the median bias of the new method was smaller compared to that of the existing method (-0.95 g/L vs. -1.3 g/L, P < 0.0001). CONCLUSIONS: The use of IS based integration using CZE and IS-CE is promising especially for small and beta migrating M-proteins.

Comparison of ACCEL ELISA COVID-19 with Elecsys Anti-SARS-CoV-2 for Screening of COVID-19.

OBJECTIVE: Although real-time reverse transcription-PCR (RT-PCR) is the gold standard for diagnosing coronavirus disease 2019 (COVID-19), simpler and faster antibody detection tests can be complementary for diagnosis of COVID-19. To manage the COVID-19 pandemic, the need for serologic testing has increased. In this report, the newly developed antibody detection assays ACCEL ELISA COVID-19 (ACCEL) and Elecsys anti-SARS-CoV-2 (Elecsys) were evaluated. METHODS: Serum samples submitted for routine laboratory testing were analyzed (66 and 161 PCR-positive and PCR-negative samples). After the samples were aliquoted, antibody detection tests were performed using ACCEL and Elecsys assays. RESULTS: When detection of viral RNA using RT-PCR was set as the reference method for diagnosis of COVID-19, the sensitivity was 83.3% and 75.8, and the specificity was 96.9 and 99.4% in ACCEL and Elecsys, respectively. The true positivity rates of ACCEL and Elecsys assays were 57.1%/42.9%, 57.1%/28.6%, 77.8%/66.7%, and 97.1%/97.1% among the specimens collected ≤3, 4-7, 8-14, and >14 days after symptom onset, respectively. CONCLUSIONS: The ACCEL assay showed high sensitivity in samples collected within 7 days after symptom onset. Because many patients are asymptomatic in the early stage of SARS-CoV-2 infection, the ACCEL assay could be a good screening tool due to high sensitivity in the early stage of SARS-CoV-2 infection.

Body fluid matrix effect evaluation on the Hitachi Labospect 008 system.

BACKGROUND: Non-standard body fluids (NSBFs) can provide essential clinical information otherwise unobtainable with conventional biological specimens. However, as most commercial chemistry reagents are only validated for serum, plasma, and urine by manufacturers, individual laboratories have to validate testing with NSBF to comply with regulatory standards. However, the heightened level of oversight and uncertainty of validation requirements to comply with regulatory standards pose a significant challenge for NSBF testing in clinical laboratories. METHODS: 28 combinations of high-volume chemistry tests requested on NSBF with established clinical utility were selected from retrospective data analysis. Specimens were analyzed with both closed and open channel chemistry reagents on a LABOSPECT 008AS platform (Hitachi High-Tech Co., Tokyo, Japan). Recovery studies were performed using a high concentration serum sample and 5 clinical NSBF samples at varying concentrations for each analyte. Acceptable performance limits were defined as 100 ± 10% of expected recovery. RESULTS: The average percent recovery ranged from 94.5% to 106.6% depending on the analyte/NSBF combination evaluated, and for each of the 28 combinations, the average percent recovery was within the predefined acceptable limits of ± 10%. CONCLUSIONS: The recovery results from this study on the LABOSPECT 008AS platform demonstrates that any systematic matrix interference of high-volume chemistry testing on NSBF samples is well within the defined limits of acceptability. This work also demonstrates recovery studies performed by an individual laboratory are practial and it is feasible to demonstrate compliance with regulatory requirements for accuracy of chemistry testing on NSBF samples.

Non-inferior long-term outcomes of adults with Philadelphia chromosome-like acute lymphoblastic leukemia.

Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) is associated with inferior outcomes in the chemotherapy setting. We hypothesized that allogeneic hematopoietic cell transplantation (allo-HCT)-based post-remission therapy would improve outcomes of this entity. We examined the frequency and long-term outcomes of adults with Ph-like ALL, particularly focusing on allo-HCT outcomes for Ph-like ALL versus non-Ph-like ALL. Ph-like ALL was determined by anchored multiplex PCR-based targeted next-generation sequencing. Of the 344 patients, 57 (16.6%) had Ph-like ALL, 197 (57.3%) had Ph-positive ALL, and 90 (26.1%) had B-other ALL. To further evaluate the prognosis of Ph-like ALL, outcome analyses were restricted to 147 patients, excluding Ph-positive ALL. The actual allo-HCT rates in complete remission were 87.7% for Ph-like ALL, 71.4% for B-other standard-risk ALL, and 70.4% for B-other poor-risk ALL. Patients with Ph-like ALL had a higher 5-year overall survival (60.6% vs 27.1%; P = 0.008) than B-other poor-risk ALL subgroup, while no difference was observed compared with B-other standard-risk ALL subgroup. Similar results were noted in a separate analysis for patients receiving allo-HCT in complete remission. In multivariate analyses, B-other poor-risk ALL was associated with poorer outcomes. Our data showed that allo-HCT-based post-remission therapy may have contributed to non-inferior outcomes of adult Ph-like ALL.

Quality Improvements in Management of Children with Acute Diarrhea Using a Multiplex-PCR-Based Gastrointestinal Pathogen Panel.

Conventional methods for etiologic diagnoses of acute gastroenteritis (AGE) are time consuming and have low positive yield leading to limited clinical value. This study aimed to investigate quality improvements in patient management, antibiotic stewardship, and in-hospital infection transmission prevention using BioFire® FilmArray® Gastrointestinal Panel (GI Panel) in children with acute diarrhea. This was a prospective study recruiting children < 19 years old with new onset diarrhea during the study period, and a matched historical cohort study of children diagnosed with AGE during the 4 years prior. Patients in the prospective cohort underwent stool testing with GI Panel and conventional methods. A total of 182 patients were included in the prospective cohort, of which 85.7% (n = 156) had community-onset and 14.3% (n = 26) had hospital-onset diarrhea. A higher pathogen positivity rate for community-onset diarrhea was observed by the GI Panel (58.3%, n = 91) compared to conventional studies (42.3%, n = 66) (p = 0.005) and historical cohort (31.4%, n = 49) (p < 0.001). The stool tests reporting time after admission was 25 (interquartile range, IQR 17-46) hours for the GI Panel, and 72 (IQR 48-96) hours for the historical cohort (p < 0.001). A significant reduction in antibiotic use was observed in the prospective cohort compared to historical cohort, 35.3% vs. 71.8%; p < 0.001), respectively. Compared to the GI Panel, norovirus ICT was only able to detect 4/11 (36.4%) patients with hospital-onset and 14/27 (51.8%) patients with community-onset diarrhea. The high positivity rate and rapid reporting time of the GI Panel had clinical benefits for children admitted for acute diarrhea, especially by reducing antibiotic use and enabling early adequate infection precaution and isolation.

Evaluation of the BioFire Gastrointestinal Panel to Detect Diarrheal Pathogens in Pediatric Patients.

Infectious diarrhea is a global pediatric health concern; therefore, rapid and accurate detection of enteropathogens is vital. We evaluated the BioFire® FilmArray® Gastrointestinal (GI) Panel with that of comparator laboratory tests. Stool samples of pediatric patients with diarrhea were prospectively collected and tested. As a comparator method for bacteria, culture, conventional PCR for diarrheagenic E. coli, and Allplex GI-Bacteria(I) Assay were tested. For discrepancy analysis, BD MAX Enteric Bacterial Panel was used. As a comparator method for virus, BD MAX Enteric Virus Panel and immunochromatography was used and Allplex GI-Virus Assay was used for discrepancy analysis. The "true positive" was defined as culture-positive and/or positive results from more than two molecular tests. Of the 184 stool samples tested, 93 (50.5%) were true positive for 128 pathogens, and 31 (16.9%) were positive for multiple pathogens. The BioFire GI Panel detected 123 pathogens in 90 of samples. The BioFire GI Panel demonstrated a sensitivity of 100% for 12 targets and a specificity of >95% for 16 targets. The overall positive rate and multiple pathogen rate among patients in the group without underlying diseases were significantly higher than those in the group with hematologic disease (57.0% vs. 28.6% (p = 0.001) and 20.4% vs. 4.8% (p = 0.02), respectively). The BioFire GI Panel provides comprehensive results within 2 h and may be useful for the rapid identification of enteropathogens.

Performance evaluation of automated BD Phoenix NMIC-500 panel for carbapenemase detection in carbapenem-resistant and carbapenem-susceptible Enterobacterales.

Rapid detection of carbapenemases and accurate reporting of carbapenem MICs is critical for appropriate treatment and infection control. We evaluated the BD Phoenix NMIC-500 panel for detection and classification of carbapenemases and antimicrobial susceptibility testing (AST) for carbapenems. A total of 235 isolates were tested; 47 carbapenemase-producing Enterobacterales, 52 non-carbapenemase-producing carbapenem-resistant Enterobacterales (non-CP-CRE), 136 carbapenem-susceptible Enterobacterales (CSE). The sensitivity of carbapenemase-producing organism (CPO) detection was 97.9%, the specificity was 100% for CSE but 32.7% for non-CP-CREs. All the 35 false-positive cases were non-CP-CREs; 23 out of the 35 were determined as untyped carbapenemase producer (CP), nine were mistyped as class B, and three were as class A. The detection rate/correct classification rate for class A, B, and D carbapenemase was 100%/78.6%, 100%/100%, and 80%/60%, respectively. To supplement the low specificity, it is suggested to report carbapenemase-producer (CP) positive results as "strongly suspicious for carbapenem resistance but carbapenemase production needs to be confirmed" and perform the confirmatory test. The EA and CA for ertapenem, imipenem, and meropenem was 99.1%/99.6%, 89.4%/90.6%, and 95.3%/95.7%. In conclusion, the BD Phoenix CPO detect panel provides advantage in that the carbapenemase test is automated and the results can be obtained within 6 h but the low specificity in CREs needs to be improved. In addition, accurate reporting of meropenem MICs will be helpful for clinicians to choose treatment options.

High incidence of RAS pathway mutations among sentinel genetic lesions of Korean pediatric BCR-ABL1-like acute lymphoblastic leukemia.

INTRODUCTION: Recent advances in genetic analysis have led to the discovery of novel genetic subtypes of precursor B-cell acute lymphoblastic leukemia (B-ALL) with prognostic relevance. In this study, we studied a cohort of pediatric B-ALL patients to retrospectively determine the incidence of patients harboring novel genetic subtypes, as well as their outcome. METHODS: B-ALL patients (N = 190) diagnosed in a single Korean hospital were included in the study. Patients' medical records were reviewed for data on established genetic abnormalities and outcome. CRLF2 expression was analyzed by quantitative RT-PCR. Anchored multiplex PCR-based enrichment was used to detect fusions and point mutations in 81 ALL-related genes. RESULTS: Incidence of established recurrent genetic subtypes was as follows: high hyperdiploidy (21.6%), ETV6-RUNX1 (21.6%), BCR-ABL1 (7.9%), KMT2A rearrangement (7.4%) TCF3-PBX1/TCF3-HLF (7.4%), and hypodiploidy (1.1%). Incidence of new genetic subtypes was as follows: BCR-ABL1-like (13.2%), ETV6-RUNX1-like (2.1%), EWSR1-ZNF384 (1.1%), and iAMP21 (1.1%). Median age at diagnosis of BCR-ABL1-like ALL was 6.8 years. According to type of genetic abnormality, BCR-ABL1-like ALL was divided into ABL class (12%), CRLF2 class (8%), JAK-STAT class (12%), and RAS class (68%). The 5-year event-free survival (EFS) of BCR-ABL1-like patients was significantly inferior to non-BCR-ABL1-like low- and standard-risk patients (71.5 ± 9.1% vs 92.5 ± 3.2%, P = .001) and comparable to non-BCR-ABL1-like high (75.2 ± 6.2%) and very high-risk patients (56.8 ± 7.4%). All four ETV6-RUNX1-like patients survived event-free. CONCLUSION: Analogous to previous studies, incidence of BCR-ABL1-like ALL in our cohort was 13.2% with outcome comparable to high and very high-risk patients. A significantly high number of RAS class mutations was a distinct feature of our BCR-ABL1-like ALL group.

Performance Evaluation of the Beckman Coulter DxN VERIS Hepatitis B Virus (HBV) Assay in Comparison With the Abbott RealTime HBV Assay.

The detection and quantification of hepatitis B virus (HBV) DNA plays an important role in diagnosing and monitoring HBV infection as well as in assessing the therapeutic response. We compared the analytical performance of a random access, fully automated HBV assay-DxN VERIS Molecular Diagnostics System (Beckman Coulter, Brea, CA, USA)-with that of Abbott RealTime HBV assay (Abbott Laboratories, Des Plaines, IL, USA). The between-day precision of the VERIS assay ranged from 0.92% (mean 4.68 log IU/mL) to 4.15% (mean 2.09 log IU/mL) for pooled sera from HBV patients. HBV DNA levels measured by the VERIS HBV assay correlated with the calculated HBV DNA levels (r²=0.9994; P<0.0001). The lower limit of quantification was estimated as 8.76 IU/mL (Probit analysis, 95% confidence interval: 7.32-12.00 IU/mL). Passing-Bablok regression analysis showed good concordance between the VERIS and RealTime assays for 187 chronic HBV samples (y=-0.2397+0.9712x; r=0.981), as well as for 20 drug-resistant HBV genotype C positive samples (y=-0.5415+0.9954x; r=0.961). The VERIS assay demonstrated performance similar to the RealTime assay and is suitable for high-throughput HBV DNA monitoring in large hospital laboratories.

Comparison of Enterococcus faecium Bacteremic Isolates from Hematologic and Non-hematologic Patients: Differences in Antimicrobial Resistance and Molecular Characteristics.

BACKGROUND: Enterococcus faecium, especially vancomycin-resistant E. faecium (VREfm), is a major concern for patients with hematologic diseases. Exposure to antibiotics including fluoroquinolone, which is used as a routine prophylaxis for patients with hematologic (MH) diseases, has been reported to be a risk factor for infection with vancomycin-resistant eneterocci. We compared the characteristics of E. faecium isolates according to their vancomycin susceptibility and patient group (MH vs non-MH patients). METHODS: A total of 120 E. faecium bacteremic isolates (84 from MH and 36 from non-MH patients) were collected consecutively, and their characteristics (susceptibility, multilocus sequence type [MLST], Tn1546 type, and the presence of virulence genes and plasmids) were determined. RESULTS: Among the vancomycin-susceptible E. faecium (VSEfm) isolates, resistance to ampicillin (97.6% vs 61.1%) and high-level gentamicin (71.4% vs 38.9%) was significantly higher in isolates from MH patients than in those from non-MH patients. Notably, hyl, esp, and pEF1071 were present only in isolates with ampicillin resistance. Among the VREfm isolates, ST230 (33.3%) and ST17 (26.2%) were predominant in MH patients, while ST17 (61.1%) was predominant in non-MH patients. Plasmid pLG1 was more prevalent in E. faecium isolates from MH patients than in those from non-MH patients, regardless of vancomycin resistance. Transposon analysis revealed five types across all VREfm isolates. CONCLUSIONS: The antimicrobial resistance profiles and molecular characteristics of E. faecium isolates differed according to the underlying diseases of patients within the same hospital. We hypothesize that the prophylactic use of fluoroquinolone might have an effect on these differences.