Diagnostic Approaches to Investigate JAK2-Unmutated Erythrocytosis Based on a Single Tertiary Center Experience.

INTRODUCTION: Erythrocytosis is attributed to various clinical and molecular factors. Many cases of JAK2-unmutated erythrocytosis remain undiagnosed. We investigated the characteristics and causes of JAK2-unmutated erythrocytosis. METHODS: We assessed the clinical and laboratory results of patients with erythrocytosis without JAK2 mutations and performed targeted next-generation sequencing (NGS) panels for somatic and germline mutations. RESULTS: In total, 117 patients with JAK2-unmutated erythrocytosis were included. The median hemoglobin and hematocrit levels were 17.9 g/dL and 53.4%, respectively. Erythropoietin levels were not below the reference range. Thrombotic events were reported in 17 patients (14.5%). Among JAK2-unmutated patients, 44 had undergone targeted panel sequencing consisting of myeloid neoplasm-related genes, and 16 had one or more reportable variants in ASXL1 (5/44), TET2, CALR, FLT3, and SH2B3 (2/44). Additional testing for germline causes revealed eight variants in seven genes in eight patients, including NF1, BPGM, EPAS1, PIEZO1, RHAG, SH2B3, and VHL genes. One NF1 pathogenic, one BPGM likely pathogenic, and six variants of undetermined significance were detected. CONCLUSION: Somatic and germline mutations were identified in 36.4% and 33.3 % of the JAK2-unmutated group; most variants had unknown clinical significance. Not all genetic causes have been identified; comprehensive diagnostic approaches are crucial for identifying the cause of erythrocytosis.

Evaluation of an eight marker-panel including long mononucleotide repeat markers to detect microsatellite instability in colorectal, gastric, and endometrial cancers.

BACKGROUND: Accurate determination of microsatellite instability (MSI) status is critical for optimal treatment in cancer patients. Conventional MSI markers can sometimes display subtle shifts that are difficult to interpret, especially in non-colorectal cases. We evaluated an experimental eight marker-panel including long mononucleotide repeat (LMR) markers for detection of MSI. METHODS: The eight marker-panel was comprised of five conventional markers (BAT-25, BAT-26, NR-21, NR-24, and NR-27) and three LMR markers (BAT-52, BAT-59 and BAT-62). MSI testing was performed against 300 specimens of colorectal, gastric, and endometrial cancers through PCR followed by capillary electrophoresis length analysis. RESULTS: The MSI testing with eight marker-panel showed 99.3% (295/297) concordance with IHC analysis excluding 3 MMR-focal deficient cases. The sensitivity of BAT-59 and BAT-62 was higher than or comparable to that of conventional markers in gastric and endometrial cancer. The mean shift size was larger in LMR markers compared to conventional markers for gastric and endometrial cancers. CONCLUSIONS: The MSI testing with eight maker-panel showed comparable performance with IHC analysis. The LMR markers, especially BAT-59 and BAT-62, showed high sensitivity and large shifts which can contribute to increased confidence in MSI classification, especially in gastric and endometrial cancers. Further study is needed with large number of samples for the validation of these LMR markers.

A simplified two-marker immunohistochemistry strategy for Lynch syndrome screening in endometrial cancer patients.

OBJECTIVE: To examine the efficacy of MSH6 and PMS2 immunohistochemistry (IHC) as a screening method for Lynch syndrome in endometrial cancer patients. METHODS: Through multidisciplinary discussions, an institutional MSH6 and PMS2 IHC-initiated cascade test (MSH6, PMS2 IHC→microsatellite instability [MSI] assay→germline mismatch repair [MMR] gene sequencing) was developed to screen for Lynch syndrome in endometrial cancer patients. Testing was performed on a consecutive cohort of 218 newly diagnosed endometrial cancer patients who underwent surgery at a tertiary hospital in the Republic of Korea between August 2018 and December 2020. The number of MMR deficiencies (MSH6 or PMS2 loss in IHC) and. RESULTS: of subsequent tests (MSI assay and germline MMR gene sequencing) were examined. RESULTS: MMR deficiency was detected in 52 of the 218 patients (24.0%). Among these 52 patients, 34 (65.0%) underwent MSI testing, of which 31 (91.0%) exhibited high MSI. Of the 31 patients with MSI-high status, 15 (48.0%) underwent germline MMR gene sequencing. Subsequently, Lynch syndrome was diagnosed in five patients (33.0%). CONCLUSION: Lynch syndrome screening using MSH6 and PMS2 IHC-initiated cascade testing is a viable strategy in the management of endometrial cancer. A simplified strategy (MSH6 and PMS2 IHC→germline MMR gene sequencing) was proposed because most women with MMR deficiencies exhibited high MSI.

Comprehensive Study of Microsatellite Instability Testing and Its Comparison With Immunohistochemistry in Gastric Cancers.

PURPOSE: In this study, polymerase chain reaction (PCR)-based microsatellite instability (MSI) testing was comprehensively analyzed and compared with immunohistochemistry (IHC) for mismatch repair (MMR) protein expression in patients with gastric cancer (GC). MATERIALS AND METHODS: In 5,676 GC cases, PCR-based MSI testing using five microsatellites (BAT-26, BAT-25, D5S346, D2S123, and D17S250) and IHC for MLH1 were performed. Re-evaluation of MSI testing/MLH1 IHC and additional IHC for MSH2, MSH6, and PMS2 were performed in discordant/indeterminate cases. RESULTS: Of the 5,676 cases, microsatellite stable (MSS)/MSI-low and intact MLH1 were observed in 5,082 cases (89.5%), whereas MSI-high (MSI-H) and loss of MLH1 expression were observed in 502 cases (8.8%). We re-evaluated the remaining 92 cases (1.6%) with a discordant/indeterminate status. Re-evaluation showed 1) 37 concordant cases (0.7%) (18 and 19 cases of MSI-H/MMR-deficient (dMMR) and MSS/MMR-proficient (pMMR), respectively), 2) 6 discordant cases (0.1%) (3 cases each of MSI-H/pMMR and MSS/dMMR), 3) 14 MSI indeterminate cases (0.2%) (1 case of dMMR and 13 cases of pMMR), and 4) 35 IHC indeterminate cases (0.6%) (22 and 13 cases of MSI-H and MSS, respectively). Finally, MSI-H or dMMR was observed in 549 cases (9.7%), of which 47 (0.8%) were additionally confirmed as MSI-H or dMMR by re-evaluation. Sensitivity was 99.3% for MSI testing and 95.4% for MMR IHC. CONCLUSIONS: Considering the low incidence of MSI-H or dMMR, discordant/indeterminate results were occasionally identified in GCs, in which case complementary testing is required. These findings could help improve the accuracy of MSI/MMR testing in daily practice.

Anti-Alpha-Toxin Antibody Responses and Clinical Outcomes of Staphylococcus aureus Bacteremia.

BACKGROUND: Alpha-toxin (AT), a major virulence factor of Staphylococcus aureus, is an important immunotherapeutic target to prevent or treat invasive S. aureus infections. Previous studies have suggested that anti-AT antibodies (Abs) may have a protective role against S. aureus bacteremia (SAB), but their function remains unclear. Therefore, we aimed to investigate the association between serum anti-AT Ab levels and clinical outcomes of SAB. METHODS: Patients from a prospective SAB cohort at a tertiary-care medical center (n = 51) were enrolled in the study from July 2016 to January 2019. Patients without symptoms or signs of infection were enrolled as controls (n = 100). Blood samples were collected before the onset of SAB and at 2- and 4-weeks post-bacteremia. Anti-AT immunoglobin G (IgG) levels were measured using an enzyme-linked immunosorbent assay. All clinical S. aureus isolates were tested for the presence of hla using polymerase chain reaction. RESULTS: Anti-AT IgG levels in patients with SAB before the onset of bacteremia did not differ significantly from those in non-infectious controls. Pre-bacteremic anti-AT IgG levels tended to be lower in patients with worse clinical outcomes (7-day mortality, persistent bacteremia, metastatic infection, septic shock), although the differences were not statistically significant. Patients who needed intensive care unit care had significantly lower anti-AT IgG levels at 2 weeks post-bacteremia (P = 0.020). CONCLUSION: The study findings suggest that lower anti-AT Ab responses before and during SAB, reflective of immune dysfunction, are associated with more severe clinical presentations of infection.

Clinical Circulating Tumor DNA Testing for Precision Oncology.

Circulating tumor DNA (ctDNA) is the portion of the cell-free DNA in the blood of cancer patients released from tumor cells via apoptosis, necrosis, or active release. From 10 mL of blood, the 4-5 mL of plasma obtained from a cancer patient contains 5-10 ng/mL of ctDNA. The plasma contains not only ctDNA of tumor origin, but also DNA from normal cells or clonal hematopoiesis. Another characteristic of ctDNA is its rapid clearance from circulation; it has a half-life of 16 minutes to 2.5 hours. Obtaining reliable results from ctDNA requires the application and approval of standardized clinical validation guidelines; however, the status of numerous ctDNA tests currently varies. The clinical use of ctDNA testing should be carefully considered based on the test's specific needs and characteristics. Here we provide the different characteristics of ctDNA tests and information regarding their validation and approval status.

Concordance of circulating tumor DNA and matched formalin-fixed paraffin-embedded tumor tissue in gastric cancer as a predictor of recurrence.

PURPOSE: Combined analysis of the variant composition of circulating tumor DNA (ctDNA) from cell-free plasma and DNA from tumor tissue could provide insight into the implications of the genetic alterations responsible for the intratumoral and intertumoral heterogeneity of gastric cancer. We aimed to evaluate the usefulness of this approach in these patients. METHODS: Cell-free plasma and formalin-fixed paraffin-embedded tumor tissue samples from 46 patients with gastric cancer were examined. Targeted deep sequencing was performed using a commercially available kit. RESULTS: The cell-free DNA (cfDNA) concentration was higher in stage II-IV versus stage I patients and in larger versus smaller tumors. Only 12 of the 36 (33.3%) alterations in the tumor tissue samples were in concordance with those in the ctDNA samples. Two variants were in concordance in stage I samples and 10 in stage II-IV samples. Actionable variants that were detected in concordance were in the stage II-IV samples. Preoperative ctDNA positivity of actionable variants was significantly associated with cfDNA concentration, lymphatic invasion, N stage, and TNM stage. Cancer recurrence was significantly associated with tumor size, lymphatic/vascular invasion, TNM stage, and ctDNA-tumor tissue variant concordance. CONCLUSION: Preoperative ctDNA genetic analysis using a multigene panel offers substantial clinical benefits when performed in conjunction with targeted deep sequencing of tumor tissue. Concordance between preoperative ctDNA and tumor tissue mutations may serve as a prognostic indicator in patients with gastric cancer.

Tissue miR-200c-3p and circulating miR-1290 as potential prognostic biomarkers for colorectal cancer.

Epithelial-mesenchymal transition (EMT)-related cancers generally elicit low immune responses. EMT is regulated by several microRNAs (miRNAs) in cancers. Thus, this study aimed to evaluate the prognostic potential of EMT-related miRNAs as biomarkers in colorectal cancer (CRC). Formalin-fixed paraffin-embedded tumor and normal tissue and plasma samples were obtained from 65 patients with pathologically confirmed CRC. In addition, plasma samples were obtained from 30 healthy volunteers. Immunohistochemical staining for E-cadherin, ZEB1, PD-1, PD-L1, CD3, CD4, CD8, Foxp3, and CD68 was conducted on tissue samples. Droplet digital polymerase chain reaction (ddPCR) analysis was performed to evaluate miR-21-5p, 34a-5p, 138-5p, 200a-3p, 200b-5p, 200c-3p, 630, 1246, and 1290 expression in tissue samples and miR-630, 1246, and 1290 expression in plasma samples. miR-21-5p, 34a-5p, 630, 1246, and 1290 expression was higher in tumor tissues than in normal tissues (P < 0.05). EMT was significantly associated with reduced tumor-infiltrating T cells. Moreover, miR-21-5p, miR-34a-5p, miR-200a-3p, and miR-200c-3p expression was negatively correlated with T cell density (P < 0.05). High tissue levels of miR-200c-3p were associated with poor overall survival (OS) (P < 0.001). CRC patients with the EMT phenotype had poor OS; however, PD-L1 positivity and abundant PD-1 positive immune cells were correlated with better OS (P < 0.05). miR-1246 and miR-1290 levels were significantly higher in the plasma of patients with CRC than in the plasma of healthy controls (P < 0.05). High plasma levels of miR-1290 were correlated with advanced stage and poor OS (P < 0.05). The tissue expression of miR-200c-3p and plasma levels of miR-1290 measured by ddPCR indicate their potential as prognostic biomarkers for CRC.

Association of HLA-DRB1 and -DQB1 Alleles with Susceptibility to IgA Nephropathy in Korean Patients.

BACKGROUND: Associations between IgA nephropathy (IgAN) and HLA-DRB1 and -DQB1 alleles have been reported in several ethnic groups. We investigated the association of HLA-DRB1 and -DQB1 alleles with the predisposition for IgAN and disease progression to end-stage kidney disease (ESKD) in Korean patients. METHODS: We analyzed HLA-DRB1 and -DQB1 genotypes in 399 IgAN patients between January 2000 and January 2019 using a LIFECODES sequence-specific oligonucleotide (SSO) typing kit (Immucor, Stamford, CT, USA) or a LABType SSO Typing Test (One Lambda, Canoga Park, CA, USA). Alleles with a significant difference in two-digit resolution were further analyzed using in-house sequence-based typing and sequence-specific primer PCR. As controls, 613 healthy hematopoietic stem cell donors were included. Kidney survival was analyzed in 281 IgAN patients with available clinical and laboratory data using Cox regression analysis. Where needed, P-values were adjusted using Bonferroni correction. RESULTS: The allele frequencies of HLA-DRB1*04:05 (corrected P [Pc]<0.001), -DQB1 *04:01 (Pc=0.048), and -DQB1*03:02 (Pc=0.021) were significantly higher in IgAN patients than in controls, whereas those of HLA-DRB1*07:01, -DRB1*15:01, -DQB1*02:02, and -DQB1*06:02 (Pc<0.001 for all) were significantly lower in IgAN patients than in controls. The allele frequency of HLA-DQB1*05:03 (Pc=0.016) was significantly lower in the ESKD group than in the non-ESKD group; however, there was no significant difference for ESKD progression between these groups. CONCLUSIONS: We report novel associations of HLA-DRB1*15:01, DQB1*02:02, -DQB1*03:02, and -DQB1*04:01 with IgAN. Further studies of HLA alleles associated with IgAN progression in a larger cohort and in various ethnic groups are needed.

Relationship among genetic polymorphism of SLCO1B1, rifampicin exposure and clinical outcomes in patients with active pulmonary tuberculosis.

AIMS: Rifampicin is a key drug for the treatment of tuberculosis (TB). Little is known for the relationship between the rifampicin pharmacokinetics and genetic polymorphisms in the Asian population. We aimed to investigate relationship between genetic polymorphism of SLCO1B1 and rifampicin exposure and its impact on clinical outcomes in Korean patients with active pulmonary TB. METHODS: From February 2016 to December 2019, patients with active pulmonary TB who were taking rifampicin for >1 week were prospectively enrolled. Serial or 1-time blood sampling was conducted to determine rifampicin concentrations. The genotype of 4 single nucleotide polymorphisms of SLCO1B1 was determined. To estimate the drug clearance and exposure, population pharmacokinetics analysis was conducted. Clinical outcomes such as time to acid-fast bacteria culture conversion, chest radiograph score changes from baseline, and all-cause mortality were also evaluated. The exposure among different SLCO1B1 genotype was compared and relationship between drug exposure and clinical outcomes were explored. RESULTS: A total of 105 patients (70 males and 35 females) were included in the final analysis. The mean age of patients was 55.4 years. The mean drug clearance and exposure were 13.6 L/h and 57.9 mg h/L, respectively. The genetic polymorphisms of SLCO1B1 were not related to rifampicin clearance or exposure. As the rifampicin exposure increased, the chest radiographs improved significantly, but the duration of acid-fast bacteria culture conversion was not related to the drug exposure. CONCLUSION: SLCO1B1 gene polymorphisms did not influence rifampicin concentrations and clinical outcomes in Korean patients with active pulmonary TB.

Comprehensive genetic features of gastric mixed adenoneuroendocrine carcinomas and pure neuroendocrine carcinomas.

We aimed to determine the pathogenesis of gastric mixed adenoneuroendocrine carcinoma (MANEC) and pure neuroendocrine carcinoma (NEC), which is largely unknown. Targeted DNA sequencing was performed on 34 tumor samples from 21 patients - 13 adenocarcinoma (ADC)/NEC components from MANECs and eight pure NECs - and 21 matched non-neoplastic gastric tissues. Mutational profiles of MANECs/NECs were compared with those of other tumors using public databases. The majority (64.1%; 59/92) of mutations in MANEC were shared by both ADC and NEC components. TP53 was the most commonly mutated gene in MANEC (69.2%, 9/13) and pure NEC (87.5%, 8/9). All TP53 mutations in MANEC were pathogenic mutations and were shared by both ADC and NEC components. A subset of TP53WT MANECs had a microsatellite-unstable phenotype or amplifications in various oncogenes including ERBB2 and NMYC, and the only TP53WT pure NEC harbored MYC amplification. Compared to NEC in other organs, NECs arising from the stomach had unique features including less frequent RB1 mutations. Differentially altered genes of MANEC ADC components were significantly associated with receptor tyrosine kinase signaling pathways, while differentially altered genes of MANEC NEC components were significantly associated with the NOTCH signaling pathway. Our data provide evidence suggesting a possible clonal origin of ADC and NEC components of MANEC, and we found that gastric MANECs and pure NECs are distinct entities with unique mutational profiles and underlying protein networks. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Toll-like receptor 2 downregulation and cytokine dysregulation predict mortality in patients with Staphylococcus aureus bacteremia.

BACKGROUND: Staphylococcus aureus bacteremia (SAB) presents heterogeneously, owing to the differences in underlying host conditions and immune responses. Although Toll-like receptor 2 (TLR2) is important in recognizing S. aureus, its function during S. aureus infection remains controversial. We aimed to examine the association of TLR2 expression and associated cytokine responses with clinical SAB outcomes. METHODS: Patients from a prospective SAB cohort at two tertiary-care medical centers were enrolled. Blood was sampled at several timepoints (≤5 d, 6-9 d, 10-13 d, 14-19 d, and ≥ 20 d) after SAB onset. TLR2 mRNA levels were determined via real-time PCR and serum tumor necrosis factor [TNF]-α, interleukin [IL]-6, and IL-10 levels were analyzed with multiplex-high-sensitivity electrochemiluminescent ELISA. RESULTS: TLR2 levels varied among 59 SAB patients. On days 2-5, TLR2 levels were significantly higher in SAB survivors than in healthy controls (p = 0.040) and slightly but not significantly higher than non-survivors (p = 0.120), and SAB patients dying within 7 d had lower TLR2 levels than survivors (P = 0.077) although statistically insignificant. IL-6 and IL-10 levels were significantly higher in non-survivors than in survivors on days 2-5 post-bacteremia (P = 0.010 and P = 0.021, respectively), and those dying within 7 d of SAB (n = 3) displayed significantly higher IL-10/TNF-α ratios than the survivors did (P = 0.007). CONCLUSION: TLR2 downregulation and IL-6 and IL-10 concentrations suggestive of immune dysregulation during early bacteremia may be associated with mortality from SAB. TLR2 expression levels and associated cytokine reactions during early-phase SAB may be potential prognostic factors in SAB, although larger studies are warranted.

Is elevated microsatellite alterations at selected tetranucleotide repeats (EMAST)-negative/MSI-high colorectal cancer a distinct subtype of the disease?

BACKGROUND AND OBJECTIVES: Microsatellite instability (MSI) plays a prognostic and predictive role in colorectal cancer (CRC). Elevated microsatellite alterations at selected tetranucleotide repeats (EMAST), a novel type of MSI, was recently identified. METHODS: A retrospective analysis of a prospective cohort database was performed. Patients who attempted curative surgery for MSI-high (MSI-H) CRC and had available testing results of EMAST were included for analysis. The difference in clinical characteristics, immunohistochemistry profile, and 3-year recurrence-free and overall survival between EMAST-negative and EMAST-positive tumors was measured. RESULTS: EMAST status was successfully evaluated in 86 cases among patients who received EMAST testing, and only 16.3% (14/86) of these patients were EMAST-negative/MSI-H. Patients with EMAST-negative tumors were younger; their tumors exhibited well differentiation, less venous invasion, and greater mutS homolog 3 expression. There was no distant metastasis or cancer-specific death among EMAST-negative patients. Yet no statistically significant difference was found between the two groups in 3-year overall or recurrence-free survival. CONCLUSIONS: Patients with EMAST-negative/MSI-H CRC seem to have different clinicopathological characteristics. Future large-scale studies could clarify the role of EMAST genotype as a sub-classifier of MSI-H CRC.

Author Correction: Comparative analysis of HER2 copy number between plasma and tissue samples in gastric cancer using droplet digital PCR.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Differential prognostic impact of CD8+ T cells based on human leucocyte antigen I and PD-L1 expression in microsatellite-unstable gastric cancer.

BACKGROUND: The aim of the study was to determine the human leucocyte antigen class-I (HLA-I), programmed death-ligand 1 (PD-L1) expression and tumour-infiltrating lymphocytes (TILs) of microsatellite instability-high gastric cancer. METHODS: The HLA-I expression type was determined by immunohistochemistry of HLA-A, HLA-B, HLA-C and ß2-microglobulin in the centre of the tumour (CT) and in the invasive margin (IM) of samples from 293 patients (total loss vs. preserved type). PD-L1 expression and TIL density was examined immunohistochemically. HLA-I genotyping was also performed. RESULTS: The expression loss of the HLA-I molecules was significantly associated with low TIL density. According to survival analyses, the HLA-I expression type and PD-L1 positivity were not independent prognostic factors. The TIL density had no prognostic implication when survival analysis was performed for the whole patient group; however, high CD8+ TIL infiltration was significantly associated with good prognosis in only HLA-I-preserved-type/PD-L1-positive group (p = 0.034). The homozygosity of the HLA-I allele was more frequently observed in the total loss type group. CONCLUSIONS: We confirmed differential prognostic implication of CD8+ TILs according to the HLA-I and PD-L1 expression. Determination of the HLA-I expression could be helpful to select patients who would benefit from anti-PD-1/PD-L1 therapy.

Comparative analysis of HER2 copy number between plasma and tissue samples in gastric cancer using droplet digital PCR.

In this study, we measured the human epidermal growth factor receptor 2 (HER2) copy number in both tissue and plasma samples of gastric cancer patients by using droplet digital polymerase chain reaction (ddPCR) method. Eighty gastric cancer patients were enrolled and both formalin-fixed and paraffin-embedded tissue and preoperative plasma samples were collected. HER2 status was determined by HER2 immunohistochemistry (IHC)/silver in situ hybridization (SISH) in tissue samples and ddPCR of the target gene HER2 and the reference gene eukaryotic translation initiation factor 2C, 1 in both tissue and plasma. The concordance rate of tissue HER2 status determined by IHC/SISH and HER2 ddPCR was 90.0% (72/80), and the sensitivity and specificity of tissue ddPCR were 85.0% and 95.0%, respectively. The concordance rate of plasma ddPCR and IHC/SISH was 63.8% (51/80). The sensitivity, specificity, positive predictive value, and negative predictive value of plasma HER2 ddPCR were 37.5%, 90.0%, 79.0%, and 59.0%, respectively. As HER2 measurement by tissue ddPCR showed a high concordance rate with HER2 status by IHC/SISH, it could replace tissue IHC/SISH testing in gastric cancer. These findings may contribute to the development of tissue and plasma HER2 testing that would be useful in daily practice.

Comparison of ABO isoagglutinin titres by three different methods: tube haemagglutination, micro-column agglutination and automated immunohematology analyzer based on erythrocyte-magnetized technology.

BACKGROUND AND OBJECTIVES: ABO isoagglutinin titre is important for evaluating and monitoring ABO-incompatible (ABOi) stem cells or solid organ transplantations. There are several methods to measure the titre level, including the tube haemagglutination method, micro-column agglutination and erythrocyte-magnetized technology (EMT). However, few studies have reported isoagglutinin measured by EMT. Here, we compared the isoagglutinin titre of normal individuals obtained by an automated instrument with that obtained by conventional manual methods to evaluate the feasibility of replacing the manual method with the automated instrument. MATERIALS AND METHODS: The ABO isoagglutinin titre was measured on residual samples of healthy individuals who visited the health promotion centre of the National Cancer Center, Korea, from April to October 2015. Samples from 120 patients were collected, which included 20 males and 20 females for each blood group (A, B and O). IgM and IgG ABO isoagglutinin titres of each blood group were measured by the tube haemagglutination, micro-column agglutination and EMT techniques. The median (minimum-maximum) titres were compared, and the concordance between two methods was evaluated with the rate of results showing within one titre difference. RESULTS: The median ABO IgM and IgG titres of all blood groups obtained by the EMT method were higher than that obtained by the conventional tube haemagglutination and micro-column agglutination. CONCLUSION: The agreement between the two methods was comparable in case of IgM but low in IgG.

Cluster of Lymphadenitis due to Nontuberculous Mycobacterium in Children and Adolescents 8-15 Years of Age.

BACKGROUND: Nontuberculous mycobacteria (NTM) lymphadenitis is an under-recognized entity, and data of the true burden in children are limited. Without a high index of suspicion, diagnosis may be delayed and microbiological detection is challenging. Here, we report a cluster of NTM lymphadenitis experienced in Korean children. METHODS: Subjects under 19 years of age diagnosed with NTM lymphadenitis during November 2016-April 2017 and April 2018 were included. Electronic medical records were reviewed for clinical, laboratory and pathological findings. Information regarding underlying health conditions and environmental exposure factors was obtained through interview and questionnaires. RESULTS: A total of ten subjects were diagnosed during 18 months. All subjects were 8-15 years of age, previously healthy, male and had unilateral, nontender, cervicofacial lymphadenitis for more than 3 weeks with no significant systemic symptoms and no response to empirical antibiotics. Lymph nodes involved were submandibular (n = 8), preauricular (n = 6) and submental (n = 1). Five patients had two infected nodes and violaceous discoloration was seen in seven subjects. Biopsy specimens revealed chronic granulomatous inflammation and acid-fast bacteria culture identified Mycobacterium haemophilum in two cases and NTM polymerase chain reaction was positive in two cases. Survey revealed various common exposure sources. CONCLUSION: NTM lymphadenitis is rare but increasing in detection and it may occur in children and adolescents. Diagnosis requires high index of suspicion and communication between clinicians and the laboratory is essential for identification of NTM.

Digital polymerase chain reaction for detecting c-MYC copy number gain in tissue and cell-free plasma samples of colorectal cancer patients.

We focused on the utility of the droplet digital polymerase chain reaction (ddPCR) for detecting c-MYC gene copy number (GCN) gain in cell-free plasma and tumor tissue of colorectal cancer (CRC) patients. c-MYC GCN status was determined using dual-color silver in situ hybridization (SISH) and ddPCR in retrospective cohort 1 (192 CRC patients) and prospective cohort 2 (64 CRC patients). In cohort 1, c-MYC GCN gain was observed in 34 (17.5%) patients by SISH, and in 7 (3.6%) patients by ddPCR. c-MYC GCN by SISH significantly correlated with ddPCR results (ρ = 0.532, P < 0.001). Although 40 cases (20.7%) showed intratumoral genetic heterogeneity, it did not cause discordance in results obtained by the two methods. c-MYC GCN gain, by both SISH and ddPCR was independently correlated with worst prognosis (P = 0.002). In cohort 2, c-MYC GCN estimation in tissue by ddPCR was also significantly associated with results obtained by SISH (ρ = 0.349, P = 0.005), but correlated with plasma ddPCR with borderline significance (ρ = 0.246, P = 0.050). Additionally, detecting c-MYC GCN gain in plasma with ddPCR might have relatively low sensitivity but high specificity. Our study suggests that ddPCR can be a useful tool for detecting c-MYC GCN gain as a potential prognostic biomarker in CRC tissue samples; however, this will need further verification in plasma samples.

Effect of combining a recruitment maneuver with protective ventilation on inflammatory responses in video-assisted thoracoscopic lobectomy: a randomized controlled trial.

BACKGROUND: We hypothesized that the addition of a recruitment maneuver to protective ventilation (PVRM) would result in lower pulmonary and systemic inflammatory responses than traditional ventilation or protective ventilation (PV) alone in patients undergoing lung surgery. METHODS: Sixty patients who underwent scheduled thoracoscopic lobectomy were randomly assigned to three groups: traditional ventilation, PV, or PVRM. Ventilations were performed using a tidal volume of 10 mL/kg for the traditional ventilation group and either 8 mL/kg (two-lung) or 6 mL/kg (one-lung, OLV) with a positive end-expiratory pressure of 5 cm H2O for the PV and PVRM groups. The RM was performed 10 min after the start of OLV. Fiberoptic bronchoalveolar lavage (BAL) was performed twice in dependent and non-dependent lungs: before the start and immediately after the end of OLV. Blood samples were collected at the same time points. The levels of cytokines, including TNF-α, IL-1ß, IL-6, IL-8, and IL-10, were measured. RESULTS: After OLV, the level of TNF-α in the BAL fluid of dependent lungs was significantly higher in the PV than in the PVRM group (P = 0.049), whereas IL-1ß, IL-6, IL-8, and IL-10 levels were not significantly different among the groups. In non-dependent lung BAL fluid, no cytokines were significantly different among the groups. After OLV, IL-10 serum levels were significantly higher in the traditional ventilation than in the PVRM group (P = 0.027). CONCLUSIONS: Lower inflammatory responses in the ventilated lung and serum were observed with PVRM than with traditional ventilation or PV alone. Larger multi-center clinical trials are warranted to confirm the effects of different ventilatory strategies on postoperative outcomes.