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Abstract Metastatic gastric cancer (GC) presents significant clinical challenges due to its poor prognosis and limited treatment options. To address this, we conducted a targeted protein biomarker discovery study to identify markers predictive of metastasis in advanced GC (AGC). Serum samples from 176 AGC patients (T stage 3 or higher) were analyzed using the Olink Proteomics Target panels. Patients were retrospectively categorized into nonmetastatic, metastatic, and recurrence groups, and differential protein expression was assessed. Machine learning and gene set enrichment analysis (GSEA) methods were applied to discover biomarkers and predict prognosis. Four proteins (MUC16, CAIX, 5’-NT, and CD8A) were significantly elevated in metastatic GC patients compared to the control group. Additionally, GSEA indicated that the response to interleukin-4 and hypoxia-related pathways were enriched in metastatic patients. Random forest classification and decision-tree modeling showed that MUC16 could be a predictive marker for metastasis in GC patients. Additionally, ELISA validation confirmed elevated MUC16 levels in metastatic patients. Notably, high MUC16 levels were independently associated with metastatic progression in T3 or higher GC. These findings suggest the potential of MUC16 as a clinically relevant biomarker for identifying GC patients at high risk of metastasis.

Abstract Acinetobacter baumannii (AB) has emerged as a major pathogen in vulnerable and severely ill patients. It remains unclear whether early mortality (EM) due to AB bacteremia is because of worse clinical characteristics of the infected patients or the virulence of the pathogen. In this study, we aimed to investigate the effect of AB virulence on EM due to bacteremia. This retrospective study included 138 patients with AB bacteremia (age: ≥ 18 years) who were admitted to a tertiary care teaching hospital in South Korea between 2015 and 2019. EM was defined as death occurring within 7 days of bacteremia onset. The AB clinical isolates obtained from the patients’ blood cultures were injected into 15 Galleria mellonella larvae each, which were incubated for 5 days. Clinical isolates were classified into high- and low-virulence groups based on the number of dead larvae. Patients’ clinical data were combined and subjected to multivariate Cox regression analyses to identify the risk factors for EM. In total, 48/138 (34.8%) patients died within 7 days of bacteremia onset. The Pitt bacteremia score was the only risk factor associated with EM. In conclusion, AB virulence had no independent effect on EM in patients with AB bacteremia.

IntroductionThe rapid spread of COVID-19 worldwide within 2 months demonstrated the vulnerability of the world’s population to infectious diseases. In 2015, the Global Antimicrobial Resistance and Use Surveillance System (GLASS) was launched to combat antimicrobial resistance (AMR). However, there has been no comprehensive assessment of the decade-long global battle against AMR based on GLASS data.MethodsSouth Korea established Kor-GLASS (Korean-GLASS) to proactively monitor data quality and enable international collaborations. A unique feature of Kor-GLASS is the quality control center (QCC), which uses network hubs and ensures standardized, high-quality data through interlaboratory proficiency testing (IPT) and external quality assessment (EQA). In addition, the QCC multifaceted endeavors for integrated data quality management.ResultsSince 2020, high-quality AMR data have indicated fluctuating antibiotic resistance rates in South Korea. This trend does not align with the decrease in antibiotic usage seen in humans but coincides with non-human antibiotic sales, indicating a need for greater monitoring of non-human antibiotic resistance. Comprehensive and robust management taking account of the intricate interplay among humans, animals, and the environment is essential. Kor-GLASS has been expanded into a “One Health” multiagency collaborative initiative.DiscussionAlthough a standardized solution is not suitable for all countries, it must align with the local context and international standards. A centralized top-down management structure such as that of the QCC is essential to ensure continuous data quality coordination. Sustained efforts and surveillance systems are crucial for monitoring and managing AMR and safeguarding human health.

Abstract Galactomannan (GM) is a polysaccharide cell wall component released by Aspergillus spp., and an immunoenzymatic GM assay is used for the diagnosis of invasive pulmonary aspergillosis. We evaluated the cause of strong positivity for GM in patients with no typical signs of aspergillosis. Repeat assays were performed using different instruments and reagent lots, but there were no differences in results among the assays. Patients with strongly positive GM results were investigated. Medication histories revealed that 14 of 23 patients had been administered total parenteral nutrition solution from one manufacturer and 4 patients had been administered dextrose solution from a different manufacturer before being tested. The results of GM assays conducted on samples of dextrose solution and the glucose fraction of the total parenteral nutrition solution were strongly positive, confirming the causes of the false-positive reactions. We hypothesize that a trace amount of GM was introduced into the glucose-containing solutions because glucoamylase, which is necessary for the saccharification step of glucose synthesis, was derived from Aspergillus niger. To enhance patient care and prevent unnecessary antifungal prescriptions, healthcare providers and manufacturers of healthcare products need to be aware of the possibility of false-positive reactions for GM.

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.

Abstract 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.

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.

ABSTRACTBacterial contamination of blood products poses a significant risk in transfusion medicine. Platelets are particularly vulnerable to bacterial growth because they must be stored at room temperature with constant agitation for >5 days. The limitations of bacterial detection using conventional methods, such as blood cultures and lateral flow assays, include the long detection times, low sensitivity, and the requirement for substantial volumes of blood components. To address these limitations, we assessed the performance of a bacterial enrichment technique using antibiotic-conjugated magnetic nanobeads (AcMNBs) and real-time PCR for the detection of bacterial contamination in plasma. AcMNBs successfully captured >80% of four bacterial strains, including Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Klebsiella pneumoniae, in both plasma and phosphate-buffered saline. After 24-h incubation with bacterial enrichment, S. aureus and B. cereus were each detected at 101 CFU/mL in all trials (5/5), E. coli at 101 CFU/mL in 1/5 trials, and K. pneumoniae at 10² CFU/mL in 4/5 trials. Additionally, without incubation, the improvement was also achieved in samples with bacterial enrichment, S. aureus at 10² CFU/mL and B. cereus at 101 CFU/mL in 1/5 trials each, E. coli at 10³ CFU/mL in 3/5 trials, and K. pneumoniae at 10¹ CFU/mL in 2/5 trials. Overall, the findings from this study strongly support the superiority of bacterial enrichment in detecting low-level bacterial contamination in plasma when employing AcMNBs and PCR.IMPORTANCEThe study presents a breakthrough approach to detect bacterial contamination in plasma, a critical concern in transfusion medicine. Traditional methods, such as blood cultures and lateral flow assays, are hampered by slow detection times, low sensitivity, and the need for large blood sample volumes. Our research introduces a novel technique using antibiotic-conjugated magnetic nanobeads combined with real-time PCR, enhancing the detection of bacteria in blood products, especially platelets. This method has shown exceptional efficiency in identifying even low levels of four different species of bacteria in plasma. The ability to detect bacterial contamination rapidly and accurately is vital for ensuring the safety of blood transfusions and can significantly reduce the risk of infections transmitted through blood products. This advancement is a pivotal step in improving patient outcomes and elevating the standards of care in transfusion medicine.

Abstract The exploration of oral microbiome has been increasing due to its relatedness with various systemic diseases, but standardization of saliva sampling for microbiome analysis has not been established, contributing to the lack of data comparability. Here, we evaluated the factors that influence the microbiome data. Saliva samples were collected by the two collection methods (passive drooling and mouthwash) using three saliva-preservation methods (OMNIgene, DNA/RNA shield, and simple collection). A total of 18 samples were sequenced by both Illumina short-read and Nanopore long-read next-generation sequencing (NGS). The component of the oral microbiome in each sample was compared with alpha and beta diversity and the taxonomic abundances, to find out the effects of factors on oral microbiome data. The alpha diversity indices of the mouthwash sample were significantly higher than that of the drooling group with both short-read and long-read NGS, while no significant differences in microbial diversities were found between the three saliva-preservation methods. Our study shows mouthwash and simple collection are not inferior to other sample collection and saliva-preservation methods, respectively. This result is promising since the convenience and cost-effectiveness of mouthwash and simple collection can simplify the saliva sample preparation, which would greatly help clinical operators and lab workers.

ABSTRACTObjectives The oral microbiome is closely associated with systemic diseases, indicating the presence of bacteremia and inflammatory mediators in the systemic circulation. Our research aims to investigate the relationship between the oral microbiome and other microbial habitats.Methods We analyzed 180 specimens from 36 patients, including saliva, buccal swab, plaque, stool, and blood samples from a healthy group (Non_PD, n = 18) and a periodontitis group (PD, n = 18). The final analysis included 147 specimens, with varying sample sizes for each group. Metagenomic analysis was performed using prokaryotic 16S rRNA on the MiSeq platform (Illumina).Results PD saliva showed significant richness differences (P's

IntroductionRecent studies have proposed several plausible mechanisms supporting the association between periodontal disease and systemic disease. However, characterizing the microbial communities in individuals with periodontal disease before onset of other diseases is an important first step in determining how the altered microbial state contributes to disease progression. This study established microbiome profiles for five body habitats of carefully selected, otherwise healthy individuals with periodontal disease.MethodsBlood, oral (buccal mucosa, dental plaque, and saliva), and stool samples were collected from ten healthy subjects with periodontal disease. Using 16S rRNA metagenomics, the taxonomic and functional compositions of microbiomes were investigated.ResultsThe most predominant phylum in blood and stool was Bacillota. Pseudomonadota accounted for the largest proportion of microbes in the buccal mucosa and saliva, whereas Bacteroidota were the most prevalent in dental plaque. Differential abundance analysis revealed that 12 phyla and 139 genera were differentially abundant between body habitats. Comparison of alpha diversity showed that the blood microbiome has the most diverse community close to neither oral nor stool microbiomes. We also predicted the functional configurations of the microbiome in otherwise healthy subjects with periodontal disease. Principal coordinate analysis based on functional abundance revealed distinct clustering of the microbial communities between different body habitats, as also observed for taxonomic abundance. In addition, 13 functional pathways, including lipopolysaccharide biosynthesis, glutathione metabolism, and proteasome, showed differential expression between habitats.DiscussionOur results offer insight into the effects of the microbiome on systemic health and disease in people with periodontal disease.

Abstract Bacterial contamination of blood products is a major problem in transfusion medicine, in terms of both morbidity and mortality. Platelets (PLTs) are stored at room temperature (under constant agitation) for more than 5 days, and bacteria can thus grow significantly from a low level to high titers. However, conventional methods like blood culture and lateral flow assay have disadvantages such as long detection time, low sensitivity, and the need for a large volume of blood components. We used real-time polymerase chain reaction (PCR) assays with antibiotic-conjugated magnetic nanobeads (MNBs) to detect enriched Gram-positive and -negative bacteria. The MNBs were coated with polyethylene glycol (PEG) to prevent aggregation by blood components. Over 80% of all bacteria were captured by the MNBs, and the levels of detection were 101 colony forming unit [CFU]/mL and 102 CFU/mL for Gram-positive and -negative bacteria, respectively. The detection time is

Abstract Background Given the increasing incidence of Clostridioides difficile infections in Korea, there has been an increase in inappropriate testing for C. difficile, which has rendered overdiagnosis of asymptomatic colonisers common. We aimed to investigate the appropriateness of C. difficile testing and the related factors. Methods We retrospectively reviewed the medical records of patients who were admitted to a 1300-bed tertiary-care teaching hospital in Korea and were tested for C. difficile infection from September 2019 to November 2019. We performed logistic regression analysis to investigate factors related to inappropriate testing. Further, a survey was conducted on physicians to assess the knowledge and ordering patterns of C. difficile testing. Results We included 715 tests from 520 patients in the analysis. Testing was classified as hospital-onset and community-onset and subclassified as appropriate and inappropriate following an algorithmic method. Among the 715 tests, 576 (80.6%) and 139 (19.6%) tests were classified as hospital-onset and community-onset, respectively. Among the hospital-onset tests, 297 (52%) were considered inappropriate. The risk of inappropriate testing increased when C. difficile tests were conducted in the emergency room (OR 24.96; 95% CI 3.12–199.98) but decreased in intensive care units (OR 0.36, 95% CI 0.19–0.67). The survey was conducted on 61 physicians. Internal medicine physicians had significantly higher scores than non-internal medicine physicians (7.1 vs. 5.7, p = 0.001). The most frequently ordered combination of tests was toxin + glutamate dehydrogenase (47.5%), which was consistent with the ordered tests. Conclusion Almost half of the C. difficile tests were performed inappropriately. The patient being located in the emergency room and intensive care unit increased and decreased the risk of inappropriate testing, respectively. In a questionnaire survey, we showed that internal medicine physicians were more knowledgeable about C. difficile testing than non-internal medicine physicians. There is a need to implement the diagnostic stewardship for C. difficile, especially through educational interventions for emergency room and non-internal medicine physicians.

Abstract 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

ABSTRACT To optimize antibiotic use, the US CDC has outlined core elements of antimicrobial stewardship programs (ASP). However, they are difficult to implement in limited-resource settings. We report on the successful implementation of a series of ASP with insufficient number of infectious diseases specialists. We retrospectively collected data regarding antibiotic administration and culture results of all patients admitted to a tertiary care teaching hospital, Seoul National University Bundang Hospital (SNUBH), from January 2010 to December 2019. Trends of antibiotic use and antibiotic resistance rates were compared with those from Korean national data. Trend analyses were performed using nonparametric, two-sided, correlated seasonal Mann–Kendall tests. Total antibiotic agent usage has significantly decreased with ASP implementation at SNUBH since 2010. National claim data from tertiary care hospitals have revealed an increase in the use of all broad-spectrum antibiotics except for third-generation cephalosporins (3GC). In contrast, at SNUBH, glycopeptide and fluoroquinolone use gradually decreased, and 3GC and carbapenem use did not significantly change. Furthermore, the rate of colonization with methicillin-resistant Staphylococcus aureus showed a consistently decreasing trend, while that with 3GC- and fluoroquinolone-resistant Escherichia coli significantly increased. Unlike the national rate, the rate of colonization with antibiotic resistant-Klebsiella pneumoniae did not increase and that of 3GC- and fluoroquinolone-resistant Pseudomonas aeruginosa significantly decreased. Stepwise implementation of core ASP elements was effective in reducing antibiotic use despite a lack of sufficient manpower. Long-term multidisciplinary teamwork is necessary for successful and sustainable ASP implementation. IMPORTANCE Antimicrobial stewardship programs aimed to optimize antibiotic use are difficult to implement in limited-resource settings. Our study indicates that stepwise implementation of core antimicrobial stewardship program elements was effective in reducing antibiotic use in a tertiary care hospital despite the lack of sufficient manpower.

Recently, the number of patients with coronavirus disease 2019 (COVID-19) who have tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), via the reverse transcription polymerase chain reaction (RT-PCR) test, after recovery has increased; this has caused a dilemma regarding the medical measures and policies. We evaluated the dynamics of viral load and anti-SARS-CoV-2 antibodies in four patients with positive RT-PCR results after recovery. In all patients, the highest levels of immunoglobulin G (IgG) and IgM antibodies were reached after about a month of the onset of the initial symptoms. Then, the IgG titers plateaued, and the IgM titers decreased, regardless of RT-PCR results. The IgG and IgM levels did not increase after the post-negative positive RT-PCR results in any of the patients. Our results reinforced that the post-negative positive RT-PCR results may be due to the detection of RNA particles rather than reinfection in individuals who have recovered from COVID-19.

Abstract 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.

Vancomycin-resistant enterococci (VRE) are nosocomial pathogens with genetic plasticity and widespread antimicrobial resistance (AMR). To prevent the spread of VRE in the hospital setting, molecular epidemiological approaches such as pulsed-field gel electrophoresis and multilocus sequence typing have been implemented for pathogen outbreak surveillance. However, due to the insufficient discriminatory power of these methods, whole-genome sequencing (WGS), which enables high-resolution analysis of entire genomic sequences, is being used increasingly. Herein, we performed WGS of VRE using both short-read next-generation sequencing (SR-NGS) and long-read next-generation sequencing (LR-NGS). Since standardized workflows and pipelines for WGS-based bacterial epidemiology are lacking, we established three-step pipelines for SR- and LR-NGS, as a standardized WGS-based approach for strain typing and AMR profiling. For strain typing, we analyzed single-nucleotide polymorphisms (SNPs) of VRE isolates and constructed SNP-based maximum-likelihood phylogenies. The phylogenetic trees constructed using short and long reads showed good correspondence. Still, SR-NGS exhibited higher sensitivity for detecting nucleotide substitutions of bacterial sequences. During AMR profiling, we examined AMR genes and resistance-conferring mutations. We also assessed the concordance between genotypic and phenotypic resistance, which was generally better for LR-NGS than SR-NGS. Further validation of our pipelines based on outbreak cases is necessary to ensure the overall performance of pipelines.

Zika virus (ZIKV) emerged as a serious public health problem since the first major outbreak in 2007. Current ZIKV diagnostic methods can successfully identify known ZIKV but are impossible to track the origin of viruses and pathogens other than known ZIKV strains. We planned to determine the ability of Whole Genome Sequencing (WGS) in clinical epidemiology by evaluating whether it can successfully detect the origin of ZIKV in a suspected case of laboratory-acquired infection (LAI). ZIKV found in the patient sample was sequenced with nanopore sequencing technology, followed by the production of the phylogenetic tree, based on the alignment of 38 known ZIKV strains with the consensus sequence. The closest viral strain with the consensus sequence was the strain used in the laboratory, with a percent identity of 99.27%. We think WGS showed its time-effectiveness and ability to detect the difference between strains to the level of a single base. Additionally, to determine the global number of LAIs, a literature review of articles published in the last 10 years was performed, and 53 reports of 338 LAIs were found. The lack of a universal reporting system was worrisome, as in the majority of cases (81.1%), the exposure route was unknown.

Background/Aims Current evidence supports lung ultrasound as a point-of-care alternative diagnostic tool for various respiratory diseases. We sought to determine the utility of lung ultrasound for early detection of pneumonia and for assessment of respiratory failure among patients with coronavirus disease 2019 (COVID-19). Methods Six patients with confirmed COVID-19 by reverse transcription-polymerase chain reaction were enrolled. All had undergone chest X-ray and chest computed tomography (CT) on the day of admission and underwent multiple point-of-care lung ultrasound scans over the course of their hospitalization. Results Lung ultrasound detected early abnormal findings of representative B-lines in a patient with a normal chest X-ray, corresponding to ground-glass opacities on the chest CT scan. The ultrasound findings improved as her clinical condition improved and her viral load decreased. In another minimally symptomatic patient without significant chest X-ray findings, the ultrasound showed B-lines, an early sign of pneumonia before abnormalities were detected on the chest CT scan. In two critically ill patients, ultrasound was performed to assess for evaluation of disease severity. In both patients, the clinicians conducted emergency rapid sequence intubation based on the ultrasound findings without awaiting the laboratory results and radiological reports. In two children, ultrasound was used to assess the improvement in their pneumonia, thus avoiding further imaging tests such as chest CT. Conclusions Lung ultrasound is feasible and useful as a rapid, sensitive, and affordable point-of-care screening tool to detect pneumonia and assess the severity of respiratory failure in patients hospitalized with COVID-19.

To monitor national antimicrobial resistance (AMR), the Korea Global AMR Surveillance System (Kor-GLASS) was established. This study analyzed bloodstream infection (BSI) cases from Kor-GLASS phase I from January 2017 to December 2019. Nine non-duplicated Kor-GLASS target pathogens, including Staphylococcus aureus, Enterococcus faecalis, Enterococcus faecium, Streptococcus pneumoniae, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter spp., and Salmonella spp., were isolated from blood specimens from eight sentinel hospitals. Antimicrobial susceptibility testing, AMR genotyping, and strain typing were carried out. Among the 20,041 BSI cases, 15,171 cases were caused by one of the target pathogens, and 12,578 blood isolates were collected for the study. Half (1,059/2,134) of S. aureus isolates were resistant to cefoxitin, and 38.1% (333/873) of E. faecium isolates were resistant to vancomycin. Beta-lactamase-non-producing ampicillin-resistant and penicillin-resistant E. faecalis isolates by disk diffusion method were identified, but the isolates were confirmed as ampicillin-susceptible by broth microdilution method. Among E. coli, an increasing number of isolates carried the blaCTX–M–27 gene, and the ertapenem resistance in 1.4% (30/2,110) of K. pneumoniae isolates was mostly (23/30) conferred by K. pneumoniae carbapenemases. A quarter (108/488) of P. aeruginosa isolates were resistant to meropenem, and 30.5% (33/108) of those carried acquired carbapenemase genes. Over 90% (542/599) of A. baumannii isolates were imipenem-resistant, and all except one harbored the blaOXA–23 gene. Kor-GLASS provided comprehensive AMR surveillance data, and the defined molecular mechanisms of resistance helped us to better understand AMR epidemiology. Comparative analysis with other GLASS-enrolled countries is possible owing to the harmonized system provided by GLASS.

Aims This study established blood group analysis methods using whole-genome sequencing (WGS) data and conducted blood group analyses to determine the domestic allele frequency using public data from the Korean whole sequence analysis of the Korean Reference Genome Project conducted by the Korea Disease Control and Prevention Agency (KDCA). Materials and methods We analyzed the differences between the human reference sequences (hg19) and the conventional reference cDNA sequences of blood group genes using the Clustal Omega website, and established blood group analysis methods using WGS data for 41 genes, including 39 blood group genes involved in 36 blood group antigens, as well as the GATA1 and KLF1 genes, which are erythrocyte-specific transcription factor genes. Using CLC genomics Workbench 11.0 (Qiagen, Aarhus, Denmark), variant analysis was performed on these 41 genes in 250 Korean WGS data sets, and each blood group’s genotype was predicted. The frequencies for major alleles were also investigated and compared with data from the Korean rare blood program (KRBP) and the Erythrogene database (East Asian and all races). Results Among the 41 blood group-related genes, hg19 showed variants in the following genes compared to the conventional reference cDNA: GYPA, RHD, RHCE, FUT3, ACKR1, SLC14A1, ART4, CR1, and GCNT2. Among 250 WGS data sets from the Korean Reference Genome Project, 70.6 variants were analyzed in 205 samples; 45 data samples were excluded due to having no variants. In particular, the FUT3, GNCT2, B3GALNT1, CR1, and ACHE genes contained numerous variants, with averages of 21.1, 13.9, 13.4, 9.6, and 7.0, respectively. Except for some blood groups, such as ABO and Lewis, for which it was difficult to predict the alleles using only WGS data, most alleles were successfully predicted in most blood groups. A comparison of allele frequencies showed no significant differences compared to the KRBP data, but there were differences compared to the Erythrogene data for the Lutheran, Kell, Duffy, Yt, Scianna, Landsteiner-Wiener, and Cromer blood group systems. Numerous minor blood group systems that were not available in the KRBP data were also included in this study. Conclusions We successfully established and performed blood group analysis using Korean public WGS data. It is expected that blood group analysis using WGS data will be performed more frequently in the future and will contribute to domestic data on blood group allele frequency and eventually the supply of safe blood products.

We incorporated nationwide Candida antifungal surveillance into the Korea Global Antimicrobial Resistance Surveillance System (Kor-GLASS) for bacterial pathogens. We prospectively collected and analyzed complete non-duplicate blood isolates and information from nine sentinel hospitals during 2020–2021, based on GLASS early implementation protocol for the inclusion of Candida species. Candida species ranked fourth among 10,758 target blood pathogens and second among 4050 hospital-origin blood pathogens. Among 766 Candida blood isolates, 87.6% were of hospital origin, and 41.3% occurred in intensive care unit patients. Adults > 60 years of age accounted for 75.7% of cases. Based on species-specific clinical breakpoints, non-susceptibility to fluconazole, voriconazole, caspofungin, micafungin, and anidulafungin was found in 21.1% (154/729), 4.0% (24/596), 0.1% (1/741), 0.0% (0/741), and 0.1% (1/741) of the isolates, respectively. Fluconazole resistance was determined in 0% (0/348), 2.2% (3/135, 1 Erg11 mutant), 5.3% (7/133, 6 Pdr1 mutants), and 5.6% (6/108, 4 Erg11 and 1 Cdr1 mutants) of C. albicans, C. tropicalis, C. glabrata, and C. parapsilosis isolates, respectively. An echinocandin-resistant C. glabrata isolate harbored an F659Y mutation in Fks2p. The inclusion of Candida species in the Kor-GLASS system generated well-curated surveillance data and may encourage global Candida surveillance efforts using a harmonized GLASS system.

The microbiological characteristics of Staphylococcus aureus causing infective endocarditis (IE) have not been investigated thoroughly. We compared the characteristics of S. aureus isolates from patients with and without IE. Cases of S. aureus bacteremia (SAB) were collected from 10 hospitals over 7 years. Cases of native valve IE were matched with non-IE controls according to the following criteria: central-line-associated infection, community-acquired infection, methicillin susceptibility, and if possible, the primary site of infection. Genes coding virulence factors were analyzed using multiplex polymerase chain reactions. Fibrinogen and fibronectin-binding properties were assessed using in vitro binding assays. The fibronectin-binding protein A gene (fnbpA) was sequenced. Of 2,365 cases of SAB, 92 had IE. After matching, 37 pairs of S. aureus isolates from the IE cases and non-IE controls were compared; fnbpA was detected in 91.9% of the IE isolates and 100% of the non-IE isolates (p = 0.24). While the fibrinogen binding ratio was similar (1.07 ± 0.33 vs. 1.08 ± 0.26, p = 0.89), the fibronectin-binding ratio was significantly higher in the IE-group (1.31 ± 0.42 vs. 1.06 ± 0.31, p = 0.01). The proportions of major single-nucleotide polymorphisms in fnbpA were as follows: E652D (2.9% vs. 2.7%), H782Q (65.6% vs. 60.6%), and K786N (65.6% vs. 72.7%). The fibronectin-binding ratio was positively correlated with the number of SNPs present in IE cases (p < 0.001) but not in the non-IE controls (p = 0.124). Fibronectin-binding might play a key role in SAB IE. However, the degree of binding may be mediated by genetic variability between isolates.

Serological weak D is a reaction of 2+ or less to anti-D reagent and includes weak D and partial D phenotypes. Although identifying the RhD subtype is important for transfusion safety, serological tests are insufficient for defining the RhD subtype, and molecular tests are needed. To analyze the molecular characteristics of D variants in Koreans to facilitate the formulation of individualized transfusion strategies, molecular tests such as RhD genotyping using real-time polymerase chain reaction (PCR) and partial-D and/or weak-D sequence-specific amplification (SSP) were performed on 105 Korean Rare Blood Program (KRBP) patients exhibiting serological weak D. In total, 58 out of 68 serologically determined weak D KRBP patients were typed as having weak D or partial D phenotypes via RhD genotyping. In detail, eight (13.8%) were typed as partial DVa or DBS, nine (15.5%) as weak D type 15, and four others (6.8%) as partial DVI, partial DVII, weak D type 2, or weak D type 41 or 45, whereas the rest (n = 37, 63.8%) was typed as having either weak D or partial D. This suggests that serological weak D Koreans who require transfusion should be treated as D-negative.

BACKGROUND:The T-SPOT.TB can be read by an ELISPOT plate imager as an alternative to a labor-intensive and time-consuming manual reading, but its accuracy has not been sufficiently discussed to date. METHODS:1,423 test results obtained from manual reading using a microscope and an ELISPOT plate imager were compared. The agreement of qualitative test results was assessed using Cohen's kappa coefficient. The relationship of spot counts was studied using Bland-Altman analysis. RESULTS:The overall percent agreement of the qualitative test results was 95.43% with a kappa coefficient of 0.91. Positive test results with the maximum net spot count of 8 and borderline test results showed relatively high discordance. The agreement of spot counts in panel A, panel B, and nil control was good, and variability did not increase with higher spot counts. On the basis of study findings, a novel strategy for interpreting the test results by an ELISPOT plate imager was proposed. CONCLUSIONS:To increase diagnostic accuracy, positive test results with the maximum net spot count of 8 and borderline test results should be manually confirmed. Our strategy could be a practical guide for laboratories to build their own strategies for interpreting the test results by an ELISPOT plate imager.

Carbapenem-resistant Acinetobacter baumannii (CRAB) outbreaks in hospital settings challenge the treatment of patients and infection control. Understanding the relatedness of clinical isolates is important in distinguishing outbreak isolates from sporadic cases. This study investigated 11 CRAB isolates from a hospital outbreak by whole-genome sequencing (WGS), utilizing various bioinformatics tools for outbreak analysis. The results of multilocus sequence typing (MLST), single nucleotide polymorphism (SNP) analysis, and phylogenetic tree analysis by WGS through web-based tools were compared, and repetitive element polymerase chain reaction (rep-PCR) typing was performed. Through the WGS of 11 A. baumannii isolates, three clonal lineages were identified from the outbreak. The coexistence of blaOXA-23, blaOXA-66, blaADC-25, and armA with additional aminoglycoside-inactivating enzymes, predicted to confer multidrug resistance, was identified in all isolates. The MLST Oxford scheme identified three types (ST191, ST369, and ST451), and, through whole-genome MLST and whole-genome SNP analyses, different clones were found to exist within the MLST types. wgSNP showed the highest discriminatory power with the lowest similarities among the isolates. Using the various bioinformatics tools for WGS, CRAB outbreak analysis was applicable and identified three discrete clusters differentiating the separate epidemiologic relationships among the isolates.

Although viruses are known to be the second most common etiological factor in community-acquired pneumonia (CAP), the respiratory viral profile of the patients with healthcare-associated pneumonia (HCAP) has not yet been elucidated. We investigated the prevalence and the clinical impact of respiratory virus infection in adult patients with HCAP.Patients admitted with HCAP or CAP, between January and December 2016, to a tertiary referral hospital in Korea, were prospectively enrolled, and virus identification was performed using reverse-transcription polymerase chain reaction (RT-PCR).Among 452 enrolled patients (224 with HCAP, 228 with CAP), samples for respiratory viruses were collected from sputum or endotracheal aspirate in 430 (95.1%) patients and from nasopharyngeal specimens in 22 (4.9%) patients. Eighty-seven (19.2%) patients had a viral infection, and the proportion of those with viral infection was significantly lower in the HCAP than in the CAP group (13.8% vs 24.6%, p = 0.004). In both the HCAP and CAP groups, influenza A was the most common respiratory virus, followed by entero-rhinovirus. The seasonal distributions of respiratory viruses were also similar in both groups. In the HCAP group, the viral infection resulted in a similar length of hospital stay and in-hospital mortality as viral-bacterial coinfection and bacterial infection, and the CAP group showed similar results.The prevalence of viral infection in patients with HCAP was lower than that in patients with CAP, and resulted in a similar prognosis as viral-bacterial coinfection or bacterial infection.

BackgroundSerotonin-also known as 5-hydroxytryptamine or 5-HT-can induce nausea and vomiting (NV) by peripheral mechanisms via the activation of 5-HT3 receptors. In this study, we observed perioperative NV, including intraoperative NV, and changes in serum 5-HT concentrations. We evaluated the relationship between perioperative NV and serum 5-HT levels in patients undergoing cesarean section under epidural anesthesia, and carried out a pilot study to determine if further studies on a larger scale were justified.MethodsTwenty-eight patients who were scheduled for cesarean section under epidural anesthesia were included in the study. Patients were assigned to 2 groups according to the occurrence of NV after induction, i.e., an NV-positive or an NV-negative group. Serum 5-HT concentrations were measured before induction, at the time that NV occurred (in the case of the NV-positive group) or 5 min after the umbilical cord clamping (in the case of the NV-negative group) during surgery, and at 2 h postoperatively.ResultsNV occurred in 10 of the 28 patients. No significant differences in serum 5-HT concentrations were found within or between the two groups.ConclusionsThis study suggests that there is no correlation between serum 5-HT concentration and the occurrence of perioperative NV in patients undergoing cesarean section under epidural anesthesia, and the findings do not seem to support further investigations regarding a possible relationship between serum 5-HT concentration and perioperative NV.

Currently, there are two types of interferon-gamma release assays (IGRAs) in use for the detection of tuberculosis (TB) infection, the QuantiFERON-TB Gold In-Tube test (GFT-GIT) and T-SPOT.TB. Owing to contradictory reports regarding whether the results of these IGRAs are affected by the age of the patient, we aimed to determine if these two tests have age-related differences in sensitivity. We retrospectively reviewed the medical records of diagnosed TB patients who were tested using either QFT-GIT or T-SPOT.TB from February 2008 to December 2013. The positivity of the two tests was analyzed and compared with true TB infection, which was defined as active TB based on either a positive Mycobacterium culture or a positive TB polymerase chain reaction. The QFT-GIT group included 192 TB patients, and the T-SPOT.TB group included 212 TB patients. Of the patients with pulmonary TB, 76 (39.6%) were in the QFT-GIT group and 143 (67.5%) in the T-SPOT.TB group. The overall sensitivity was 80.2% for QFT-GIT and 91.0% for T.SPOT.TB. The sensitivities of QFT-GIT and T-SPOT.TB according to age group were as follows: 70 years, 68.3% and 85.7%, respectively. The trend of age-related changes in sensitivity was significant for both QFT-GIT (p = 0.004) and T.SPOT.TB (p = 0.039). However, only QFT-GIT was significantly related to age in the multivariate analysis. QFT-GIT, but not T-SPOT.TB, was significantly affected by patient age.

Anti-EGFR antibody-based treatment is an important therapeutic strategy for advanced colorectal cancer (CRC); despite this, several mutations--including KRAS, BRAF, and PIK3CA mutations, and HER2 amplification--are associated with the mechanisms underlying the development of resistance to anti-EGFR therapy. The aim of our study was to investigate the frequencies and clinical implications of these genetic alterations in advanced CRC.KRAS, BRAF, and PIK3CA mutations were determined by Cobas real-time polymerase chain reaction (PCR) in 191 advanced CRC patients with distant metastasis. Microsatellite instability (MSI) status was determined by a fragmentation assay and HER2 amplification was assessed by silver in situ hybridization. In addition, KRAS mutations were investigated by the Sanger sequencing method in 97 of 191 CRC cases.Mutations in KRAS, BRAF, and PIK3CA were found in 104 (54.5%), 6 (3.1%), and 25 (13.1%) cases of advanced CRC, respectively. MSI-high status and HER2 amplification were observed in 3 (1.6%) and 16 (8.4%) cases, respectively. PIK3CA mutations were more frequently found in KRAS mutant type (18.3%) than KRAS wild type (6.9%) (P = 0.020). In contrast, HER2 amplifications and BRAF mutations were associated with KRAS wild type with borderline significance (P = 0.052 and 0.094, respectively). In combined analyses with KRAS, BRAF and HER2 status, BRAF mutations or HER2 amplifications were associated with the worst prognosis in the wild type KRAS group (P = 0.004). When comparing the efficacy of detection methods, the results of real time PCR analysis revealed 56 of 97 (57.7%) CRC cases with KRAS mutations, whereas Sanger sequencing revealed 49 cases (50.5%).KRAS mutations were found in 54.5% of advanced CRC patients. Our results support that subgrouping using PIK3CA and BRAF mutation or HER2 amplification status, in addition to KRAS mutation status, is helpful for managing advanced CRC patients.

Osteoporosis-related fractures are one of the complications of Graves' disease. This study hypothesized that the different actions of thyroid-stimulating hormone receptor (TSHR) antibodies, both stimulating and blocking activities in Graves' disease patients might oppositely impact bone turnover. Newly diagnosed premenopausal Graves' disease patients were enrolled (n = 93) and divided into two groups: patients with TSHR antibodies with thyroid-stimulating activity (stimulating activity group, n = 83) and patients with TSHR antibodies with thyroid-stimulating activity combined with blocking activity (blocking activity group, n = 10). From the stimulating activity group, patients who had matched values for free T4 and TSH binding inhibitor immunoglobulin (TBII) to the blocking activity group were further classified as stimulating activity-matched control (n = 11). Bone turnover markers BS-ALP, Osteocalcin, and C-telopeptide were significantly lower in the blocking activity group than in the stimulating activity or stimulating activity-matched control groups. The TBII level showed positive correlations with BS-ALP and osteocalcin levels in the stimulating activity group, while it had a negative correlation with the osteocalcin level in the blocking activity group. In conclusion, the activation of TSHR antibody-activated TSH signaling contributes to high bone turnover, independent of the actions of thyroid hormone, and thyroid-stimulation blocking antibody has protective effects against bone metabolism in Graves' disease.

To determine whether newer combination cytokine treatment with granulocyte colony-stimulating factor (G-CSF) and darbepoetin can improve efficacy of stem cell therapy, we evaluated safety and peripheral blood stem/progenitor cell (PBSC) mobilizing effects of combination cytokine in comparison with G-CSF alone in patients with acute myocardial infarction (AMI). We randomized 60 patients with AMI into two groups under 2:1 ratio; combination treatment with darbepoetin and G-CSF ( n = 41: Combicytokine group) and the G-CSF alone ( n = 19: G-CSF group). After coronary angioplasty, G-CSF was treated for 3 days with dose of 10 μg/kg/day in both groups. Only in the combicytokine group, additional single intravenous injection of 4.5 μg/kg of darbepoetin was administrated immediate after coronary angioplasty. Combination cytokine treatment was well tolerated as was G-CSF alone. PBSCs were obtained by apheresis for intracoronary infusion after completion of cytokine treatment and were analyzed by flow cytometry. The purity of proangiogenic cells was higher in combination cytokine group than the G-CSF group. Specifically, proportion of CD34 + /KDR + endothelial progenitor cells, CD3 + /CD31+ angiogenic T cells and Tie2 + /CXCR4 + cells in apheresis products were higher in the combicytokine group. These meant that the combicytokine treatment recruited PBSCs in higher purity and fewer unwanted inflammatory cells than G-CSF alone in apheresis products. Combination treatment with darbepoetin and G-CSF is safe and more efficient to mobilize and recruit proangiogenic cells than G-CSF alone in patients with AMI. (Trial registration: www.ClinicalTrials.gov identifier: NCT00501917)

Background: Enterovirus D68 (EV-D68) is a re-emerging pathogen that is particularly common in children and may cause asthma-like respiratory infection and acute flaccid myelitis. However, in Korea, EV-D68 has never been reported thus far. This study aimed to identify EV-D68 from nasopharyngeal aspirates (NPAs) in Korean children with a respiratory tract infection. Materials and Methods: The EV-D68 reference strain was purchased and blindly used to assess the detection ability of three commercial and one in-house mRT-PCR kit in 2018. Then, we selected children whose specimens were positive for human rhinovirus (HRV) and/or enterovirus (EV) by Allplex mRT-PCR (Seegene, Inc., Seoul, Korea) from April to December 2022. Total RNA was extracted from NPAs, and a partial 5’-UTR gene was amplified and sequenced for the identification of HRV/EV species. Additionally, PCR targeting the VP1 gene was performed to assess EV-D68- positive NPAs, followed by sequencing. Phylogenetic analysis and comparison of amino acid sequence alignments were performed using a partial VP1 gene of our and recent international EV-D68 strains. Results: Among the mRT-PCR kits tested, only the in-house kit was able to detect EV-D68 in 2018. However, we detected three EV-D68 strains among children hospitalized with fever and/or respiratory symptoms in September - December 2022 who tested positive for EV by the Allplex kit. Two of them were healthy toddlers with lower respiratory infections accompanied by new-onset wheezing but no neurologic complications. Among 34 children with lower respiratory infection who tested positive for HRV during the same period, EV-D68 was not detected. Phylogenetic analysis revealed that the first Korean EV-D68 belonged to subclade B3. Amino acid sequence alignment of international subclade B3 EV-D68 strains also showed that our strain is genetically more related to those from Europe than those from Japan. Conclusion: We first detected EV-D68 in three Korean children who had EV detected by the Allplex mRT-PCR kit in 2022. EV-D68 also circulated in Korea in fall 2022, but the prevalence and severity seemed to be lower than those in previous reports from other countries. [ABSTRACT FROM AUTHOR]