The endemicity of highly pathogenic avian influenza (HPAI) A(H5N1) viruses in Asia has led to the generation of reassortant H5 strains with novel gene constellations. A newly emerged HPAI A(H5N8) virus caused poultry outbreaks in the Republic of Korea in 2014. Because newly emerging high-pathogenicity H5 viruses continue to pose public health risks, it is imperative that their pathobiological properties be examined. Here, we characterized A/mallard duck/Korea/W452/2014 (MDk/W452(H5N8)), a representative virus, and evaluated its pathogenic and pandemic potential in various animal models. We found that MDk/W452(H5N8), which originated from the reassortment of wild bird viruses harbored by migratory waterfowl in eastern China, replicated systemically and was lethal in chickens, but appeared to be attenuated, albeit efficiently transmitted, in ducks. Despite predominant attachment to avian-like virus receptors, MDk/W452(H5N8) also exhibited detectable human virus-like receptor binding and replicated in human respiratory tract tissues. In mice, MDk/W452(H5N8) was moderately pathogenic and had limited tissue tropism relative to previous HPAI A(H5N1) viruses. It also induced moderate nasal wash titers in inoculated ferrets; additionally, it was recovered in extrapulmonary tissues and one of three direct-contact ferrets seroconverted without shedding. Moreover, domesticated cats appeared to be more susceptible than dogs to virus infection. With their potential to become established in ducks, continued circulation of A(H5N8) viruses could alter the genetic evolution of pre-existing avian poultry strains. Overall, detailed virological investigation remains a necessity given the capacity of H5 viruses to evolve to cause human illness with few changes in the viral genome.

Targeted inactivation of Runx3 in mouse lung induced mucinous and nonmucinous adenomas and markedly shortened latency of adenocarcinoma formation induced by oncogenic K-Ras. RUNX3 was frequently inactivated in K-RAS mutated human lung adenocarcinomas. A functional genetic screen of a fly mutant library and molecular analysis in cultured cell lines revealed that Runx3 forms a complex with BRD2 in a K-Ras-dependent manner in the early phase of the cell cycle; this complex induces expression of p14(ARF)/p19(Arf) and p21(WAF/CIP). When K-Ras was constitutively activated, the Runx3-BRD2 complex was stably maintained and expression of both p14(ARF) and p21(WAF/CIP) was prolonged. These results provide a missing link between oncogenic K-Ras and the p14(ARF)-p53 pathway, and may explain how cells defend against oncogenic K-Ras.

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While the seroprevalence of SARS-CoV-2 in healthy people does not differ significantly among age groups, those aged 65 years or older exhibit strikingly higher COVID-19 mortality compared to younger individuals. To further understand differing COVID-19 manifestations in patients of different ages, three age groups of ferrets are infected with SARS-CoV-2. Although SARS-CoV-2 is isolated from all ferrets regardless of age, aged ferrets (≥3 years old) show higher viral loads, longer nasal virus shedding, and more severe lung inflammatory cell infiltration, and clinical symptoms compared to juvenile (≤6 months) and young adult (1–2 years) groups. Furthermore, direct contact ferrets co-housed with the virus-infected aged group shed more virus than direct-contact ferrets co-housed with virus-infected juvenile or young adult ferrets. Transcriptome analysis of aged ferret lungs reveals strong enrichment of gene sets related to type I interferon, activated T cells, and M1 macrophage responses, mimicking the gene expression profile of severe COVID-19 patients. Thus, SARS-CoV-2-infected aged ferrets highly recapitulate COVID-19 patients with severe symptoms and are useful for understanding age-associated infection, transmission, and pathogenesis of SARS-CoV-2.

Highly sensitive methods, such as PNA clamping, may be superior to direct sequencing for the detection of EGFR mutations in diagnostic specimens with a low proportion of tumor cells. Direct sequencing may be more representative when diagnostic specimens with a high proportion of tumor cells are available.

Circulating cell-free microRNAs (miRNAs) are potential cancer biomarkers. The aim of this study was to identify miRNAs that are differentially expressed between benign pleural effusion (BPE) and lung adenocarcinoma-associated malignant pleural effusion (LA-MPE). The expression level of cell-free miRNA was investigated in 107 patients with pleural effusion. Microarrays were used to screen 160 miRNAs in a discovery set comprising 20 effusion samples (ten BPEs and ten LA-MPEs). Real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to validate the profiling results obtained for the discovery set and those obtained for a validation set comprising 42 BPEs and 45 LA-MPEs. The area under the receiver operating characteristic curve (AUC) was used to evaluate the diagnostic performance of the identified miRNAs and other common tumor markers, such as carcinoembryonic antigen (CEA) and cytokeratin fragment (CYFRA) 21-1. Microarray profiling showed that miR-198 was significantly downregulated in LA-MPE compared with BPE (p 5 0.002). The miRNA microarray analysis results were confirmed by qRT-PCR (p < 0.001) using the validation set. The AUCs for miR-198, CEA and CYFRA 21-1 in the validation set were 0.887, 0.898 and 0.836, respectively. The diagnostic performance of miR-198 was comparable with that of CEA, but better than that of CYFRA 21-1. The AUC for all three markers combined was 0.926 (95% confidence interval, 0.843-0.973) with a sensitivity of 89.2% and a specificity of 85.0%. The present study suggests that cell-free miR-198 from patients with pleural effusion might have diagnostic potential for differentiating LA-MPE from BPE.

Highly pathogenic avian influenza (HPAI) A(H5N6) and A(H5N8) virus infections resulted in the culling of more than 37 million poultry in the Republic of Korea during the 2016/17 winter season. Here we characterize two representative viruses, A/Environment/Korea/W541/2016 [Em/W541(H5N6)] and A/Common Teal/Korea/W555/2017 [CT/W555(H5N8)], and evaluate their zoonotic potential in various animal models. Both Em/W541(H5N6) and CT /W555(H5N8) are novel reassortants derived from various gene pools of wild bird viruses present in migratory waterfowl arising from eastern China. Despite strong preferential binding to avian virus–type receptors, the viruses were able to grow in human respiratory tract tissues. Em/W541(H5N6) was found to be highly pathogenic in both chickens and ducks, while CT/W555(H5N8) caused lethal infections in chickens but did not induce remarkable clinical illness in ducks. In mice, both viruses appeared to be moderately pathogenic and displayed limited tissue tropism relative to HPAI H5N1 viruses. Em/W541(H5N6) replicated to moderate levels in the upper respiratory tract of ferrets and was detected in the lungs, brain, spleen, liver, and colon. Unexpectedly, two of three ferrets in direct contact with Em/W541(H5N6)-infected animals shed virus and seroconverted at 14 dpi. CT/W555(H5N8) was less pathogenic than the H5N6 virus in ferrets and no transmission was detected. Given the co-circulation of different, phenotypically distinct, subtypes of HPAI H5Nx viruses for the first time in South Korea, detailed virologic investigations are imperative given the capacity of these viruses to evolve and cause human infections.

Liquid-based cytology (LBC) is being increasingly used for body fluid specimens and has improved diagnostic accuracy when compared to conventional smears. We compared the diagnostic accuracy and cellular morphologic features between CellprepPlus® LBC and ThinPrep® LBC in effusion cytology. One hundred and eighty body fluid specimens, consisting of 119 pleural fluid specimens, 59 peritoneal fluid specimens, and 2 pericardial fluid specimens, were obtained from 166 patients. Equal volumes of body fluid from each specimen were used in the CellprepPlus® and ThinPrep® preparations. Sensitivity, specificity, and positive and negative predictive values were evaluated. In addition, we selected 16 specimens from patients with metastatic adenocarcinoma, confirmed them by both LBC preparations, and measured the size of the nucleus in the tumor cells in these specimens. The sensitivity of the CellprepPlus® and ThinPrep® methods was 73.1% and 50.0%, respectively. The specificity and positive predictive values were 100% for both LBC methods, and the negative predictive values of the CellprepPlus® and ThinPrep® methods were 90.9% and 83.3%, respectively. The average nuclear size of the tumor cells was calculated as 20.87 μm using the CellprepPlus® method and 15.08 μm using the ThinPrep® method (P < 0.05). The CellprepPlus® method provided better diagnostic accuracy of effusion cytology compared to the ThinPrep® method and revealed the characteristic morphological features of tumor cells, including large and hypochromatic nuclei, prominent nucleoli, distinct nuclear membranes, and high cellularity.