Collection and transportation of SARS-CoV-2 and influenza A virus diagnostic samples: Optimizing the usage of guanidine-based chaotropic salts for enhanced biosafety and viral genome preservation.

Many virus lysis/transport buffers used in molecular diagnostics, including the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, contain guanidine-based chaotropic salts, primarily guanidine hydrochloride (GuHCl) or guanidine isothiocyanate (GITC). Although the virucidal effects of GuHCl and GITC alone against some enveloped viruses have been established, standardized data on their optimum virucidal concentrations against SARS-CoV-2 and effects on viral RNA stability are scarce. Thus, we aimed to determine the optimum virucidal concentrations of GuHCl and GITC against SARS-CoV-2 compared to influenza A virus (IAV), another enveloped respiratory virus. We also evaluated the effectiveness of viral RNA stabilization at the determined optimum virucidal concentrations under high-temperature conditions (35°C) using virus-specific real-time reverse transcription polymerase chain reaction. Both viruses were potently inactivated by 1.0 M GITC and 2.5 M GuHCl, but the GuHCl concentration for efficient SARS-CoV-2 inactivation was slightly higher than that for IAV inactivation. GITC showed better viral RNA stability than GuHCl at the optimum virucidal concentrations. An increased concentration of GuHCl or GITC increased viral RNA degradation at 35°C. Our findings highlight the need to standardize GuHCl and GITC concentrations in virus lysis/transport buffers and the potential application of these guanidine-based salts alone as virus inactivation solutions in SARS-CoV-2 and IAV molecular diagnostics.

Fusion Imaging Objectively Demonstrates Improved Pancreas Visualization through Manipulation Techniques: A Prospective Interventional Study.

Objective Abdominal ultrasonography (AUS) is used to screen for abdominal diseases owing to its low cost, safety, and accessibility. However, the detection rate of pancreatic disease using AUS is unsatisfactory. We evaluated the visualization area of the pancreas and the efficacy of manipulation techniques for AUS with fusion imaging. Methods Magnetic resonance imaging (MRI) volume data were obtained from 20 healthy volunteers in supine and right lateral positions. The MRI volume data were transferred to an ultrasound machine equipped with a fusion imaging software program. We evaluated the visualization area of the pancreas before and after postural changes using AUS with fusion imaging and assessed the liquid-filled stomach method using 500 ml of de-aerated water in 10 randomly selected volunteers. Patients This study included 20 healthy volunteers (19 men and 1 woman) with a mean age of 33.0 (21-37.5) years old. Results Fusion imaging revealed that the visualization area of the entire pancreas using AUS was 55%, which significantly improved to 75% with a postural change and 90% when using the liquid-filled stomach method (p=0.043). Gastrointestinal gas is the main obstacle for visualization of the pancreas. Conclusion Fusion imaging objectively demonstrated that manipulation techniques can improve pancreatic visualization.

Specificity and sensitivity of an indirect fluorescent antibody test to detect antibodies against bovine viral diarrhea virus.

Since being reported in 1979 and 2006, indirect fluorescent antibody (IFA) tests have not been reported to detect bovine viral diarrhea virus (BVDV) antibodies to our knowledge. Thus, we re-evaluated the efficacy and usefulness of IFA tests for BVDV serology. We tested 4 combinations of 2 antibody conjugates (fluorescein isothiocyanate [FITC]-conjugated rabbit IgG anti-bovine IgG; rabbit IgG F(ab')2 fragment anti-bovine IgG [F(ab')2 FITC-IgG]) and 2 washing solutions (PBS; carbonate-bicarbonate-buffered saline [CBBS]) to evaluate the specificity of an IFA test for BVDV. We compared the sensitivity of the optimal combination with virus neutralization (VN) tests and an ELISA, and compared IFA with VN titers against different genotype (subgenotype) strains. For the F(ab')2 FITC-IgG/CBBS combination, only 1 of the 156 (0.6%) 4-fold diluted cattle sera resulted in a nonspecific reaction; other combinations led to a much higher incidence (22.9-37.2%). For the F(ab')2 FITC-IgG/CBBS combination, IFA detection rates were identical (36 of 59) for BVDV1 and BVDV2 genotypes, and IFA titers against them were strongly correlated (r = 0.99). The antibody-detection rates of the IFA tests were almost identical to those of VN tests and the ELISA (κ: 0.96 and 0.89, respectively). The IFA titers against 4 strains (BVDV1a, BVDV1j, BVDV2a, and an unidentified strain) were similar, 1,024 to ≥4,096, although the VN titers were different. Thus, our IFA tests were specific and sensitive, and more useful than VN tests given that the IFA tests could evaluate the immune status of cattle using a representative strain, regardless of genotype (subgenotype).

Partial Stent-in-Stent Method with an Uncovered Self-Expandable Metallic Stent for Unresectable Malignant Hilar Bile Duct Obstruction.

(1) Background: There is controversy regarding stent placement for unresectable malignant hilar biliary obstruction (UMHBO). We mainly use the partial stent-in-stent (PSIS) method with an uncovered self-expandable metallic stent (UCSEMS) based on the drainage area and patency period. In this study, we investigated the usefulness and safety of the PSIS method. (2) Methods: In total, 59 patients who underwent the PSIS method for UMHBO at our hospital were included in the study. The technical success rate, clinical success rate, time to recurrent biliary obstruction (TRBO) and overall survival (OS) from the first placement, factors affecting TRBO and OS, and early complications within 30 days after the procedure were evaluated retrospectively. (3) Results: The technical and clinical success rates were 100% and 96.6%, respectively, with a TRBO of 121 days [95% confidence interval: 82-231] and an OS of 194 days [95% confidence interval: 113-305] after the first placement. Early complications occurred in nine patients (15.3%), including five cases of cholangitis, three cases of pancreatitis, and one case of cholecystitis. (4) Conclusions: The PSIS method for UMHBO is safe and useful with high technical and clinical success rates.

Challenges for Precise Subtyping and Sequencing of a H5N1 Clade 2.3.4.4b Highly Pathogenic Avian Influenza Virus Isolated in Japan in the 2022-2023 Season Using Classical Serological and Molecular Methods.

The continuous evolution of H5Nx highly pathogenic avian influenza viruses (HPAIVs) is a major concern for accurate diagnosis. We encountered some challenges in subtyping and sequencing a recently isolated H5N1 HPAIV strain using classical diagnostic methods. Oropharyngeal, conjunctival, and cloacal swabs collected from a dead white-tailed eagle (Haliaeetus albicilla albicilla) were screened via real-time RT-PCR targeting the influenza A virus matrix (M) gene, followed by virus isolation. The hemagglutination inhibition test was applied in order to subtype and antigenically characterize the isolate using anti-A/duck/Hong Kong/820/80 (H5N3) reference serum or anti-H5N1 cross-clade monoclonal antibodies (mAbs). Sequencing using previously reported universal primers was attempted in order to analyze the full-length hemagglutinin (HA) gene. Oropharyngeal and conjunctival samples were positive for the M gene, and high hemagglutination titers were detected in inoculated eggs. However, its hemagglutination activity was not inhibited by the reference serum or mAbs. The antiserum to a recently isolated H5N1 clade 2.3.4.4b strain inhibited our isolate but not older strains. A homologous sequence in the previously reported forward primer and HA2 region in our isolate led to partial HA gene amplification. Finally, next-generation sequencing confirmed the isolate as H5N1 clade 2.3.4.4b HPAIV, with genetic similarity to H5N1 strains circulating in Japan since November 2021.

A Novel Monochromatic Light to Detect Target Specimens Within Endoscopic Ultrasonography-guided Fine-needle Aspiration Biopsy Samples.

BACKGROUND/AIM: Endoscopic ultrasonography (EUS)-guided fine-needle aspiration biopsy (EUS-FNB) enhances the diagnostic capabilities of EUS by providing additional pathological samples. However, detecting the target specimens within the collected samples can be challenging. The aim of this study was to determine the optimal wavelength of light for detection of target specimens within EUS-FNB samples in an animal experiment. MATERIALS AND METHODS: EUS-FNB pancreatic tissue samples were collected from a male beagle (weight, 10 kg), and the samples were illuminated with monochromatic light ranging from 430 to 700 nm in 5-nm intervals. The intensities of the target specimen and blood samples were analyzed using the densitometry of the images obtained through irradiation. RESULTS: We found that transmitted monochromatic light of 605 nm most vividly enhanced the contrast between the target specimens and blood in the samples in the impression of appearance. CONCLUSION: Thus, microscopical observations under transmitted light of 605 nm are optimal for target tissue identification within EUS-FNB samples.

Safety and Diagnostic Yield of Endoscopic Ultrasound-Guided Fine-Needle Biopsy for Hypervascular Pancreatic Lesions.

Endoscopic ultrasound-guided fine-needle biopsy (EUS-FNB) is a common technique for diagnosing pancreatic lesions with high accuracy and a low incidence of procedural adverse events. However, occasional adverse events, particularly bleeding, may occur. Procedures for hypervascular lesions are considered important, but their risks are unknown. We aimed to evaluate the safety and diagnostic yield of EUS-FNB for hypervascular pancreatic solid lesions. This study included 301 patients with 308 solid pancreatic lesions who underwent EUS-FNB between May 2011 and December 2018. We performed propensity-score matching to balance clinical differences between hypervascular and hypovascular lesions and analyzed 52 lesions. We compared the safety and diagnostic performance of propensity score-matched cohorts. The sensitivity, specificity, and accuracy rates of EUS-FNB for hypervascular lesions were 94.7%, 100%, and 96.2%, and those for hypovascular lesions were 80.0%, 100%, and 84.6%, respectively. There was no difference in diagnostic performance between hypervascular and hypovascular lesions. Furthermore, adverse events occurred in only one patient (pancreatitis) in the hypovascular group. There were no significant differences in the occurrence of adverse events between hypervascular and hypovascular lesions (0% vs. 3.8%, p = 1.000). Therefore, EUS-FNB may be safe with a high diagnostic yield, even for hypervascular solid pancreatic lesions.

An Abies Extract Containing Nonvolatile Polyphenols Shows Virucidal Activity against SARS-CoV-2 That Is Enhanced in Increased pH Conditions.

Researching the beneficial health properties of wood byproducts can prevent wastage by turning them into valuable resources. In this study, the virucidal activity of two extracts from Abies sachalinensis byproducts, ASE1, and ASE2, against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was investigated. ASE1 is rich in monoterpenoid volatile compounds, whereas ASE2 contains nonvolatile polyphenols. SARS-CoV-2 solutions were mixed with ASE1 or ASE2, and viral titer reduction was evaluated. At their original acidic pH, ASE2 showed stronger virucidal activity than ASE1. The virucidal activity of ASE2 was also significantly enhanced when pH was increased to neutral or basic, which was not the case for ASE1. At a neutral pH, ASE2 induced statistically significant viral titer reduction in 1 min. HCl and NaOH solutions, which had a pH close to that of acidic and basic ASE2 test mixtures, respectively, exhibited no virucidal activity against SARS-CoV-2. Among the SARS-CoV-2 variants, Omicron showed the highest vulnerability to ASE2. Western blotting, RT-PCR, and electron microscopic analysis revealed that neutral ASE2 interacts with SARS-CoV-2 spike proteins and moderately disrupts the SARS-CoV-2 genome and viral envelope. These findings reveal the virucidal potential of ASE2.

Inhibition of Toll-like receptor 4 and Interleukin-1 receptor prevent SARS-CoV-2 mediated kidney injury.

Acute kidney injury (AKI) is a common and severe complication of the coronavirus disease 2019 (COVID-19). Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) directly affects the glomerular and tubular epithelial cells to induce AKI; however, its pathophysiology remains unclear. Here, we explored the underlying mechanisms and therapeutic targets of renal involvement in COVID-19. We developed an in vitro human kidney cellular model, including immortalized tubular epithelial and endothelial cell lines, demonstrating that SARS-CoV-2 directly triggers cell death. To identify the molecular targets in the process of SARS-CoV-2-mediated cell injury, we performed transcriptional analysis using RNA sequencing. Tubular epithelial cells were more prone to dying by SARS-CoV-2 than endothelial cells; however, SARS-CoV-2 did not replicate in renal cells, distinct from VeroE6/transmembrane protease serine 2 cells. Transcriptomic analysis revealed increased inflammatory and immune-related gene expression levels in renal cells incubated with SARS-CoV-2. Toll-like receptor (TLR) 3 in renal cells recognized viral RNA and underwent cell death. Furthermore, analysis of upstream regulators identified several key transcriptional regulators. Among them, inhibition of the interleukin-1 receptor (IL-1R) and TLR4 pathways protects tubular epithelial and endothelial cells from injury via regulation of the signal transducer and activator of transcription protein-3/nuclear factor-kB pathway. Our results reveal that SARS-CoV-2 directly injures renal cells via the proinflammatory response without viral replication, and that IL-1R and TLR4 may be used as therapeutic targets for SARS-CoV-2 mediated kidney injury.

Current status of molecular diagnostic approaches using liquid biopsy.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal cancers, and developing an efficient and reliable approach for its early-stage diagnosis is urgently needed. Precancerous lesions of PDAC, such as pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasms (IPMN), arise through multiple steps of driver gene alterations in KRAS, TP53, CDKN2A, SMAD4, or GNAS. Hallmark mutations play a role in tumor initiation and progression, and their detection in bodily fluids is crucial for diagnosis. Recently, liquid biopsy has gained attention as an approach to complement pathological diagnosis, and in addition to mutation signatures in cell-free DNA, cell-free RNA, and extracellular vesicles have been investigated as potential diagnostic and prognostic markers. Integrating such molecular information to revise the diagnostic criteria for pancreatic cancer can enable a better understanding of the pathogenesis underlying inter-patient heterogeneity, such as sensitivity to chemotherapy and disease outcomes. This review discusses the current diagnostic approaches and clinical applications of genetic analysis in pancreatic cancer and diagnostic attempts by liquid biopsy and molecular analyses using pancreatic juice, duodenal fluid, and blood samples. Emerging knowledge in the rapidly advancing liquid biopsy field is promising for molecular profiling and diagnosing pancreatic diseases with significant diversity.

Pancreatic Juice Cytology Evaluations Using Synthetic Secretin and Serial Pancreatic Juice Aspiration Cytological Examination for the Diagnosis of Pancreatic Ductal Adenocarcinoma.

Pathological examination by endoscopic ultrasound-fine needle aspiration is not possible in approximately 10% of pancreatic tumor cases. Pancreatic juice cytology (PJC) is considered an alternative diagnostic method. However, its diagnostic capability is insufficient, and PJC has been repeatedly redevised. Serial pancreatic juice aspiration cytological examination (SPACE) and secretin-loaded PJC (S-PJC) have been recently introduced as alternative diagnostic methods. This study aimed to determine the diagnostic capacity and safety of SPACE and S-PJC using a propensity score-matched analysis. The sensitivity, specificity, and accuracy were 75.0%, 100%, and 92.3% for S-PJC, respectively, and 71.4%, 100%, and 92.3% for SPACE, respectively, meaning that there was no significant difference between the groups. Four patients (15.4%) each in the S-PJC and SPACE groups experienced complications, including postendoscopic retrograde cholangiopancreatography, pancreatitis, and cholangitis. Overall, there was no difference in efficacy and safety between the SPACE and S-PJC groups.

Rapid Virucidal Activity of Japanese Saxifraga Species-Derived Condensed Tannins against SARS-CoV-2, Influenza A Virus, and Human Norovirus Surrogate Viruses.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus (IAV), and norovirus are global threats to human health. The application of effective virucidal agents, which contribute to the inactivation of viruses on hands and environmental surfaces, is important to facilitate robust virus infection control measures. Naturally derived virucidal disinfectants have attracted attention owing to their safety and eco-friendly properties. In this study, we showed that multiple Japanese Saxifraga species-derived fractions demonstrated rapid, potent virucidal activity against the SARS-CoV-2 ancestral strain and multiple variant strains, IAV, and two human norovirus surrogates: feline calicivirus (FCV) and murine norovirus (MNV). Condensed tannins were identified as active chemical constituents that play a central role in the virucidal activities of these fractions. At a concentration of 25 µg/mL, the purified condensed tannin fraction Sst-2R induced significant reductions in the viral titers of the SARS-CoV-2 ancestral strain, IAV, and FCV (reductions of ≥3.13, ≥3.00, and 2.50 log10 50% tissue culture infective doses [TCID50]/mL, respectively) within 10 s of reaction time. Furthermore, at a concentration of 100 µg/mL, Sst-2R induced a reduction of 1.75 log10 TCID50/mL in the viral titers of MNV within 1 min. Western blotting and transmission electron microscopy analyses revealed that Sst-2R produced structural abnormalities in viral structural proteins and envelopes, resulting in the destruction of viral particles. Furthermore, Saxifraga species-derived fraction-containing cream showed virucidal activity against multiple viruses within 10 min. Our findings indicate that Saxifraga species-derived fractions containing condensed tannins can be used as disinfectants against multiple viruses on hands and environmental surfaces. IMPORTANCE SARS-CoV-2, IAV, and norovirus are highly contagious pathogens. The use of naturally derived components as novel virucidal/antiviral agents is currently attracting attention. We showed that fractions from extracts of Saxifraga species, in the form of a solution as well as a cream, exerted potent, rapid virucidal activities against SARS-CoV-2, IAV, and surrogates of human norovirus. Condensed tannins were found to play a central role in this activity. The in vitro cytotoxicity of the purified condensed tannin fraction at a concentration that exhibited some extent of virucidal activity was lower than that of 70% ethanol or 2,000 ppm sodium hypochlorite solution, which are popular virucidal disinfectants. Our study suggests that Saxifraga species-derived fractions containing condensed tannins can be used on hands and environmental surfaces as safe virucidal agents against multiple viruses.

Comparison between Target Sample Check Illuminator and White Light Observation in Discriminating the Presence of Target Specimen for Endoscopic Ultrasound-Guided Fine Needle Aspiration Sample.

Endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) is an essential endoscopic tissue sampling method for diagnosing pancreatobiliary diseases; however, determining the presence of target specimens mixed in the blood by conventional observation is challenging due to the small size of the obtained sample. This study investigated the usefulness of a target sample check illuminator (TSCI) that emits a specific wavelength of light to determine the presence of target specimens. Twenty-seven patients who underwent EUS-FNA at our hospital were included. Conventional white light observation was performed for the collected samples, followed by TSCI; six people evaluated the presence of the target specimen on a 5-point scale. The target specimen discrimination score using TSCI (median: 5) was significantly higher than that using conventional white light observation (median: 1) (p < 0.001). No significant difference was observed in the discrimination score between the evaluator (novice vs. expert, p = 0.162) and puncture needle (22G vs. 25G, p = 0.196). The discriminability of TSCI in the samples obtained using EUS-FNA was significantly higher than that of conventional observation. TSCI does not depend on the evaluator or puncture needle for the identification of the target specimen; hence, it can provide a good pathological specimen and may contribute to the improvement of the diagnostic ability.

Efficacy Validation of SARS-CoV-2-Inactivation and Viral Genome Stability in Saliva by a Guanidine Hydrochloride and Surfactant-Based Virus Lysis/Transport Buffer.

To enhance biosafety and reliability in SARS-CoV-2 molecular diagnosis, virus lysis/transport buffers should inactivate the virus and preserve viral RNA under various conditions. Herein, we evaluated the SARS-CoV-2-inactivating activity of guanidine hydrochloride (GuHCl)- and surfactant (hexadecyltrimethylammonium chloride (Hexa-DTMC))-based buffer, Prep Buffer A, (Precision System Science Co., Ltd., Matsudo, Japan) and its efficacy in maintaining the stability of viral RNA at different temperatures using the traditional real-time one-step RT-PCR and geneLEAD VIII sample-to-result platform. Although Prep Buffer A successfully inactivated SARS-CoV-2 in solutions with high and low organic substance loading, there was considerable viral genome degradation at 35 °C compared with that at 4 °C. The individual roles of GuHCl and Hexa-DTMC in virus inactivation and virus genome stability at 35 °C were clarified. Hexa-DTMC alone (0.384%), but not 1.5 M GuHCl alone, exhibited considerable virucidal activity, suggesting that it was essential for potently inactivating SARS-CoV-2 using Prep Buffer A. GuHCl and Hexa-DTMC individually reduced the viral copy numbers to the same degree as Prep Buffer A. Although both components inhibited RNase activity, Hexa-DTMC, but not GuHCl, directly destroyed naked viral RNA. Our findings suggest that samples collected in Prep Buffer A should be stored at 4 °C when RT-PCR will not be performed for several days.

Virucidal Activities of Acidic Electrolyzed Water Solutions with Different pH Values against Multiple Strains of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a threat to human health. Acidic electrolyzed water (AEW) has recently been suggested to demonstrate virucidal activity. Many types of AEW with different pH values, generated by the electrolysis of different chemicals, such as sodium chloride, potassium chloride, and hydrochloric acid, are commercially available. In this study, we compared the virucidal activities of these types of AEW against SARS-CoV-2, including the ancestral strain and variant Alpha, Beta, Gamma, Delta, and Omicron strains. Virus solution (viral titer, 6.9 log10 50% tissue culture infective dose [TCID50]/mL) was mixed with AEW (free available chlorine concentration, 34.5 ppm) at mixing ratios of 1:9, 1:19, and 1:49. At mixing ratios of 1:9 and 1:19, AEW with a pH of 2.8 showed stronger virucidal activities than AEW with a pH of 4.1 to 6.5 against the SARS-CoV-2 ancestral strain in 20 s. From the strongest to the weakest virucidal activity, the AEW pH levels were as follows: pH 2.8, pH 4.1 to 5.4, pH 6.4 to 6.5. At a ratio of 1:49, the viral titers of viruses treated with all AEW solutions at pH 2.8 to 6.5 were almost below the detection limit, which was 1.25 log10 TCID50/mL. The virus inactivation efficiency of AEW was reduced in the presence of fetal bovine serum and other substances contained in the virus solution used in this study. AEW with pH values of 2.8 to 6.5 showed virucidal activity against all of the tested SARS-CoV-2 strains, including the ancestral and variant strains. These results provide useful knowledge for the effective application of AEW as a SARS-CoV-2 disinfectant. IMPORTANCE Acidic electrolyzed water (AEW) demonstrates virucidal activity against multiple viruses. Since AEW exhibits low toxicity, is inexpensive, and is environmentally friendly, it can be a useful disinfectant against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Although the pH values of currently available AEW products vary, the impact of different pH values on SARS-CoV-2 inactivation has not previously been evaluated in detail. In this study, we compared the virucidal activities of multiple AEW solutions with different pH values, under the same experimental conditions. We found that AEW solutions with lower pH values demonstrated more potent virucidal activity. Also, we showed that the extent of virus inactivation by the AEW was based on the balance of the abundance of free available chlorine, virus, and other organic substances in the mixture. AEW exhibited rapid virucidal activity against multiple SARS-CoV-2 strains. This study demonstrated the usefulness of AEW as a disinfectant which can be applied to the inactivation of SARS-CoV-2.

Current status and future perspective of linked color imaging for gastric cancer screening: a literature review.

Screening endoscopy has advanced to facilitate improvements in the detection and prognosis of gastric cancer. However, most early gastric cancers (EGCs) have subtle morphological or color features that are difficult to detect by white-light imaging (WLI); thus, even well-trained endoscopists can miss EGC when using this conventional endoscopic approach. This review summarizes the current and future status of linked color imaging (LCI), a new image-enhancing endoscopy (IEE) method, for gastric screening. LCI has been shown to produce bright images even at a distant view and provide excellent visibility of gastric cancer due to high color contrast relative to the surrounding tissue. LCI delineates EGC as orange-red and intestinal metaplasia as purple, regardless of a history of Helicobacter pylori (Hp) eradication, and contributes to the detection of superficial EGC. Moreover, LCI assists in the determination of Hp infection status, which is closely related to the risk of developing gastric cancer. Transnasal endoscopy (ultra-thin) using LCI is also useful for identifying gastric neoplastic lesions. Recently, several prospective studies have demonstrated that LCI has a higher detection ratio for gastric cancer than WLI. We believe that LCI should be used in routine upper gastrointestinal endoscopies.

Transcriptional dynamics of granulocytes in direct response to incubation with SARS-CoV-2.

Severe coronavirus disease 2019 (COVID-19) is characterized by acute respiratory distress syndrome and multiple organ dysfunction, in which the host immune response plays a pivotal role. Excessive neutrophil activation and subsequent superfluity of neutrophil extracellular traps (NETs) can lead to tissue damage, and several studies have shown the involvement of neutrophils in severe COVID-19. However, the detailed responses of each neutrophil subset to SARS-CoV-2 infection has not been fully described. To explore this issue, we incubated normal-density granulocytes (NDGs) and low-density granulocytes (LDGs) with different viral titers of SARS-CoV-2. NDGs form NETs with chromatin fibers in response to SARS-CoV-2, whereas LDGs incubated with SARS-CoV-2 display a distinct morphology with condensed nuclei and moderate transcriptional changes. Based on these transcriptional changes, we suggest that AGO2 possibly plays a role in LDG regulation in response to SARS-CoV-2.

The Usefulness of Peroral Cholangioscopy for Intrahepatic Stones.

Peroral cholangioscopy (POCS) is believed to be effective in treating intrahepatic stones; however, reports on its efficacy are few. We reviewed the results of intrahepatic stones treated with fluoroscopic guidance or POCS. This study included 26 patients who underwent endoscopic treatment for intrahepatic stones at our institution between January 2017 and December 2021. We retrospectively evaluated the procedure time and adverse events in the first session and the rate of complete stone removal. Complete stone removal was achieved in 92% (24/26); POCS was required in 16 of 26 (62%) procedures and the complete stone removal was achieved in 15 of 16 (94%) of these procedures. The POCS group had a significantly longer procedure time than the fluoroscopy group. Cholangitis incidence was high; however, no difference was noted between patients with and without POCS, and all cases were mild and treated conservatively. Endoscopic treatment for intrahepatic stones may lead to an increase in the incidence of cholangitis, requires specialized devices such as a cholangioscope, and should be performed in an established institution by experienced staff. POCS is useful for intrahepatic stones formed upstream of the stenosis and intrahepatic stones piled in the bile duct.

Viral RNA extraction using an automatic nucleic acid extractor with magnetic particles and genetic characterization of bovine viral diarrhea virus in Tokachi Province, Japan, in 2016-2017.

In this study, the viral genome extraction performance of automatic nucleic acid extractors and manual nucleic acid extraction kits was compared. We showed that compared with manual kits, the automatic extractors showed superior genome extraction performance using bovine viral diarrhea virus (BVDV) genome-positive cattle sera and bovine coronavirus/infectious bovine rhinotracheitis virus-spiked cattle nasal swabs. In addition, the subgenotyping of BVDV strains detected in Tokachi Province in Japan during 2016-2017 was performed. Results showed that most of these BVDV strains belonged to subgenotype 1b, while few strains belonged to subgenotypes 1a and 2a. This study showed the high applicability of automatic nucleic acid extractors in extracting multiple viral genomes and the dominant subgenotype of BVDV in Tokachi.