Augmenting Neutrophil Extracellular Traps with Carbonized Polymer Dots: A Potential Treatment for Bacterial Sepsis.

Sepsis is a life-threatening condition that can progress to septic shock as the body's extreme response to pathogenesis damages its own vital organs. Staphylococcus aureus (S. aureus) accounts for 50% of nosocomial infections, which are clinically treated with antibiotics. However, methicillin-resistant strains (MRSA) have emerged and can withstand harsh antibiotic treatment. To address this problem, curcumin (CCM) is employed to prepare carbonized polymer dots (CPDs) through mild pyrolysis. Contrary to curcumin, the as-formed CCM-CPDs are highly biocompatible and soluble in aqueous solution. Most importantly, the CCM-CPDs induce the release of neutrophil extracellular traps (NETs) from the neutrophils, which entrap and eliminate microbes. In an MRSA-induced septic mouse model, it is observed that CCM-CPDs efficiently suppress bacterial colonization. Moreover, the intrinsic antioxidative, anti-inflammatory, and anticoagulation activities resulting from the preserved functional groups of the precursor molecule on the CCM-CPDs prevent progression to severe sepsis. As a result, infected mice treated with CCM-CPDs show a significant decrease in mortality even through oral administration. Histological staining indicates negligible organ damage in the MRSA-infected mice treated with CCM-CPDs. It is believed that the in vivo studies presented herein demonstrate that multifunctional therapeutic CPDs hold great potential against life-threatening infectious diseases.

New Advances in Rapid Pretreatment for Small Dense LDL Cholesterol Measurement Using Shear Horizontal Surface Acoustic Wave (SH-SAW) Technology.

Atherosclerosis is an inflammatory disease of the arteries associated with alterations in lipid and other metabolism and is a major cause of cardiovascular disease (CVD). LDL consists of several subclasses with different sizes, densities, and physicochemical compositions. Small dense LDL (sd-LDL) is a subclass of LDL. There is growing evidence that sd-LDL-C is associated with CVD risk, metabolic dysregulation, and several pathophysiological processes. In this study, we present a straightforward membrane device filtration method that can be performed with simple laboratory methods to directly determine sd-LDL in serum without the need for specialized equipment. The method consists of three steps: first, the precipitation of lipoproteins with magnesium harpin; second, the collection of effluent from a 100 nm filter; and third, the quantification of sd-LDL-ApoB in the effluent with an SH-SAW biosensor. There was a good correlation between ApoB values obtained using the centrifugation (y = 1.0411x + 12.96, r = 0.82, n = 20) and filtration (y = 1.0633x + 15.13, r = 0.88, n = 20) methods and commercially available sd-LDL-C assay values. In addition to the filtrate method, there was also a close correlation between sd-LDL-C and ELISA assay values (y = 1.0483x - 4489, r = 0.88, n = 20). The filtration treatment method also showed a high correlation with LDL subfractions and NMR spectra ApoB measurements (y = 2.4846x + 4.637, r = 0.89, n = 20). The presence of sd-LDL-ApoB in the effluent was also confirmed by ELISA assay. These results suggest that this filtration method is a simple and promising pretreatment for use with the SH-SAW biosensor as a rapid in vitro diagnostic (IVD) method for predicting sd-LDL concentrations. Overall, we propose a very sensitive and specific SH-SAW biosensor with the ApoB antibody in its sensitive region to monitor sd-LDL levels by employing a simple delay-time phase shifted SH-SAW device. In conclusion, based on the demonstration of our study, the SH-SAW biosensor could be a strong candidate for the future measurement of sd-LDL.

Development of antiviral carbon quantum dots that target the Japanese encephalitis virus envelope protein.

Japanese encephalitis is a mosquito-borne disease caused by the Japanese encephalitis virus (JEV) that is prevalent in Asia and the Western Pacific. Currently, there is no effective treatment for Japanese encephalitis. Curcumin (Cur) is a compound extracted from the roots of Curcuma longa, and many studies have reported its antiviral and anti-inflammatory activities. However, the high cytotoxicity and very low solubility of Cur limit its biomedical applications. In this study, Cur carbon quantum dots (Cur-CQDs) were synthesized by mild pyrolysis-induced polymerization and carbonization, leading to higher water solubility and lower cytotoxicity, as well as superior antiviral activity against JEV infection. We found that Cur-CQDs effectively bound to the E protein of JEV, preventing viral entry into the host cells. In addition, after continued treatment of JEV with Cur-CQDs, a mutant strain of JEV was evolved that did not support binding of Cur-CQDs to the JEV envelope. Using transmission electron microscopy, biolayer interferometry, and molecular docking analysis, we revealed that the S123R and K312R mutations in the E protein play a key role in binding Cur-CQDs. The S123 and K312 residues are located in structural domains II and III of the E protein, respectively, and are responsible for binding to receptors on and fusing with the cell membrane. Taken together, our results suggest that the E protein of flaviviruses represents a potential target for the development of CQD-based inhibitors to prevent or treat viral infections.

Potential relevance of salivary legumain for the clinical diagnostic of hand, foot, and mouth disease.

The fight against hand, foot, and mouth disease (HFMD) remains an arduous challenge without existing point-of-care (POC) diagnostic platforms for accurate diagnosis and prompt case quarantine. Hence, the purpose of this salivary biomarker discovery study is to set the fundamentals for the realization of POC diagnostics for HFMD. Whole salivary proteome profiling was performed on the saliva obtained from children with HFMD and healthy children, using a reductive dimethylation chemical labeling method coupled with high-resolution mass spectrometry-based quantitative proteomics technology. We identified 19 upregulated (fold change = 1.5-5.8) and 51 downregulated proteins (fold change = 0.1-0.6) in the saliva samples of HFMD patients in comparison to that of healthy volunteers. Four upregulated protein candidates were selected for dot blot-based validation assay, based on novelty as biomarkers and exclusions in oral diseases and cancers. Salivary legumain was validated in the Singapore (n = 43 healthy, 28 HFMD cases) and Taiwan (n = 60 healthy, 47 HFMD cases) cohorts with an area under the receiver operating characteristic curve of 0.7583 and 0.8028, respectively. This study demonstrates the feasibility of a broad-spectrum HFMD POC diagnostic test based on legumain, a virus-specific host systemic signature, in saliva.

Role of Enteroviral RNA-Dependent RNA Polymerase in Regulation of MDA5-Mediated Beta Interferon Activation.

Infection by enteroviruses can cause severe neurological complications in humans. The interactions between the enteroviral and host proteins may facilitate the virus replication and be involved in the pathogenicity of infected individuals. It has been shown that human enteroviruses possess various mechanisms to suppress host innate immune responses in infected cells. Previous studies showed that infection by enterovirus 71 (EV71) causes the degradation of MDA5, which is a critical cytoplasmic pathogen sensor in the recognition of picornaviruses for initiating transcription of type I interferons. In the present study, we demonstrated that the RNA-dependent RNA polymerase (RdRP; also denoted 3Dpol) encoded by EV71 interacts with the caspase activation and recruitment domains (CARDs) of MDA5 and plays a role in the inhibition of MDA5-mediated beta interferon (IFN-ß) promoter activation and mRNA expression. In addition, we found that the 3Dpol protein encoded by coxsackievirus B3 also interacted with MDA5 and downregulated the antiviral signaling initiated by MDA5. These findings indicate that enteroviral RdRP may function as an antagonist against the host antiviral innate immune response.IMPORTANCE Infection by enteroviruses causes severe neurological complications in humans. Human enteroviruses possess various mechanisms to suppress the host type I interferon (IFN) response in infected cells to establish viral replication. In the present study, we found that the enteroviral 3Dpol protein (or RdRP), which is a viral RNA-dependent RNA polymerase for replicating viral RNA, plays a role in the inhibition of MDA5-mediated beta interferon (IFN-ß) promoter activation. We further demonstrated that enteroviral 3Dpol protein interacts with the caspase activation and recruitment domains (CARDs) of MDA5. These findings indicate that enteroviral RdRP functions as an antagonist against the host antiviral response.

Hijacking of multiple phospholipid biosynthetic pathways and induction of membrane biogenesis by a picornaviral 3CD protein.

RNA viruses induce specialized membranous structures for use in genome replication. These structures are often referred to as replication organelles (ROs). ROs exhibit distinct lipid composition relative to other cellular membranes. In many picornaviruses, phosphatidylinositol-4-phosphate (PI4P) is a marker of the RO. Studies to date indicate that the viral 3A protein hijacks a PI4 kinase to induce PI4P by a mechanism unrelated to the cellular pathway, which requires Golgi-specific brefeldin A-resistance guanine nucleotide exchange factor 1, GBF1, and ADP ribosylation factor 1, Arf1. Here we show that a picornaviral 3CD protein is sufficient to induce synthesis of not only PI4P but also phosphatidylinositol-4,5-bisphosphate (PIP2) and phosphatidylcholine (PC). Synthesis of PI4P requires GBF1 and Arf1. We identified 3CD derivatives: 3CDm and 3CmD, that we used to show that distinct domains of 3CD function upstream of GBF1 and downstream of Arf1 activation. These same 3CD derivatives still supported induction of PIP2 and PC, suggesting that pathways and corresponding mechanisms used to induce these phospholipids are distinct. Phospholipid induction by 3CD is localized to the perinuclear region of the cell, the outcome of which is the proliferation of membranes in this area of the cell. We conclude that a single viral protein can serve as a master regulator of cellular phospholipid and membrane biogenesis, likely by commandeering normal cellular pathways.

Translation control of Enterovirus A71 gene expression.

Upon EV-A71 infection of a host cell, EV-A71 RNA is translated into a viral polyprotein. Although EV-A71 can use the cellular translation machinery to produce viral proteins, unlike cellular translation, which is cap-dependent, the viral RNA genome of EV-A71 does not contain a 5' cap and the translation of EV-A71 protein is cap-independent, which is mediated by the internal ribosomal entry site (IRES) located in the 5' UTR of EV-A71 mRNA. Like many other eukaryotic viruses, EV-A71 manipulates the host cell translation devices, using an elegant RNA-centric strategy in infected cells. During viral translation, viral RNA plays an important role in controlling the stage of protein synthesis. In addition, due to the cellular defense mechanism, viral replication is limited by down-regulating translation. EV-A71 also utilizes protein factors in the host to overcome antiviral responses or even use them to promote viral translation rather than host cell translation. In this review, we provide an introduction to the known strategies for EV-A71 to exploit cellular translation mechanisms.

MicroRNA 876-5p modulates EV-A71 replication through downregulation of host antiviral factors.

BACKGROUND: Human enterovirus 71 (EV-A71) is a non-enveloped virus that has a single stranded positive sense RNA genome. In a previous study, we showed that miR-876-5p upregulation was observed in the serum of patients with severe EV-A71 infection. Micro-876-5p (miR-876-5p) is a circulating miRNA that can be identified to modulate EV-A71 infections through both in vitro and in vivo studies. However, the regulatory mechanisms that involve miR-876-5p in the EV-A71 infection cycle remain unclear. METHODS: We demonstrated that miR-876-5p facilitated EV-A71 replication and expression by overexpression and knocking-down of miR-876-5p through the transfection of miR-876-5p plasmid and miR-876-5p inhibitor. Although miR-876-5p suppressed CREB5 expression, luciferase reporter assay confirmed this. We also evaluated the role of miR-876-5p in the EV-A71 infection cycle by CREB5 mediated by transfection with an anti-miR-876-5P inhibitor or in combination with an si-CREB5 plasmid. RESULTS: MicroR-876-5p was upregulated in EV-A71-infected neuroblastoma cells. Overexpression of miR-876-5p or knockdown of cyclic-AMP responsive element binding protein 5 (CREB5) promoted EV-A71 replication. The downregulation of miR-876-5p inhibited the accumulation of viral RNA and the production of viral proteins. Interestingly, CREB5 overexpression also suppressed EV-A71 replication. Our in vitro studies reveal that miR-876-5p directly targets CREB5. Finally, downregulation of CREB5 protein abated the inhibitory effect of anti-miR-876-5p and induced inhibitory effect of EV-A71 replication. CONCLUSIONS: Our results suggest that intracellular miR-876-5p promotes EV-A71 replication indirectly by targeting the host CREB5 protein.

High Amplification of the Antiviral Activity of Curcumin through Transformation into Carbon Quantum Dots.

It is demonstrated that carbon quantum dots derived from curcumin (Cur-CQDs) through one-step dry heating are effective antiviral agents against enterovirus 71 (EV71). The surface properties of Cur-CQDs, as well as their antiviral activity, are highly dependent on the heating temperature during synthesis. The one-step heating of curcumin at 180 °C preserves many of the moieties of polymeric curcumin on the surfaces of the as-synthesized Cur-CQDs, resulting in superior antiviral characteristics. It is proposed that curcumin undergoes a series of structural changes through dehydration, polymerization, and carbonization to form core-shell CQDs whose surfaces remain a pyrolytic curcumin-like polymer, boosting the antiviral activity. The results reveal that curcumin possesses insignificant inhibitory activity against EV71 infection in RD cells [half-maximal effective concentration (EC50 ) >200 µg mL-1 ] but exhibits high cytotoxicity toward RD cells (half-maximal cytotoxic concentration (CC50 ) <13 µg mL-1 ). The EC50 (0.2 µg mL-1 ) and CC50 (452.2 µg mL-1 ) of Cur-CQDs are >1000-fold lower and >34-fold higher, respectively, than those of curcumin, demonstrating their far superior antiviral capabilities and high biocompatibility. In vivo, intraperitoneal administration of Cur-CQDs significantly decreases mortality and provides protection against virus-induced hind-limb paralysis in new-born mice challenged with a lethal dose of EV71.

Cytotoxicity of 11-epi-Sinulariolide Acetate Isolated from Cultured Soft Corals on HA22T Cells through the Endoplasmic Reticulum Stress Pathway and Mitochondrial Dysfunction.

Natural compounds from soft corals have been increasingly used for their antitumor therapeutic properties. This study examined 11-epi-sinulariolide acetate (11-epi-SA), an active compound isolated from the cultured soft coral Sinularia flexibilis, to determine its potential antitumor effect on four hepatocellular carcinoma cell lines. Cell viability was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the results demonstrated that 11-epi-SA treatment showed more cytotoxic effect toward HA22T cells. Protein profiling of the 11-epi-SA-treated HA22T cells revealed substantial protein alterations associated with stress response and protein synthesis and folding, suggesting that the mitochondria and endoplasmic reticulum (ER) play roles in 11-epi-SA-initiated apoptosis. Moreover, 11-epi-SA activated caspase-dependent apoptotic cell death, suggesting that mitochondria-related apoptosis genes were involved in programmed cell death. The unfolded protein response signaling pathway-related proteins were also activated on 11-epi-SA treatment, and these changes were accompanied by the upregulated expression of growth arrest and DNA damage-inducible protein (GADD153) and CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP), the genes encoding transcription factors associated with growth arrest and apoptosis under prolonged ER stress. Two inhibitors, namely salubrinal (Sal) and SP600125, partially abrogated 11-epi-SA-related cell death, implying that the protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK)-activating transcription factor (ATF) 6-CHOP or the inositol-requiring enzyme 1 alpha (IRE1α)-c-Jun N-terminal kinase (JNK)-cJun signal pathway was activated after 11-epi-SA treatment. In general, these results suggest that 11-epi-SA exerts cytotoxic effects on HA22T cells through mitochondrial dysfunction and ER stress cell death pathways.

13-acetoxysarcocrassolide induces apoptosis on human gastric carcinoma cells through mitochondria-related apoptotic pathways: p38/JNK activation and PI3K/AKT suppression.

13-acetoxysarcocrassolide (13-AC), an active compound isolated from cultured Formosa soft coral Sarcophyton crassocaule, was found to possess anti-proliferative and apoptosis-inducing activities against AGS (human gastric adenocarcinoma cells) gastric carcinoma cells. The anti-tumor effects of 13-AC were determined by MTT assay, colony formation assessment, cell wound-healing assay, TUNEL/4,6-Diamidino-2-phenylindole (DAPI) staining, Annexin V-fluorescein isothiocyanate/propidium iodide (PI) staining and flow cytometry. 13-AC inhibited the growth and migration of gastric carcinoma cells in a dose-dependent manner and induced both early and late apoptosis as assessed by flow cytometer analysis. 13-AC-induced apoptosis was confirmed through observation of a change in ΔΨm, up-regulated expression levels of Bax and Bad proteins, down-regulated expression levels of Bcl-2, Bcl-xl and Mcl-1 proteins, and the activation of caspase-3, caspase-9, p38 and JNK. Furthermore, inhibition of p38 and JNK activity by pretreatment with SB03580 (a p38-specific inhibitor) and SP600125 (a JNK-specific inhibitor) led to rescue of the cell cytotoxicity of 13-AC-treated AGS cells, indicating that the p38 and the JNK pathways are also involved in the 13-AC-induced cell apoptosis. Together, these results suggest that 13-AC induces cell apoptosis against gastric cancer cells through triggering of the mitochondrial-dependent apoptotic pathway as well as activation of the p38 and JNK pathways.

Human foreskin fibroblast-like stromal cells can differentiate into functional hepatocytic cells.

Foreskin fibroblast-like stromal cells (FDSCs) are progenitors isolated from human tissue that can differentiate into diverse cell types. Many types of stem cells can differentiate into hepatocyte-like cells, which could be used for drug testing or in liver regeneration therapy, but whether FDSCs can be converted into functional hepatocytes is unknown. FDSCs show divergent properties when cultured in distinct media, forming spheres in Dulbecco's modified Eagle's medium (DMEM) containing F12, epidermal growth factor (EGF), and basic fibroblast growth factor (b-FGF), but have fibroblast-like morphology when cultured in DMEM-based growth medium. Both cell populations express the typical mesenchymal stem cell markers CD90, CD105, and CD73, but the p75 neurotrophin receptor (p75NTR) was detected only in FDSC spheres. Both types of FDSCs can differentiate into hepatocyte-like cells, which express typical liver markers, including albumin and hepatocyte paraffin 1 (Hep Par1), along with liver-specific biological activities. When plasmids containing the human hepatitis B virus (HBV) genome were transfected transiently into FDSCs, differentiated hepatocyte-like cells secrete large amounts of HBe and HBs antigens. FDSCs could be used for clinical hepatic therapy and/or serve as a model of HBV.

Application of Shear Horizontal Surface Acoustic Wave (SH-SAW) Immunosensor in Point-of-Care Diagnosis.

Point-of-care testing (POCT), also known as on-site or near-patient testing, has been exploding in the last 20 years. A favorable POCT device requires minimal sample handling (e.g., finger-prick samples, but plasma for analysis), minimal sample volume (e.g., one drop of blood), and very fast results. Shear horizontal surface acoustic wave (SH-SAW) biosensors have attracted a lot of attention as one of the effective solutions to complete whole blood measurements in less than 3 min, while providing a low-cost and small-sized device. This review provides an overview of the SH-SAW biosensor system that has been successfully commercialized for medical use. Three unique features of the system are a disposable test cartridge with an SH-SAW sensor chip, a mass-produced bio-coating, and a palm-sized reader. This paper first discusses the characteristics and performance of the SH-SAW sensor system. Subsequently, the method of cross-linking biomaterials and the analysis of SH-SAW real-time signals are investigated, and the detection range and detection limit are presented.

The Upf1 protein restricts EV-A71 viral replication.

Enterovirus A71 (EV-A71) is transmitted through the respiratory tract, gastrointestinal system, and fecal-oral routes. The main symptoms caused by EV-A71 are hand, foot, and mouth disease (HFMD) or vesicular sore throat. Upf1 (Up-frameshift protein 1) was reported to degrade mRNA containing early stop codons, known as nonsense-mediated decay (NMD). Upf1 is also involved in the NMD mechanism as a host factor detrimental to viral replication. In this study, we dissected the potential roles of Upf1 in the EV-A71-infected cells. Upf1 was virulently down-regulated in three different EV-A71-infected cells, RD, Hela, and 293T, implying that Upf1 is a host protein unfavorable for EV-A71 replication. Knockdown of Upf1 protein resulted in increased viral RNA expression and production of progeny virus, and conversely, overexpression of Upf1 protein resulted in decreased viral RNA expression and production of progeny virus. Importantly, we observed increased RNA levels of asparagine synthetase (ASNS), one of the indicator substrates for the NMD mechanism, which indirectly suggests that EV-A71 infection of cells suppresses NMD activity in the host. The results shown in this study are useful for subsequent analysis of the relationship between the NMD/Upf1 mechanism and other picornaviruses, which may lead to the development of anti-picornavirus drugs.

Induction of apoptosis by sinulariolide from soft coral through mitochondrial-related and p38MAPK pathways on human bladder carcinoma cells.

Sinulariolide, an isolated compound from the soft coral Sinularia flexibilis, possesses the anti-proliferative, anti-migratory and apoptosis-inducing activities against the TSGH bladder carcinoma cell. The anti-tumor effects of sinulariolide were determined by 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, cell migration assay and flow cytometry, respectively. Sinulariolide inhibited the growth and migration of bladder carcinoma cells in a dose-dependent manner, as well as induced both early and late apoptosis as determined by the flow cytometer. Also, the sinulariolide-induced apoptosis is related to the mitochondrial-mediated apoptosis via caspase-dependent pathways, elucidated by the loss of mitochondrial membrane potential, release of cytochrome C, activation of caspase-3/-9, Bax and Bad, as well as suppression of Bcl-2/Bcl-xL/Mcl-1. Detection of the PARP-1 cleaved product suggested the partial involvement of caspase-independent pathways. Moreover, inhibition of p38MAPK activity leads to the rescue of the cell cytotoxicity of sinulariolide-treated TSGH cells, indicating that the p38MAPK pathway is also involved in the sinulariolide-induced cell apoptosis. Altogether, these results suggest that sinulariolide induces apoptosis against bladder cancer cells through mitochondrial-related and p38MAPK pathways.

Measurements of Anti-SARS-CoV-2 Antibody Levels after Vaccination Using a SH-SAW Biosensor.

To prevent the COVID-19 pandemic that threatens human health, vaccination has become a useful and necessary tool in the response to the pandemic. The vaccine not only induces antibodies in the body, but may also cause adverse effects such as fatigue, muscle pain, blood clots, and myocarditis, especially in patients with chronic disease. To reduce unnecessary vaccinations, it is becoming increasingly important to monitor the amount of anti-SARS-CoV-2 S protein antibodies prior to vaccination. A novel SH-SAW biosensor, coated with SARS-CoV-2 spike protein, can help quantify the amount of anti-SARS-CoV-2 S protein antibodies with 5 µL of finger blood within 40 s. The LoD of the spike-protein-coated SAW biosensor was determined to be 41.91 BAU/mL, and the cut-off point was determined to be 50 BAU/mL (Youden's J statistic = 0.94733). By using the SH-SAW biosensor, we found that the total anti-SARS-CoV-2 S protein antibody concentrations spiked 10−14 days after the first vaccination (p = 0.0002) and 7−9 days after the second vaccination (p = 0.0116). Furthermore, mRNA vaccines, such as Moderna or BNT, could achieve higher concentrations of total anti-SARS-CoV-2 S protein antibodies compared with adenovirus vaccine, AZ (p < 0.0001). SH-SAW sensors in vitro diagnostic systems are a simple and powerful technology to investigate the local prevalence of COVID-19.

Identification of heat-shock protein 90 beta in Japanese encephalitis virus-induced secretion proteins.

Five host cellular proteins were identified in the secretion medium from Japanese encephalitis virus (JEV)-infected baby hamster kidney-21 (BHK-21) cells, including three molecular chaperones: Hsp70, GRP78 and Hsp90. Hsp90 isoforms were characterized further. Hsp90α was observed to be retained inside the nuclei, whereas Hsp90ß associated with virus particles during assembly and was released into the secretion medium upon JEV infection. The association of Hsp90ß and viral E protein was demonstrated by using sucrose-density fractionation and Western blot analysis. Moreover, JEV viral RNA replication was not affected by treatment with geldanamycin, an Hsp90 inhibitor, but impaired virus infectivity that was determined by a plaque-forming assay. Our results show that Hsp90ß, not Hsp90α, is present in the JEV-induced secretion medium and is required for JEV infectivity in BHK-21 cells.

A Novel Rapid Test to Detect Anti-SARS-CoV-2 N Protein IgG Based on Shear Horizontal Surface Acoustic Wave (SH-SAW).

Since the Coronavirus disease 2019 (COVID-19) pandemic outbreak, many methods have been used to detect antigens or antibodies to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), including viral culture, nucleic acid test, and immunoassay. The shear-horizontal surface acoustic wave (SH-SAW) biosensor is a novel pathogen detection platform with the advantages of high sensitivity and short detection time. The objective of this study is to develop a SH-SAW biosensor to detect the anti-SARS-CoV-2 nucleocapsid antibody. The rabbit sera collected from rabbits on different days after SARS-CoV-2 N protein injection were evaluated by SH-SAW biosensor and enzyme-linked immunosorbent assay (ELISA). The results showed that the SH-SAW biosensor achieved a high correlation coefficient (R = 0.9997) with different concentrations (34.375-1100 ng/mL) of the "spike-in" anti-N protein antibodies. Compared to ELISA, the SH-SAW biosensor has better sensitivity and can detect anti-N protein IgG signals earlier than ELISA on day 6 (p < 0.05). Overall, in this study, we demonstrated that the SH-SAW biosensor is a promising platform for rapid in vitro diagnostic (IVD) testing, especially for antigen or antibody testing.

High Incidence and Early Onset of Urinary Tract Cancers in Patients with BK Polyomavirus Associated Nephropathy.

Over-immunosuppressed kidney transplant recipients are susceptible to malignancies and BK polyomavirus (BKPyV)-associated nephropathy (BKPyVAN). This study aimed to verify the association between BKPyV infection and urinary tract cancers (UTC). A total of 244 kidney transplant recipients were enrolled at Chang Gung Memorial Hospital from June 2000 to February 2020. Biopsy-proven BKPyVAN patients (n = 17) had worse kidney function (eGFR: 26 ± 13.7 vs. 47.8 ± 31.0 mL/min/1.73 m2). The 5-year allograft survival rates for patients with and without BKPyVAN were 67% and 93%, respectively (p = 0.0002), while the 10-year patient survival was not different between the two groups. BKPyVAN patients had a significantly higher incidence of UTC compared to the non-BKPyVAN group (29.4% vs. 6.6%). Kaplan-Meier analysis showed that the UTC-free survival rate was significantly lower in BKPyVAN patients, and the onset of UTC was significantly shorter in BKPyVAN patients (53.4 vs. 108.9 months). The multivariate logistic regression analysis demonstrated that age (RR = 1.062) and BKVAN (RR = 6.459) were the most significant risk factors for the development of UTC. Our study demonstrates that BKPyVAN patients have greater allograft losses, higher incidence, a lower cancer-free survival rate, and an earlier onset with a higher relative risk of developing UTC compared to non-BKPyVAN patients.

Evaluation of ViroTrack Sero Zika IgG/IgM, a New Rapid and Quantitative Zika Serological Diagnostic Assay.

Dengue virus (DENV) and Zika virus (ZIKV) belong to the flavivirus genus and are antigenically closely related. They also share the same mosquito vector and can cause similar symptoms upon infection. However, DENV and ZIKV infections lead to different clinical sequelae and treatments; therefore, clinicians need rapid and accurate diagnostics capable of distinguishing between the two diseases. METHODS: We employed the immuno-magnetic assay technology on a microfluidic cartridge (ViroTrack Sero Zika IgG/IgM) for diagnosis of ZIKV infection based on the aggregation of magnetic nanoparticles. We carried out three serological studies including samples from the Dominican Republic, USA, and Nicaragua, aimed at detecting ZIKV-specific IgG and IgM using the ViroTrack Sero Zika IgG/IgM test. RESULTS: The seroconversion results were comparable with ZIKV IgG and IgM reactivity measured by the commercial ZIKV ELISA kit. The sensitivity and specificity for both ZIKV IgG and IgM tested by the ViroTrack Sero Zika IgG/IgM was approximately 98% and 93%, respectively. CONCLUSION: Serological detection of ZIKV infection by the new ViroTrack Sero Zika IgG/IgM test shows promising performance and limited cross-reactivity with DENV.