A rapid, high-throughput, viral infectivity assay using automated brightfield microscopy with machine learning.

Infectivity assays are essential for the development of viral vaccines, antiviral therapies, and the manufacture of biologicals. Traditionally, these assays take 2-7 days and require several manual processing steps after infection. We describe an automated viral infectivity assay (AVIATM), using convolutional neural networks (CNNs) and high-throughput brightfield microscopy on 96-well plates that can quantify infection phenotypes within hours, before they are manually visible, and without sample preparation. CNN models were trained on HIV, influenza A virus, coronavirus 229E, vaccinia viruses, poliovirus, and adenoviruses, which together span the four major categories of virus (DNA, RNA, enveloped, and non-enveloped). A sigmoidal function, fit between virus dilution curves and CNN predictions, results in sensitivity ranges comparable to or better than conventional plaque or TCID50 assays, and a precision of ∼10%, which is considerably better than conventional infectivity assays. Because this technology is based on sensitizing CNNs to specific phenotypes of infection, it has potential as a rapid, broad-spectrum tool for virus characterization, and potentially identification.

Color-variable dual-dyed photodynamic antimicrobial polyethylene terephthalate (PET)/cotton blended fabrics.

The urgent demand for scalable, potent, color variable, and comfortable antimicrobial textiles as personal protection equipment (PPE) to help reduce infection transmission in hospitals and healthcare facilities has significantly increased since the start of the COVID-19 pandemic. Here, we explored photodynamic antimicrobial polyethylene terephthalate/cotton (TC) blended fabrics comprised of photosensitizer-conjugated cotton fibers and polyethylene terephthalate (PET) fibers dyed with disperse dyes. A small library of TC blended fabrics was constructed wherein the PET fibers were embedded with traditional disperse dyes dominating the fabric color, thereby enabling variable color expression, while the cotton fibers were covalently coupled with the photosensitizer thionine acetate as the microbicidal agent. Physical (SEM, CLSM, TGA, XPS and mechanical strength) and colorimetric (K/S and CIELab values) characterization methods were employed to investigate the resultant fabrics, and photooxidation studies with DPBF demonstrated the ability of these materials to generate reactive oxygen species (i.e., singlet oxygen) upon visible light illumination. The best results demonstrated a photodynamic inactivation of 99.985% (~ 3.82 log unit reduction, P = 0.0021) against Gram-positive S. aureus, and detection limit inactivation (99.99%, 4 log unit reduction, P ≤ 0.0001) against Gram-negative E. coli upon illumination with visible light (60 min; ~ 300 mW/cm2; λ ≥ 420 nm). Enveloped human coronavirus 229E showed a photodynamic susceptibility of ~ 99.99% inactivation after 60 min illumination (400-700 nm, 65 ± 5 mW/cm2). The presence of the disperse dyes on the fabrics showed no significant effects on the aPDI results, and furthermore, appeared to provide the photosensitizer with some measure of protection from photobleaching, thus improving the photostability of the dual-dyed fabrics. Taken together, these results suggest the feasibility of low cost, scalable and color variable thionine-conjugated TC blended fabrics as potent self-disinfecting textiles.

Effect of BIO-PLYTM, a Platelet-Rich Plasma Derived Biologic on PRRSV-2-Infected Macrophages.

Porcine Reproductive and Respiratory Syndrome (PRRS) is the one of the most devastating diseases impacting the swine industry worldwide. Control and prevention methods rely on biosafety measures and vaccination. As an RNA virus with a high rate of mutation, vaccines are only partially effective against circulating and newly emerging strains. To reduce the burden of this disease, research on alternative control methods is needed. Here, we assess the in vitro antiviral effect of a novel platelet-rich plasma-derived biologic termed BIO-PLYTM (for the BIOactive fraction of Platelet-rich plasma LYsate) from both swine and equine origin. Our results show that BIO-PLYTM significantly reduces the amount of PRRSV viral load determined by RT-qPCR and the number of infectious viral particles measured by TCID50 in infected porcine alveolar and parenchymal macrophages. This study also showed limited toxicity of BIO-PLYTM in vitro and aspects of its immunomodulatory capacity evaluating the regulation of reactive oxygen species and cytokines production in infected cells. Finally, this study presents promising data on the effect of BIO-PLYTM on other RNA viruses such as human A influenza viruses and coronavirus.

On-demand, remote and lossless manipulation of biofluid droplets.

The recent global outbreaks of epidemics and pandemics have shown us that we are severely under-prepared to cope with infectious agents. Exposure to infectious agents present in biofluids (e.g., blood, saliva, urine etc.) poses a severe risk to clinical laboratory personnel and healthcare workers, resulting in hundreds of millions of hospital-acquired and laboratory-acquired infections annually. Novel technologies that can minimize human exposure through remote and automated handling of infectious biofluids will mitigate such risk. In this work, we present biofluid manipulators, which allow on-demand, remote and lossless manipulation of virtually any liquid droplet. Our manipulators are designed by integrating thermo-responsive soft actuators with superomniphobic surfaces. Utilizing our manipulators, we demonstrate on-demand, remote and lossless manipulation of biofluid droplets. We envision that our biofluid manipulators will not only reduce manual operations and minimize exposure to infectious agents, but also pave the way for developing inexpensive, simple and portable robotic systems, which can allow point-of-care operations, particularly in developing nations.

Vandetanib Blocks the Cytokine Storm in SARS-CoV-2-Infected Mice.

The portfolio of SARS-CoV-2 small molecule drugs is currently limited to a handful that are either approved (remdesivir), emergency approved (dexamethasone, baricitinib, paxlovid, and molnupiravir), or in advanced clinical trials. Vandetanib is a kinase inhibitor which targets the vascular endothelial growth factor receptor (VEGFR), the epidermal growth factor receptor (EGFR), as well as the RET-tyrosine kinase. In the current study, it was tested in different cell lines and showed promising results on inhibition versus the toxic effect on A549-hACE2 cells (IC50 0.79 µM) while also showing a reduction of >3 log TCID50/mL for HCoV-229E. The in vivo efficacy of vandetanib was assessed in a mouse model of SARS-CoV-2 infection and statistically significantly reduced the levels of IL-6, IL-10, and TNF-α and mitigated inflammatory cell infiltrates in the lungs of infected animals but did not reduce viral load. Vandetanib also decreased CCL2, CCL3, and CCL4 compared to the infected animals. Vandetanib additionally rescued the decreased IFN-1ß caused by SARS-CoV-2 infection in mice to levels similar to that in uninfected animals. Our results indicate that the FDA-approved anticancer drug vandetanib is worthy of further assessment as a potential therapeutic candidate to block the COVID-19 cytokine storm.

Flavonols and dihydroflavonols inhibit the main protease activity of SARS-CoV-2 and the replication of human coronavirus 229E.

Since December 2019, the deadly novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused the current COVID-19 pandemic. To date, vaccines are available in the developed countries to prevent the infection of this virus; however, medicines are necessary to help control COVID-19. Human coronavirus 229E (HCoV-229E) causes the common cold. The main protease (Mpro) is an essential enzyme required for the multiplication of these two viruses in the host cells, and thus is an appropriate candidate to screen potential medicinal compounds. Flavonols and dihydroflavonols are two groups of plant flavonoids. In this study, we report docking simulation with two Mpro enzymes and five flavonols and three dihydroflavonols, in vitro inhibition of the SARS-CoV-2 Mpro, and in vitro inhibition of the HCoV 229E replication. The docking simulation results predicted that (+)-dihydrokaempferol, (+)- dihydroquercetin, (+)-dihydromyricetin, kaempferol, quercetin, myricentin, isoquercitrin, and rutin could bind to at least two subsites (S1, S1', S2, and S4) in the binding pocket and inhibit the activity of SARS-CoV-2 Mpro. Their affinity scores ranged from -8.8 to -7.4 (kcal/mol). Likewise, these compounds were predicted to bind and inhibit the HCoV-229E Mpro activity with affinity scores ranging from -7.1 to -7.8 (kcal/mol). In vitro inhibition assays showed that seven available compounds effectively inhibited the SARS-CoV-2 Mpro activity and their IC50 values ranged from 0.125 to 12.9 µM. Five compounds inhibited the replication of HCoV-229E in Huh-7 cells. These findings indicate that these antioxidative flavonols and dihydroflavonols are promising candidates for curbing the two viruses.

Chalcones from Angelica keiskei (ashitaba) inhibit key Zika virus replication proteins.

Zika virus (ZIKV) is a dangerous human pathogen and no antiviral drugs have been approved to date. The chalcones are a group of small molecules that are found in a number of different plants, including Angelica keiskei Koidzumi, also known as ashitaba. To examine chalcone anti-ZIKV activity, three chalcones, 4-hydroxyderricin (4HD), xanthoangelol (XA), and xanthoangelol-E (XA-E), were purified from a methanol-ethyl acetate extract from A. keiskei. Molecular and ensemble docking predicted that these chalcones would establish multiple interactions with residues in the catalytic and allosteric sites of ZIKV NS2B-NS3 protease, and in the allosteric site of the NS5 RNA-dependent RNA-polymerase (RdRp). Machine learning models also predicted 4HD, XA and XA-E as potential anti-ZIKV inhibitors. Enzymatic and kinetic assays confirmed chalcone inhibition of the ZIKV NS2B-NS3 protease allosteric site with IC50s from 18 to 50 µM. Activity assays also revealed that XA, but not 4HD or XA-E, inhibited the allosteric site of the RdRp, with an IC50 of 6.9 µM. Finally, we tested these chalcones for their anti-viral activity in vitro with Vero cells. 4HD and XA-E displayed anti-ZIKV activity with EC50 values of 6.6 and 22.0 µM, respectively, while XA displayed relatively weak anti-ZIKV activity with whole cells. With their simple structures and relative ease of modification, the chalcones represent attractive candidates for hit-to-lead optimization in the search of new anti-ZIKV therapeutics.

A Liquid Metal Mediated Metallic Coating for Antimicrobial and Antiviral Fabrics.

Fabrics are widely used in hospitals and many other settings for bedding, clothing, and face masks; however, microbial pathogens can survive on surfaces for a long time, leading to microbial transmission. Coatings of metallic particles on fabrics have been widely used to eradicate pathogens. However, current metal particle coating technologies encounter numerous issues such as nonuniformity, processing complexity, and poor adhesion. To overcome these issues, an easy-to-control and straightforward method is reported to coat a wide range of fabrics by using gallium liquid metal (LM) particles to facilitate the deposition of liquid metal copper alloy (LMCu) particles. Gallium particles coated on the fabric provide nucleation sites for forming LMCu particles at room temperature via galvanic replacement of Cu2+ ions. The LM helps promote strong adhesion of the particles to the fabric. The presence of the LMCu particles can eradicate over 99% of pathogens (including bacteria, fungi, and viruses) within 5 min, which is significantly more effective than control samples coated with only Cu. The coating remains effective over multiple usages and against contaminated droplets and aerosols, such as those encountered in facemasks. This facile coating method is promising for generating robust antibacterial, antifungal, and antiviral fabrics and surfaces.

Toward Universal Photodynamic Coatings for Infection Control.

The dual threats posed by the COVID-19 pandemic and hospital-acquired infections (HAIs) have emphasized the urgent need for self-disinfecting materials for infection control. Despite their highly potent antimicrobial activity, the adoption of photoactive materials to reduce infection transmission in hospitals and related healthcare facilities has been severely hampered by the lack of scalable and cost-effective manufacturing, in which case high-volume production methods for fabricating aPDI-based materials are needed. To address this issue here, we examined the antimicrobial efficacy of a simple bicomponent spray coating composed of the commercially-available UV-photocrosslinkable polymer N-methyl-4(4'-formyl-styryl)pyridinium methosulfate acetal poly(vinyl alcohol) (SbQ-PVA) and one of three aPDI photosensitizers (PSs): zinc-tetra(4-N-methylpyridyl)porphine (ZnTMPyP4+), methylene blue (MB), and Rose Bengal (RB). We applied these photodynamic coatings, collectively termed SbQ-PVA/PS, to a variety of commercially available materials. Scanning electron microscopy (SEM) and time-of-flight secondary ion mass spectrometry (ToF-SIMS) confirmed the successful application of the coatings, while inductively coupled plasma-optical emission spectroscopy (ICP-OES) revealed a photosensitizer loading of 0.09-0.78 nmol PS/mg material. The antimicrobial efficacy of the coated materials was evaluated against methicillin-susceptible Staphylococcus aureus ATCC-29213 and human coronavirus strain HCoV-229E. Upon illumination with visible light (60 min, 400-700 nm, 65 ± 5 mW/cm2), the coated materials inactivated S. aureus by 97-99.999% and HCoV-229E by 92-99.999%, depending on the material and PS employed. Photobleaching studies employing HCoV-229E demonstrated detection limit inactivation (99.999%) even after exposure for 4 weeks to indoor ambient room lighting. Taken together, these results demonstrate the potential for photodynamic SbQ-PVA/PS coatings to be universally applied to a wide range of materials for effectively reducing pathogen transmission.

Rapid and Repetitive Inactivation of SARS-CoV-2 and Human Coronavirus on Self-Disinfecting Anionic Polymers.

While the ongoing COVID-19 pandemic affirms an urgent global need for effective vaccines as second and third infection waves are spreading worldwide and generating new mutant virus strains, it has also revealed the importance of mitigating the transmission of SARS-CoV-2 through the introduction of restrictive social practices. Here, it is demonstrated that an architecturally- and chemically-diverse family of nanostructured anionic polymers yield a rapid and continuous disinfecting alternative to inactivate coronaviruses and prevent their transmission from contact with contaminated surfaces. Operating on a dramatic pH-drop mechanism along the polymer/pathogen interface, polymers of this archetype inactivate the SARS-CoV-2 virus, as well as a human coronavirus surrogate (HCoV-229E), to the minimum detection limit within minutes. Application of these anionic polymers to frequently touched surfaces in medical, educational, and public-transportation facilities, or personal protection equipment, can provide rapid and repetitive protection without detrimental health or environmental complications.

Repurposing the Ebola and Marburg Virus Inhibitors Tilorone, Quinacrine, and Pyronaridine: In Vitro Activity against SARS-CoV-2 and Potential Mechanisms.

Severe acute respiratory coronavirus 2 (SARS-CoV-2) is a newly identified virus that has resulted in over 2.5 million deaths globally and over 116 million cases globally in March, 2021. Small-molecule inhibitors that reverse disease severity have proven difficult to discover. One of the key approaches that has been widely applied in an effort to speed up the translation of drugs is drug repurposing. A few drugs have shown in vitro activity against Ebola viruses and demonstrated activity against SARS-CoV-2 in vivo. Most notably, the RNA polymerase targeting remdesivir demonstrated activity in vitro and efficacy in the early stage of the disease in humans. Testing other small-molecule drugs that are active against Ebola viruses (EBOVs) would appear a reasonable strategy to evaluate their potential for SARS-CoV-2. We have previously repurposed pyronaridine, tilorone, and quinacrine (from malaria, influenza, and antiprotozoal uses, respectively) as inhibitors of Ebola and Marburg viruses in vitro in HeLa cells and mouse-adapted EBOV in mice in vivo. We have now tested these three drugs in various cell lines (VeroE6, Vero76, Caco-2, Calu-3, A549-ACE2, HUH-7, and monocytes) infected with SARS-CoV-2 as well as other viruses (including MHV and HCoV 229E). The compilation of these results indicated considerable variability in antiviral activity observed across cell lines. We found that tilorone and pyronaridine inhibited the virus replication in A549-ACE2 cells with IC50 values of 180 nM and IC50 198 nM, respectively. We used microscale thermophoresis to test the binding of these molecules to the spike protein, and tilorone and pyronaridine bind to the spike receptor binding domain protein with K d values of 339 and 647 nM, respectively. Human Cmax for pyronaridine and quinacrine is greater than the IC50 observed in A549-ACE2 cells. We also provide novel insights into the mechanism of these compounds which is likely lysosomotropic.

Impact of lung morphology on clinical outcomes with riociguat in patients with pulmonary hypertension and idiopathic interstitial pneumonia: A post hoc subgroup analysis of the RISE-IIP study.

BACKGROUND: Riociguat in Patients with Symptomatic Pulmonary Hypertension associated with Idiopathic Interstitial Pneumonias (RISE-IIP), a randomized, controlled, phase 2b trial of riociguat for pulmonary hypertension associated with idiopathic interstitial pneumonia, was terminated early due to increased mortality in riociguat-treated patients. Baseline characteristics of enrolled patients demonstrated a low diffusing capacity of the lung for carbon monoxide (DLCO) with preserved lung volumes at baseline, suggesting the presence of combined pulmonary fibrosis and emphysema (CPFE) in some patients. This post hoc analysis of RISE-IIP was undertaken to explore lung morphology, assessed by high-resolution computed tomography, and associated clinical outcomes. METHODS: Available baseline/pre-baseline high-resolution computed tomography scans were reviewed centrally by 2 radiologists. The extent of emphysema and fibrosis was retrospectively scored and combined to provide the total CPFE score. RESULTS: Data were available for 65/147 patients (44%), including 15/27 fatal cases (56%). Of these, 41/65 patients (63%) had CPFE. Mortality was higher in patients with CPFE (12/41; 29%) than those without (3/24; 13%). Fourteen patients with CPFE had emphysema > fibrosis (4 died). No relationship was observed between CPFE score, survival status, and treatment assignment. A low DLCO, short 6-min walking distance, and high forced vital capacity:DLCO ratio at baseline also appeared to be risk factors for mortality. CONCLUSIONS: High parenchymal lung disease burden and the presence of more emphysema than fibrosis might have predisposed patients with pulmonary hypertension associated with idiopathic interstitial pneumonia to poor outcomes in RISE-IIP. Future studies of therapy for group 3 pulmonary hypertension should include centrally adjudicated imaging for morphologic phenotyping and disease burden evaluation during screening.

Vandetanib Reduces Inflammatory Cytokines and Ameliorates COVID-19 in Infected Mice.

The portfolio of SARS-CoV-2 small molecule drugs is currently limited to a handful that are either approved (remdesivir), emergency approved (dexamethasone, baricitinib) or in advanced clinical trials. We have tested 45 FDA-approved kinase inhibitors in vitro against murine hepatitis virus (MHV) as a model of SARS-CoV-2 replication and identified 12 showing inhibition in the delayed brain tumor (DBT) cell line. Vandetanib, which targets the vascular endothelial growth factor receptor (VEGFR), the epidermal growth factor receptor (EGFR), and the RET-tyrosine kinase showed the most promising results on inhibition versus toxic effect on SARS-CoV-2-infected Caco-2 and A549-hACE2 cells (IC50 0.79 µM) while also showing a reduction of > 3 log TCID50/mL for HCoV-229E. The in vivo efficacy of vandetanib was assessed in a mouse model of SARS-CoV-2 infection and statistically significantly reduced the levels of IL-6, IL-10, TNF-α, and mitigated inflammatory cell infiltrates in the lungs of infected animals but did not reduce viral load. Vandetanib rescued the decreased IFN-1ß caused by SARS-CoV-2 infection in mice to levels similar to that in uninfected animals. Our results indicate that the FDA-approved vandetanib is a potential therapeutic candidate for COVID-19 positioned for follow up in clinical trials either alone or in combination with other drugs to address the cytokine storm associated with this viral infection.

Photodynamic Coatings on Polymer Microfibers for Pathogen Inactivation: Effects of Application Method and Composition.

A substantial increase in the risk of hospital-acquired infections (HAIs) has greatly impacted the global healthcare industry. Harmful pathogens adhere to a variety of surfaces and infect personnel on contact, thereby promoting transmission to new hosts. This is particularly worrisome in the case of antibiotic-resistant pathogens, which constitute a growing threat to human health worldwide and require new preventative routes of disinfection. In this study, we have incorporated different loading levels of a porphyrin photosensitizer capable of generating reactive singlet oxygen in the presence of O2 and visible light in a water-soluble, photo-cross-linkable polymer coating, which was subsequently deposited on polymer microfibers. Two different application methods are considered, and the morphological and chemical characteristics of these coated fibers are analyzed to detect the presence of the coating and photosensitizer. To discern the efficacy of the fibers against pathogenic bacteria, photodynamic inactivation has been performed on two different bacterial strains, Staphylococcus aureus and antibiotic-resistant Escherichia coli, with population reductions of >99.9999 and 99.6%, respectively, after exposure to visible light for 1 h. In response to the current COVID-19 pandemic, we also confirm that these coated fibers can inactivate a human common cold coronavirus serving as a surrogate for the SARS-CoV-2 virus.

Repurposing the Ebola and Marburg Virus Inhibitors Tilorone, Quinacrine and Pyronaridine: In vitro Activity Against SARS-CoV-2 and Potential Mechanisms.

SARS-CoV-2 is a newly identified virus that has resulted in over 1.3 M deaths globally and over 59 M cases globally to date. Small molecule inhibitors that reverse disease severity have proven difficult to discover. One of the key approaches that has been widely applied in an effort to speed up the translation of drugs is drug repurposing. A few drugs have shown in vitro activity against Ebola virus and demonstrated activity against SARS-CoV-2 in vivo . Most notably the RNA polymerase targeting remdesivir demonstrated activity in vitro and efficacy in the early stage of the disease in humans. Testing other small molecule drugs that are active against Ebola virus would seem a reasonable strategy to evaluate their potential for SARS-CoV-2. We have previously repurposed pyronaridine, tilorone and quinacrine (from malaria, influenza, and antiprotozoal uses, respectively) as inhibitors of Ebola and Marburg virus in vitro in HeLa cells and of mouse adapted Ebola virus in mouse in vivo . We have now tested these three drugs in various cell lines (VeroE6, Vero76, Caco-2, Calu-3, A549-ACE2, HUH-7 and monocytes) infected with SARS-CoV-2 as well as other viruses (including MHV and HCoV 229E). The compilation of these results indicated considerable variability in antiviral activity observed across cell lines. We found that tilorone and pyronaridine inhibited the virus replication in A549-ACE2 cells with IC 50 values of 180 nM and IC 50 198 nM, respectively. We have also tested them in a pseudovirus assay and used microscale thermophoresis to test the binding of these molecules to the spike protein. They bind to spike RBD protein with K d values of 339 nM and 647 nM, respectively. Human C max for pyronaridine and quinacrine is greater than the IC 50 hence justifying in vivo evaluation. We also provide novel insights into their mechanism which is likely lysosomotropic.

BODIPY-embedded electrospun materials in antimicrobial photodynamic inactivation.

Drug-resistant pathogens, particularly those that result in hospital acquired infections (HAIs), have emerged as a critical priority for the World Health Organization. To address the need for self-disinfecting materials to counter the threat posed by the transmission of these pathogens from surfaces to new hosts, here we investigated if a cationic BODIPY photosensitizer, embedded via electrospinning into nylon and polyacrylonitrile (PAN) nanofibers, was capable of inactivating both bacteria and viruses via antimicrobial photodynamic inactivation (aPDI). Materials characterization, including fiber morphology and the degree of photosensitizer loading, was assessed by scanning electron microscopy (SEM), thermal gravimetric analysis (TGA), and UV-visible diffuse reflectance spectroscopy (UV-Vis DRS), and demonstrated that the materials were comprised of nanofibers (125-215 nm avg. diameter) that were thermostable to >300 °C. The antimicrobial potencies of the resultant Nylon-BODIPY(+) and PAN-BODIPY(+) nanofiber materials were evaluated against four strains of bacteria recognized by the World Health Organization as either critical or high priority pathogens: Gram-positive strains methicillin-resistant S. aureus (MRSA; ATCC BAA-44) and vancomycin-resistant E. faecium (VRE; ATCC BAA-2320), and Gram-negative strains multidrug-resistant A. baumannii (MDRAB; ATCC BAA-1605) and NDM-1 positive K. pneumoniae (KP; ATCC BAA-2146). Our results demonstrated the detection limit (99.9999%; 6 log units reduction in CFU mL-1) photodynamic inactivation of three strains upon illumination (30-60 min; 40-65 ± 5 mW cm-2; 400-700 nm): MRSA, VRE, and MDRAB, but only minimal inactivation (47-75%) of KP. Antiviral studies employing PAN-BODIPY(+) against vesicular stomatitis virus (VSV), a model enveloped virus, revealed complete inactivation. Taken together, the results demonstrate the potential for electrospun BODIPY(+)-embedded nanofiber materials as the basis for pathogen-specific anti-infective materials, even at low photosensitizer loadings.

Photodynamic Polymers as Comprehensive Anti-Infective Materials: Staying Ahead of a Growing Global Threat.

To combat the global threat posed by surface-adhering pathogens that are becoming increasingly drug-resistant, we explore the anti-infective efficacy of bulk thermoplastic elastomer films containing ∼1 wt % zinc-tetra(4- N-methylpyridyl)porphine (ZnTMPyP4+), a photoactive antimicrobial that utilizes visible light to generate singlet oxygen. This photodynamic polymer is capable of inactivating five bacterial strains and two viruses with at least 99.89% and 99.95% success, respectively, after exposure to noncoherent light for 60 min. Unlike other anti-infective methodologies commonly requiring oxidizing chemicals, carcinogenic radiation, or toxic nanoparticles, our approach is nonspecific and safe/nontoxic, and sustainably relies on the availability of just oxygen and visible light.

Dengue virus NS1 enhances viral replication and pro-inflammatory cytokine production in human dendritic cells.

Dengue virus (DV) has become the most prevalent arthropod borne virus due to globalization and climate change. It targets dendritic cells during infection and leads to production of pro-inflammatory cytokines and chemokines. Several DV non-structural proteins (NS) modulate activation of human dendritic cells. We investigated the effect of DV NS1 on human monocyte-derived dendritic cells (mo-DCs) during dengue infection. NS1 is secreted into the serum of infected individuals where it interacts with various immune mediators and cell types. We purified secreted DV1 NS1 from supernatants of 293T cells that over-express the protein. Upon incubation with mo-DCs, we observed NS1 uptake and enhancement of early DV1 replication. As a consequence, mo-DCs that were pre-exposed to NS1 produced more pro-inflammatory cytokines in response to subsequent DV infection compared to DCs exposed to heat-inactivated NS1 (HNS1). Therefore the presence of exogenous NS1 is able to modulate dengue infection in mo-DCs.

The adaptor molecule Trif contributes to murine host defense during Leptospiral infection.

Leptospirosis is a zoonotic disease and is caused by pathogenic species of the Leptospira genus, including Leptospira interrogans (L. interrogans). Humans, domestic and wild animals are susceptible to acute or chronic infection. The innate immune response is a critical defense mechanism against Leptospira interrogans, and has been investigated in mouse models. Murine Toll-like receptors (TLRs) have been shown to be key factors in sensing and responding to L. interrogans infection. Specifically, TLR2, TLR4 and the TLR adaptor molecule MyD88 are essential for host defense against L. interrogans; however, the role of the TLR adaptor molecule TIR-domain-containing adaptor-inducing interferon ß (TRIF) in the response to L. interrogans has not been previously determined. In the present study, TRIF was found to play an important role during leptospiral infection. Following challenge with L. interrogans, Trif(-/-) mice exhibited delayed weight gain compared to wild-type mice. Moreover, Trif(-/-) mice exhibited an increase in L. interrogans burden in the kidneys, lungs, and blood at early time points (less than 7days post infection). Multiple components of the innate immune responses were dampened in response to leptospiral infection including transcription and production of cytokines, and the humoral response, which suggested that TRIF contributes to expression and production of cytokines important for the host defense against L. interrogans.