Virus Recovery Research Articles

Surveillance of SARS-CoV-2, rotavirus, norovirus genogroup II, and human adenovirus in wastewater as an epidemiological tool to anticipate outbreaks of COVID-19 and acute gastroenteritis in a city without a wastewater treatment plant in the Peruvian Highlands

The coronavirus disease 2019 (COVID-19) pandemic has demonstrated that Wastewater Based Epidemiology is a fast and economical alternative for monitoring severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the community level in high-income countries. In the present study, wastewater from a city in the Peruvian Highlands, which lacks a wastewater treatment plant, was monitored for one year to assess the relationship between the concentration of SARS-CoV-2 and the reported cases of COVID-19 in the community. Additionally, we compared the relationship between rotavirus (RV), norovirus genogroup II (NoV GGII), and human adenovirus (HAdV) with the number of reported cases of acute gastroenteritis. Before commencing the analysis of the samples, the viral recovery efficacy of three processing methods was determined in spiked wastewater with SARS-CoV-2. This evaluation demonstrated the highest recovery rate with direct analysis (72.2 %), as compared to ultrafiltration (50.8 %) and skimmed milk flocculation (5.6 %). Wastewater monitoring revealed that 72 % (36/50) of the samples tested positive for SARS-CoV-2, with direct analysis yielding the highest detection frequency and quantification of SARS-CoV-2. Furthermore, a strong correlation was observed between the concentration of SARS-CoV-2 in wastewater and the reported cases of COVID-19, mainly when we shift the concentration of SARS-CoV-2 by two weeks, which allows us to anticipate the onset of the fourth and fifth waves of the pandemic in Peru up to two weeks in advance. All samples processed using the skimmed milk flocculation method tested positive and showed high concentrations of RV, NoV GGII, and HAdV. In fact, the highest RV concentrations were detected up to four weeks before outbreaks of acute gastroenteritis reported in children under four years of age. In conclusion, the results of this study suggest that periodic wastewater monitoring is an excellent epidemiological tool for surveillance and can anticipate outbreaks of infectious diseases, such as COVID-19, in low- and middle-income countries.

Methods for virus recovery from environmental surfaces to monitor infectious viral contamination

Accurate quantification of infectious contaminants on environmental surfaces, particularly infectious viruses, is essential for contact transmission risk assessment; however, difficulties in recovering viruses from surfaces using swabs complicates this quantification process. Herein, we identified the factors that significantly affected virus recovery rates and developed an ideal swab method that yielded the highest rate of virus recovery. We comprehensively analyzed the effects of swab type (cotton/polyester), swab water content (wet/dry conditions), surface material, and surface area on the rates of viral RNA and infectious virus recovery.The virus recovery rate was significantly lower than the viral RNA recovery rate (P < 0.01), indicating difficulty in the quantification of infectious viruses. The virus recovery rate was significantly higher under wet conditions than that under dry conditions (P < 0.006), and the virus recovery rate obtained using cotton swabs was significantly higher than that using polyester swabs (P < 0.0001). Furthermore, the virus recovery rate had a strong negative correlation (correlation coefficient >0.8) with the target surface area. The maximum surface area where the virus recovery rate was ≥10% (MSA-10%) was identified as the maximum quantifiable area. For influenza virus recovery, MSA-10% on polyvinyl chloride (PVC) sheet, PVC leather, stainless steel, silicone, glass, and polycarbonate surfaces was 66.7, 193, 60.2, 144, 105, and 15.6 cm2, respectively. For feline calicivirus recovery, MSA-10% on PVC sheet, PVC leather, stainless steel, silicone, glass, and polycarbonate surfaces was 210, 111, 2120, 250, 322, and 180 cm2, respectively.The most accurate and ideal method for quantifying infectious viruses on environmental surfaces with the highest recovery rates meets three specifications: “wet conditions,” “the use of cotton swabs,” and “a target surface area of approximately 10 cm2.

Age-differential CD13 and interferon expression in airway epithelia affect SARS-CoV-2 infection - Effects of vitamin D

Young age and high vitamin D plasma levels have been associated with lower SARS-CoV-2 infection risk and favourable disease outcomes. This study investigated mechanisms associated with differential responses to SARS-CoV-2 across age groups and effects of vitamin D. Nasal epithelia were collected from healthy children and adults and cultured for four weeks at the air-liquid interface with and without vitamin D. Gene expression and DNA methylation were investigated. Surface protein expression was confirmed by immunofluorescence while vitamin D receptor recruitment to the DNA was analysed through chromatin immunoprecipitation. HEp-2 cells were used for protein co-immunoprecipitation and luciferase reporter assays. Compared to children, airway epithelia from adults show higher viral RNA recovery following infection. This was associated with higher ANPEP/CD13, reduced type I interferon expression, and differential DNA methylation. In cells from adults, exposure to vitamin D reduced TTLL-12 expression, a negative regulator of the interferon response. This was mediated by vitamin D receptor recruitment to TTLL12, where it instructs DNA methylation through DNA methyltransferase 1. This study links age-dependent differential expression of CD13 and type I interferon to variable infection of upper airway epithelia. Furthermore, it provides molecular evidence for vitamin D reducing viral replication by inhibiting TTLL-12.

Synthesis and Morphological Control of Fe Metal Particles By Controlling the Metal Complex Condition in Basic Solution

Coronavirus outbreaks have recently necessitated rapid virus recovery and vaccine developments. Magnetic beads are essential components to recover virus in an immunoprecipitation process. Pure iron particles with a diameter of 1 µm can be used as core materials of high-performance magnetic beads.Thus, the development of effective and green synthesis method of Fe nanoparticles is important. However, Fe nanoparticles were synthesized only in acidic solution by using strong reducing reagent, such as NaBH4, while it cannot be achieved in the basic solution since iron hydroxide, which is easily formed in this condition, cannot be reduced into metals. However, former is not suitable for commercial synthesis method because of its intense reducing process. Thus, the reducing rout of Fe species under basic condition should be developed.Here, until now, we tried to synthesis the various metal/alloy nanoparticles, such as Pd20Te7 nanoparticles and uniform Cu-In (CI) alloy nanoparticles, by utilizing the restrict metallic complexes controlling method in an aqueous solution under room temperature. In this method, reduction potential of the metal species can be controlled under the room temperature, consequently metal/alloy nanomaterials with uniform and well crystallized structure can be successfully synthesized.So, in this study, we proposed the synthesis method of iron particles under basic conditions, and morphological conditions control of iron particles by utilizing the “gel-sol” method using gel networks, which is a safe and simple approach to suppress the chain crystallization.Calculation results indicated that Fe complex can be restricted into one species as the function of pH, complexing reagent species and/also concentration. For example, Fe complex can be restricted into single complex in the basic condition (for example 8<pH<11).By using this homogeneous condition, metal Fe nanoparticles were successfully synthesized. The addition of gel networks into the synthesis system can effectively suppress the chain crystallization of iron particles. Furthermore, in order to obtain iron particles with organic-free surface, we proposed a novel method to decompose and remove the gel networks.Other results will present in our presentation. This work was supported by JSPS KAKENHI Grant Number 21H03628 and DOWA holdings.[1] H. Takahashi et.,al, Applied Catalysis A: General 392 (2011) 80–85

Alternative probe hybridization buffers for target RNA depletion and viral sequence recovery in NGS for poultry samples

Non-targeted next generation sequencing (NGS) is widely applied to identify the diversity of pathogens in field samples. However, abundance of host RNA (especially rRNA) and other environmental nucleic acids can reduce the abundance of pathogen specific reads of interest, reduce depth of coverage and increase surveillance costs. We presently deplete chicken- and selected bacterial-specific rRNAs in poultry field RNA samples with complementary DNA probes in a commercially available probe hybridization buffer followed by digestion of the RNA:DNA hybrids with RNase H. Because the current buffer is an expensive special order reagent of proprietary composition, we tested in-house and other commercially available buffers and identified a viable alternative that yields equivalent host rRNA depletion and viral-specific reads in poultry samples as the current special order reagent but at a reduced cost.

Effects of the nozzle-to-nozzle distance on the performance of the water condensation growth-based virus aerosol concentrator

Efficient collection of airborne viruses is crucial for monitoring and preventing the spread of respiratory diseases. The laminar flow water condensation-based growth tube collector (GTC) is a microbial sampler extensively used to collect virus aerosol particles owing to its high recovery of viable viruses. The GTC involves multiple nozzles and eight tubes for sampling flow rates of 6–8 L per minute (LPM). To allow the GTC to operate at a higher flow rate, optimizing GTC performance through investigation of the effects of nozzle-to-nozzle distance is essential. Herein, we examined these effects on the collection efficiency and viability of airborne MS2 viruses collected by the growth-based virus aerosol concentrator (GVC), which is a lab-made single-tubed GTC, at flow rates of up to 6 LPM per tube. The flow interactions between adjacent jets were analyzed by observing the particle deposition patterns. The collection efficiencies of MS2 virus aerosol particles and the enrichment ratio of the GVC increased with increasing nozzle-to-nozzle distance. The infectious virus concentration relative to that in the virus suspension (RIVC) ranged from 0.0093 to 0.095 for 5 min of sampling. At flow rates of 1 and 6 LPM, the RIVC decreased as the nozzle-to-nozzle distance decreased, which may be ascribed to the higher shear stress at shorter nozzle-to-nozzle distances applied to the viruses collected through the cross-flows driven by adjacent jets. These measured RIVC values are much larger than those collected by the BioSampler. These findings can provide important insights into optimizing multiple nozzles in high-flow-rate GTCs.

Establishment of a new reverse genetics system for respiratory syncytial virus under the control of RNA polymerase II.

A reverse genetics system for the respiratory syncytial virus (RSV), which causes acute respiratory illness, is an effective tool for understanding the pathogenicity of RSV. To date, a method dependent on T7 RNA polymerase is commonly used for RSV. Although this method is well established and recombinant RSV is well rescued from transfected cells, the requirement for artificial supply of T7 RNA polymerase limits its application. To overcome this, we established a reverse genetics system dependent on RNA polymerase II, which is more convenient for the recovery of recombinant viruses from various cell lines. First, we identified human cell lines with high transfection efficiency in which RSV can replicate effectively. Two human cell lines, Huh-7 and 293T, permitted the propagation of recombinant green fluorescent protein-expressing RSV. Our minigenome system revealed that efficient transcription and replication of RSV occurred in both Huh-7 and 293T cells. We then confirmed that recombinant green fluorescent protein-expressing RSV was rescued in both Huh-7 and 293T cells. Furthermore, the growth capability of viruses rescued from Huh-7 and 293T cells was similar to that of recombinant RSV rescued using the conventional method. Thus, we succeeded in establishing a new reverse genetics system for RSV that is dependent on RNA polymerase II.

Challenges detecting SARS-CoV-2 in Costa Rican domestic wastewater and river water

This study presents the development of a SARS-CoV-2 detection method for domestic wastewater and river water in Costa Rica, a middle-income country in Central America. Over a three-year period (November to December 2020, July to November 2021, and June to October 2022), 80 composite wastewater samples (43 influent and 37 effluent) were collected from a Wastewater Treatment Plant (SJ-WWTP) located in San José, Costa Rica. Additionally, 36 river water samples were collected from the Torres River near the SJ-WWTP discharge site. A total of three protocols for SARS-CoV-2 viral concentration and RNA detection and quantification were analyzed. Two protocols using adsorption-elution with PEG precipitation (Protocol A and B, differing in the RNA extraction kit; n = 82) were used on wastewater samples frozen prior to concentration, while wastewater (n = 34) collected in 2022 were immediately concentrated using PEG precipitation. The percent recovery of Bovine coronavirus (BCoV) was highest using the Zymo Environ Water RNA (ZEW) kit with PEG precipitation executed on the same day as collection (mean 6.06 % ± 1.37 %). It was lowest when samples were frozen and thawed, and viruses were concentrated using adsorption-elution and PEG concentration methods using the PureLink™ Viral RNA/DNA Mini (PLV) kit (protocol A; mean 0.48 % ± 0.23 %). Pepper mild mottle virus and Bovine coronavirus were used as process controls to understand the suitability and potential impact of viral recovery on the detection/quantification of SARS-CoV-2 RNA. Overall, SARS-CoV-2 RNA was detected in influent and effluent wastewater samples collected in 2022 but not in earlier years when the method was not optimized. The burden of SARS-CoV-2 at the SJ-WWTP decreased from week 36 to week 43 of 2022, coinciding with a decline in the national COVID-19 prevalence rate. Developing comprehensive nationwide surveillance programs for wastewater-based epidemiology in low-middle-income countries involves significant technical and logistical challenges.

Translation Arrest: A Key Player in Plant Antiviral Response.

Plants evolved several mechanisms to protect themselves against viruses. Besides recessive resistance, where compatible host factors required for viral proliferation are absent or incompatible, there are (at least) two types of inducible antiviral immunity: RNA silencing (RNAi) and immune responses mounted upon activation of nucleotide-binding domain leucine-rich repeat (NLR) receptors. RNAi is associated with viral symptom recovery through translational repression and transcript degradation following recognition of viral double-stranded RNA produced during infection. NLR-mediated immunity is induced upon (in)direct recognition of a viral protein by an NLR receptor, triggering either a hypersensitive response (HR) or an extreme resistance response (ER). During ER, host cell death is not apparent, and it has been proposed that this resistance is mediated by a translational arrest (TA) of viral transcripts. Recent research indicates that translational repression plays a crucial role in plant antiviral resistance. This paper reviews current knowledge on viral translational repression during viral recovery and NLR-mediated immunity. Our findings are summarized in a model detailing the pathways and processes leading to translational arrest of plant viruses. This model can serve as a framework to formulate hypotheses on how TA halts viral replication, inspiring new leads for the development of antiviral resistance in crops.

Treatment Outcomes of Dolutegravir- Versus Efavirenz-Based Highly Active Antiretroviral Therapy Regimens Among Treatment-Naive People Living With HIV.

In India, following the implementation of the dolutegravir (DTG) based regimen, only a few studies compared the outcomes of DTG and efavirenz (EFV) based regimens. Therefore, this study aimed to assess virological suppression and gain in CD4+ counts of DTG and EFV-based antiretroviral therapy (ART) regimens. A retrospective study was conducted and the entire sample (n=140) was divided into two major classes as DTG group (n=70) and EFV group (n=70) further classified as tenofovir/lamivudine/dolutegravir (TLD) and tenofovir/lamivudine/efavirenz (TLE) regimen. Data was collected on socio-demographic characteristics, laboratory measures, and clinical and drug-related variables.For quantitative and qualitative data analysis, respectively, the T-tests and Chi-square tests were applied. The mean CD4+ gain was comparable in both regimens after six months of ART but significant after 12 months of ART in the TLD group. Viral load suppression was achieved in 55.71% of clients in the TLE group after six months of ART while in the TLD group, 88.57% of clients achieved virologic suppression which was highly significant. Clients who remained on the DTG-based regimen gained significantly more weight at 12 months (mean +6.15 kg) as compared to the EFV-based regimen (mean +1.85 kg). After 12 months of ART, the majority of laboratory variables were unaffected by either regimen with the exception of serumcreatinine and random blood sugar (RBS) in the TLD group. Our study provides real-life evidence of better outcomes of therapy with DTG over EFV in terms of viral load suppression but immunologic recovery is equivalent in EFV-based regimens after six months of treatment. We recommend the use of DTG only in clients with a high baseline viral load as it costs approximately twice as much as EFV when cost-effectiveness is taken into account.

VirMAP for cancer: Characterization of the intratumoral virome in virally associated cancers and a resource for investigators.

2568 Background: A major barrier to understanding the role of viruses in cancer progression has been the lack of sensitivity of current tools to assemble viral genomes in an untargeted way. Here, we utilize our novel VirMAP algorithm to identify eukaryotic and prokaryotic viruses associated with cancers, using RNA sequencing data from The Cancer Genome Atlas (TCGA). Methods: VirMAP was run on 16 TCGA RNASeq datasets of primary tumor tissues, with a focus on cancers with known viral associations (including cervical, head and neck, liver, stomach, esophagus and ovarian). Previously published survival data were used for exploratory analysis. Overall survival (OS) was assessed using the Kaplan-Meier method, and p-values were calculated using log-rank test. Multivariate analysis (MVA) was done using Cox proportional hazards model to control for age, sex, stage, and histology. Results: Viral sequence recovery (among 537 billion sequencing reads from 6185 patients) was at least equal to previously published algorithms such as Kraken, but with higher taxonomic resolution; overall recovery was 0.0041% from 16 tumor types. Cancers with high viral loads such as cervical cancer demonstrated high recovery rates (0.012% of all reads mapped to viral taxa, followed by 0.004% for liver, 0.0018% head and neck, 0.0016% stomach, 0.0009% esophageal, and 0.0002% ovarian). There was a wide range of viral signature richness, with cervical cancer showing the highest viral read count predominated by 18 different HPV serotypes; in contrast, stomach cancer had lower abundance but contained over 300 unique viral species. Exploratory analysis revealed novel correlations between viral signatures and OS. In cervical cancer, presence of HPV45 predicted worse survival (median OS 837 days vs 4086, p=0.0043); MVA demonstrated hazard ratio (HR) of 5.7 [2.6-11.4, p&lt;0.0001]. In stomach cancer, patients often had more than one viral taxon present (mean 3.7, range 0-18); presence of seven or more unique taxa predicted worse OS (median OS 403 days vs 1153, p=0.0001). MVA showed HR of 1.1 for every additional taxon (p=0.0004). Across all 16 tumor types, the detection of phage virus was associated with worse OS (1073 days vs. 2094 days for non-phage virus and 2131 days for no virus, p&lt;0.0001). Conclusions: VirMAP performs deep characterization of the tumor-associated virome in both high and low abundance settings, with generation of partial genome reconstructions and strain level taxonomic classification including phages. Further work is ongoing to optimize viral read normalization based on genome length, as well as subset analysis with disease-specific modifiers such as histology, race, and exposures. Utilizing publicly available platforms such as TCGA, clinical and correlative published data can be leveraged to better understand the nuanced pleotropic effects of the tumor virome on cancer progression.

Zika virus RNA persistence and recovery in water and wastewater: An approach for Zika virus surveillance in resource-constrained settings

During the 2015–2016 Zika virus (ZIKV) epidemic in the Americas, serological cross-reactivity with other flaviviruses and relatively high costs of nucleic acid testing in the region hindered the capacity for widespread diagnostic testing. In such cases where individual testing is not feasible, wastewater monitoring approaches may offer a means of community-level public health surveillance. To inform such approaches, we characterized the persistence and recovery of ZIKV RNA in experiments where we spiked cultured ZIKV into surface water, wastewater, and a combination of both to examine the potential for detection in open sewers serving communities most affected by the ZIKV outbreak, such as those in Salvador, Bahia, Brazil. We used reverse transcription droplet digital PCR to quantify ZIKV RNA. In our persistence experiments, we found that the persistence of ZIKV RNA decreased with increasing temperature, significantly decreased in surface water versus wastewater, and significantly decreased when the initial concentration of virus was lowered by one order of magnitude. In our recovery experiments, we found higher percent recovery of ZIKV RNA in pellets versus supernatants from the same sample, higher recoveries in pellets using skimmed milk flocculation, lower recoveries of ZIKV RNA in surface water versus wastewater, and lower recoveries from a freeze thaw. We also analyzed samples collected from Salvador, Brazil during the ZIKV outbreak (2015–2016) that consisted of archived samples obtained from open sewers or environmental waters thought to be contaminated by sewage. Although we did not detect any ZIKV RNA in the archived Brazil samples, results from these persistence and recovery experiments serve to inform future wastewater monitoring efforts in open sewers, an understudied and important application of wastewater monitoring.

Simple Bioparticle Filtration Device Based on an Ultralow-Fouling Zwitterionic Polyurethane Membrane for Rapid Large-Volume Separation of Plasma and Viruses from Whole Blood

Plasma separation from whole blood is oftent required as an essential first step when performing blood tests with a viral assay. However, developing a point-of-care plasma extraction device with a large output and high virus recovery remains a significant obstacle to the success of on-site viral load tests. Here, we report a portable, easy-to-use, cost-efficient, membrane-filtration-based plasma separation device that enables rapid large-volume plasma extraction from whole blood, designed for point-of-care virus assays. The plasma separation is realized by a low-fouling zwitterionic polyurethane-modified cellulose acetate (PCBU-CA) membrane. The zwitterionic coating on the cellulose acetate membrane can decrease surface protein adsorption by 60% and increase plasma permeation by 46% compared with a pristine membrane. The PCBU-CA membrane, with its ultralow-fouling properties, enables rapid plasma separation. The device can yield a total of 1.33 mL plasma from 10 mL whole blood in 10 min. The extracted plasma is cell-free and exhibits a low hemoglobin level. In addition, our device demonstrated a 57.8% T7 phage recovery in the separated plasma. The results of real-time polymerase chain reaction analysis confirmed that the nucleic acid amplification curve of the plasma extracted by our device is comparable to that obtained by centrifugation. With its high plasma yield and good phage recovery, our plasma separation device provides an excellent replacement for traditional plasma separation protocols for point-of-care virus assays and a broad spectrum of clinical tests.

Efficient adeno-associated virus serotype 5 capture with affinity functionalized nanofiber adsorbents.

Adeno-associated viruses (AAVs) are one of the most promising tools for gene therapy applications. These vectors are purified using affinity and ion exchange chromatography, typically using packed beds of resin adsorbents. This leads to diffusion and pressure drop limitations that affect process productivity. Due to their high surface area and porosity, electrospun nanofiber adsorbents offer mass transfer and flow rate advantages over conventional chromatographic media. The present work investigated the use of affinity cellulose-based nanofiber adsorbents for adeno-associated virus serotype 5 (AAV5) capture, evaluating dynamic binding capacity, pressure drop, and AAV5 recovery at residence times (RT) less than 5s. The dynamic binding capacity was found to be residence time-dependent, but nevertheless higher than 1.0 × 1014TP mL-1 (RT = 1.6s), with a pressure drop variation of 0.14MPa obtained after loading more than 2,000 column volumes of clarified AAV5 feedstock. The single affinity chromatography purification step using these new affinity adsorbents resulted in 80% virus recovery, with the removal of impurities comparable to that of bead-based affinity adsorbents. The high binding capacity, virus recovery and reduced pressure drop observed at residence times in the sub-minute range can potentially eliminate the need for prior concentration steps, thereby reducing the overall number of unit operations, process time and costs.

Infection tracing and organ tropism of Siniperca chuatsi rhabdovirus expressing enhanced green fluorescent protein

Siniperca chuatsi Rhabdovirus (SCRV) is one of the important pathogens that infect mandarin fish. Recovery of recombinant viruses expressing fluorescent protein is an important method for virology research. In this study, the enhanced green fluorescent protein (EGFP) gene was inserted between the leader and the N gene of the SCRV genome. The recombinant SCRV of rS3-LeNEGFP expressing the EGFP was rescued, and the virus titer was as high as 2 × 106 PFU/mL in EPC cells. The insertion of the EGFP gene in the genome of SCRV was stable. EGFP was expressed efficiently after 10 passages in EPC cells. In vitro growth characterization, pathogenicity and organ tropism in vivo were studied. Compared to the parental rS3, rS3-LeNEGFP was defective in vitro, but not attenuated in vivo. The mortality of red zebrafish caused by rS3-LeNEGFP was the same as the parental SCRV rS3. The earliest expression of EGFP was detectable at 2 h, and clear fluorescence spots appeared at 8 h post-infection in EPC cells. The liver, heart, and brain were the major organ targets of rS3-LeNEGFP in vivo, and the kidney, gill, and spermatocyte were also invaded. The organ infection in vitro was different from that in vivo. This study is of great significance for revealing the pathogenesis of SCRV and developing vaccines and antiviral drugs for SCRV in the future.

Aromatic amino acid metabolites influence interferon signaling and Influenza infectivity

Abstract The gut microbiota extensively metabolizes the three aromatic amino acids. Previously we found that a tyrosine metabolite can alter type I interferon (IFN) signaling. Herein we screened 17 metabolites of these amino acids for effects on IFN signaling alone and with LPS, IFN-β, and Influenza utilizing IFN reporter human lung epithelial cells (A549) and monocytes (THP1). While the tryptophan metabolites reduced IFN signaling in both cell types, tyrosine and phenylalanine metabolites had varied effects, depending on cell type. The cells were also treated with pooled metabolites from each family and a composite treatment with all three family metabolites. The composite treatment reduced IFN signaling in both cell types, suggesting a tryptophan metabolite effect dominance. Strikingly, when pretreated with composite treatment and infected with influenza, cells had reduced viral recovery and IFN signaling in both cell types. RNA sequencing validated earlier observation of reduced viral loads and IFN signaling. Gene silencing utilizing single siRNAs for upregulated genes identified (EGR2, ATP6V0D2, SPOCK1, and IL31RA) did not completely abrogate the repressed IFN signaling. However, these targets demonstrated linkage to TGF-beta signaling, and treatment with a TGF-beta inhibitor and gene silencing of all four gene targets led to a significant reversal of IFN signaling suppression. Our in vivo work demonstrated that intranasal administration of these metabolites prior to influenza infection leads to reduced animal weight loss, viral titers, and histological inflammation. Our work implicates that microbial metabolites alter IFN signaling through TGF-beta signaling and can promote beneficial outcomes during influenza infection. The study was supported by grant from NIH (K08 AI135097-02) and Burroughs Wellcome Fund, awarded to Dr. Ashley L. Steed

Anti-COVID-19 Potential of Ellagic Acid and Polyphenols of Punica granatum L.

Pomegranate (Punica granatum L.) is a rich source of polyphenols, including ellagitannins and ellagic acid. The plant is used in traditional medicine, and its purified components can provide anti-inflammatory and antioxidant activity and support of host defenses during viral infection and recovery from disease. Current data show that pomegranate polyphenol extract and its ellagitannin components and metabolites exert their beneficial effects by controlling immune cell infiltration, regulating the cytokine secretion and reactive oxygen and nitrogen species production, and by modulating the activity of the NFκB pathway. In vitro, pomegranate extracts and ellagitannins interact with and inhibit the infectivity of a range of viruses, including SARS-CoV-2. In silico docking studies show that ellagitannins bind to several SARS-CoV-2 and human proteins, including a number of proteases. This warrants further exploration of polyphenol-viral and polyphenol-host interactions in in vitro and in vivo studies. Pomegranate extracts, ellagitannins and ellagic acid are promising agents to target the SARS-CoV-2 virus and to restrict the host inflammatory response to viral infections, as well as to supplement the depleted host antioxidant levels during the stage of recovery from COVID-19.

Editorial for the special issue “Control‐theoretic approaches for systems in the life sciences”

Editorial for the special issue “Control<scp>‐theoretic</scp> approaches for systems in the life sciences”

A multidisciplinary approach to the identification of the protein-RNA connectome in double-stranded RNA virus capsids.

How multi-segmented double-stranded RNA (dsRNA) viruses correctly incorporate their genomes into their capsids remains unclear for many viruses, including Bluetongue virus (BTV), a Reoviridae member, with a genome of 10 segments. To address this, we used an RNA-cross-linking and peptide-fingerprinting assay (RCAP) to identify RNA binding sites of the inner capsid protein VP3, the viral polymerase VP1 and the capping enzyme VP4. Using a combination of mutagenesis, reverse genetics, recombinant proteins and in vitro assembly, we validated the importance of these regions in virus infectivity. Further, to identify which RNA segments and sequences interact with these proteins, we used viral photo-activatable ribonucleoside crosslinking (vPAR-CL) which revealed that the larger RNA segments (S1-S4) and the smallest segment (S10) have more interactions with viral proteins than the other smaller segments. Additionally, using a sequence enrichment analysis we identified an RNA motif of nine bases that is shared by the larger segments. The importance of this motif for virus replication was confirmed by mutagenesis followed by virus recovery. We further demonstrated that these approaches could be applied to a related Reoviridae member, rotavirus (RV), which has human epidemic impact, offering the possibility of novel intervention strategies for a human pathogen.

Influence of membrane pore-size on the recovery of endogenous viruses from wastewater using an adsorption-extraction method

The ongoing COVID-19 pandemic has emphasized the significance of wastewater surveillance in monitoring and tracking the spread of infectious diseases, including SARS-CoV-2. The wastewater surveillance approach detects genetic fragments from viruses in wastewater, which could provide an early warning of outbreaks in communities. In this study, we determined the concentrations of four types of endogenous viruses, including non-enveloped DNA (crAssphage and human adenovirus 40/41), non-enveloped RNA (enterovirus), and enveloped RNA (SARS-CoV-2) viruses, from wastewater samples using the adsorption-extraction (AE) method with electronegative HA membranes of different pore sizes (0.22, 0.45, and 0.80 µm). Our findings showed that the membrane with a pore size of 0.80 µm performed comparably to the membrane with a pore size of 0.45 µm for virus detection/quantitation (repeated measurement one-way ANOVA; p > 0.05). We also determined the recovery efficiencies of indigenous crAssphage and pepper mild mottle virus, which showed recovery efficiencies ranging from 50% to 94% and from 20% to 62%, respectively. Our results suggest that the use of larger pore size membranes may be beneficial for processing larger sample volumes, particularly for environmental waters containing low concentrations of viruses. This study offers valuable insights into the application of the AE method for virus recovery from wastewater, which is essential for monitoring and tracking infectious diseases in communities.