Viral Material Research Articles

Virality in the fast lane: How businesses can succeed with integrity on TikTok and Instagram

In the quick-paced environment of social media, TikTok and Instagram have turned into crucial instruments for brands to attain digital popularity. This paper examines ways businesses can thrive in this setting while preserving their brand’s honesty. It dives into various plans like employing emotional narratives, gentle incorporation of the brand, collaborations with influencers and realizing each platform’s distinct algorithms’ importance. The algorithms particular to the users of TikTok and Instagram assist businesses in reaching specific audience groups, but they can be problematic when it comes to maintaining uniform brand communications. The research adds a case review of Coca-Cola’s “Happiness Factory” advertising scheme as an example; this shows how combining emotional connection with understated branding may bring about viral achievement which increases loyalty and recognition towards the brand. Further, it explores reputation hazards related to forceful marketing techniques within our paper and suggests ways on blending viral material effectively with ethical plus strategic management for brands. In the end, for success in the digital marketing field, companies must always adjust and create new ideas to remain important while maintaining their basic principles.

Sewer transport conditions and their role in the decay of endogenous SARS-CoV-2 and pepper mild mottle virus from source to collection

This study presents a comprehensive analysis of the decay patterns of endogenous SARS-CoV-2 and Pepper mild mottle virus (PMMoV) within wastewaters spiked with stool from infected patients expressing COVID-19 symptoms, and hence explores the decay of endogenous SARS-CoV-2 and PMMoV targets in wastewaters from source to collection of the sample. Stool samples from infected patients were used as endogenous viral material to more accurately mirror real-world decay processes compared to more traditionally used lab-propagated spike-ins. As such, this study includes data on early decay stages of endogenous viral targets in wastewaters that are typically overlooked when performing decay studies on wastewaters harvested from wastewater treatment plants that contain already-degraded endogenous material. The two distinct sewer transport conditions of dynamic suspended sewer transport and bed and near-bed sewer transport were simulated in this study at temperatures of 4 °C, 12 °C and 20 °C to elucidate decay under these two dominant transport conditions within wastewater infrastructure. The dynamic suspended sewer transport was simulated over 35 h, representing typical flow conditions, whereas bed and near-bed transport extended to 60 days to reflect the prolonged settling of solids in sewer systems during reduced flow periods. In dynamic suspended sewer transport, no decay was observed for SARS-CoV-2, PMMoV, or total RNA over the 35-h period, and temperature ranging from 4 °C to 20 °C had no noticeable effect. Conversely, experiments simulating bed and near-bed transport conditions revealed significant decreases in SARS-CoV-2 and total RNA concentrations by day 2, and PMMoV concentrations by day 3. Only PMMoV exhibited a clear trend of increasing decay constant with higher temperatures, suggesting that while temperature influences decay dynamics, its impact may be less significant than previously assumed, particularly for endogenous RNA that is bound to dissolved organic matter in wastewater. First order decay models were inadequate for accurately fitting decay curves of SARS-CoV-2, PMMoV, and total RNA in bed and near-bed transport conditions. F-tests confirmed the superior fit of the two-phase decay model compared to first order decay models across temperatures of 4 °C–20 °C. Finally, and most importantly, total RNA normalization emerged as an appropriate approach for correcting the time decay of SARS-CoV-2 exposed to bed and near-bed transport conditions. These findings highlight the importance of considering decay from the point of entry in the sewers, sewer transport conditions, and normalization strategies when assessing and modelling the impact of viral decay rates in wastewater systems. This study also emphasizes the need for ongoing research into the diverse and multifaceted factors that influence these decay rates, which is crucial for accurate public health monitoring and response strategies.

Clinical, magnetic resonance imaging and histopathological findings of a congenital hypomyelinating leukodystrophy in a Holstein Friesian calf

AbstractA 12‐day‐old, male Holstein Friesian calf was presented to the hospital due to progressive ataxia and tremors since birth. A diffuse central nervous system disorder primarily affecting white matter was suspected based on clinical neurological examination. Blood tests were unremarkable, except for positive serology for Schmallenberg virus. Magnetic resonance imaging of the brain demonstrated severe, diffuse abnormalities primarily affecting the white matter regions of the encephalon and cranial cervical spinal cord, consistent with leukoencephalomyelopathy. Euthanasia was performed at 8 weeks of age after no clinical improvement. A postmortem examination was performed. Histopathological examination of the central nervous system revealed changes compatible with hypomyelinating leukodystrophy, with no signs of infectious agents observed. The disorder was considered most likely to be hereditary, as Schmallenberg virus infections during pregnancy in cattle have not been reported to result in such clinical, magnetic resonance imaging or histopathological findings, and no viral genomic material was identified with polymerase chain reaction.

Peningkatan Motivasi dan Hasil Belajar Kognitif Kelas X melalui Model Problem Based Learning

Learning outcomes are students' abilities that have been achieved after following a learning process that includes cognitive, affective and psychomotor aspects. Student learning outcomes are influenced by several factors, both internal and external factors. Internal factors are factors that originate from students themselves which can influence learning outcomes, one of which is students' learning motivation. The problem based learning model is designed to guide active learning and critical thinking. In PBL, students who are faced with problems are required to be able to explore for themselves through observation skills, arguing in discussions, and carrying out practices in accordance with scientific procedures. Based on the results of observations in class X7 of SMA Negeri 10 Semarang, the majority of students were still passive during learning activities. This became the background for carrying out classroom action research regarding increasing student motivation and learning outcomes through the problem based learning model. This research method is PTK with a model according to Kemmis & McTaggart which consists of three stages, namely planning, implementation and observation, and reflection. Data collection techniques include observation, distributing questionnaires, and evaluation questions in the form of daily assessments. The research results showed that there was an increase in learning motivation with a percentage of 72% in cycle I to 83% in cycle II. Furthermore, the percentage of completeness of student learning outcomes in cycle I was 77% and cycle II increased to 94%. Thus, it can be concluded that there is an increase in motivation and learning outcomes for viral material through the Problem Based Learning model.

Analytical validation of a semi-automated methodology for quantitative measurement of SARS-CoV-2 RNA in wastewater collected in northern New England.

Wastewater-based epidemiology (WBE) can be used to monitor the community presence of infectious disease pathogens of public health concern such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Viral nucleic acid has been detected in the stool of SARS-CoV-2-infected individuals. Asymptomatic SARS-CoV-2 infections make community monitoring difficult without extensive and continuous population screening. In this study, we validated a procedure that includes manual pre-processing, automated SARS-CoV-2 RNA extraction and detection workflows using both reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) and reverse transcriptase droplet digital PCR (RT-ddPCR). Genomic RNA and calibration materials were used to create known concentrations of viral material to determine the linearity, accuracy, and precision of the wastewater extraction and SARS-CoV-2 RNA detection. Both RT-qPCR and RT-ddPCR perform similarly in all the validation experiments, with a limit of detection of 50 copies/mL. A wastewater sample from a care facility with a known outbreak was assessed for viral content in replicate, and we showed consistent results across both assays. Finally, in a 2-week survey of two New Hampshire cities, we assessed the suitability of our methods for daily surveillance. This paper describes the technical validation of a molecular assay that can be used for long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.IMPORTANCEThis paper describes the technical validation of a molecular assay that can be used for the long-term monitoring of SARS-CoV-2 in wastewater as a potential tool for community surveillance to assist with public health efforts.

Development booklet 3-dimensional image-based digital as learning media on virus materials for senior high school

Learning media is something that is used to increase student’s interest in learning activities. Limited learning media makes biology material difficult for students to understand. Virus material is a type of biological material that has a lot of material and images that are difficult to understand. So, learning media needed that is concise and has attractive images in the form of booklet 3-dimensional image-based digital as a learning medium for viral material for Senior High School. This research aims to test the feasibility and practicality booklet 3-dimensional image-based digital as a learning medium for virus material at Kartika High School 1-5 Padang. This research is a development using Thiagarajan's 4-D development model. Based on the research results obtained as follows: material expert validation results 90.15% in the very good category, media experts 94.69% in the very good category, teacher practicality test results 95.83% in the very good category, student practicality test 89.10% in the very good category. Can be concluded that booklet digital material developed on viral material is feasible and capable of increasing student interest..

Variations in the persistence of 5′-end genomic and subgenomic SARS-CoV-2 RNAs in wastewater from aircraft, airports and wastewater treatment plants

Wastewater sequencing has become a powerful supplement to clinical testing in monitoring SARS-CoV-2 infections in the post-COVID-19 pandemic era. While its applications in measuring the viral burden and main circulating lineages in the community have proved their efficacy, the variations in sequencing quality and coverage across the different regions of the SARS-CoV-2 genome are not well understood. Furthermore, it is unclear how different sample origins, viral extraction and concentration methods and environmental factors impact the reads sequenced from wastewater. Using high-coverage, amplicon-based, paired-end read sequencing of viral RNA extracted from wastewater collected directly from aircraft, pooled from different aircraft and airport buildings or from regular wastewater plants, we assessed the genome coverage across the sample groups with a focus on the 5′-end region covering the leader sequence and investigated whether it was possible to detect subgenomic RNA from viral material recovered from wastewater. We identified distinct patterns in the persistence of the different genomic regions across the different types of wastewaters and the existence of chimeric reads mapping to non-amplified regions. Our findings suggest that preservation of the 5′-end of the genome and the ability to detect subgenomic RNA reads, though highly susceptible to environment and sample processing conditions, may be indicative of the quality and amount of the viral RNA present in wastewater.

Non-viral systems for intracellular delivery of genome editing tools.

A hallmark of the last decades is an extensive development of genome editing systems and technologies propelling genetic engineering to the next level. Specific and efficient delivery of genome editing tools to target cells is one of the key elements of such technologies. Conventional vectors are not always suitable for this purpose due to a limited cargo volume, risks related to cancer and immune reactions, toxicity, a need for high-purity viral material and quality control, as well as a possibility of integration of the virus into the host genome leading to overexpression of the vector components and safety problems. Therefore, the search for novel approaches to delivering proteins and nucleic acids into cells is a relevant priority. This work reviews abiotic vectors and systems for delivering genome editing tools into target cells, including liposomes and solid lipid particles, other membrane-based vesicles, cell-penetrating peptides, micelles, dendrimers, carbon nanotubes, inorganic, polymer, metal and other nanoparticles. It considers advantages, drawbacks and preferred applications of such systems as well as suitability thereof for the delivery of genome editing systems. A particular emphasis is placed on metal-organic frameworks (MOFs) and their potential in the targeted intracellular delivery of proteins and polynucleotides. It has been concluded that further development of MOF-based vectors and technologies, as well as combining MOFs with other carriers can result in safe and efficient delivery systems, which would be able to circulate in the body for a long time while recognizing target cells and ensuring cell-specific delivery and release of intact cargoes and, thereby, improving the genome editing outcome.

The Creation of A Three-Tier Multiple-Choice Diagnostic Test Instrument to Identifying High School Students' Misconceptions Regarding Biological Virus Material

This study aims to develop a diagnostic test instrument with three tiers focusing on viral material. Research and Development (R&D) constitutes a form of applied research. The participants for the Field Test were Class X students at SMAN 22 Bone, while those for the subsequent test were Class X students at SMAN 19 Bone. Interviews and documentation serve as the primary research methods employed in this study. A three-tier multiple-choice format is characterized by three levels of questioning within the multiple-choice framework. The resultant diagnostic test instrument comprises three tiers: the first tier includes questions, the second tier offers alternative explanations, and the third tier assesses respondents' confidence levels in selecting answers and providing justifications. The outcomes of this research include answer keys, test questions, and question grids as integral components of the instrument. This study demonstrates the validity of the developed instrument through validation conducted by expert validators. The reliability test yielded a robust average value of 0.91. Only 24 out of the 40 questions were deemed valid during the field test phase. Findings revealed that students exhibited an average misconception rate of 47.2%, spanning two indicators derived from the 40 questions. Notably, misconceptions were prominent in the discussion sub-topic concerning the earliest discovery of viruses and methods for distinguishing viruses and bacteria, with an average misconception rate of 56.8% in viral material. Developing a three-tier diagnostic test effectively identified students' misconceptions regarding viral material, indicating suitability and efficacy for educational application.

Within-host evolution of SARS-CoV-2: how often are de novo mutations transmitted from symptomatic infections?

Despite a relatively low mutation rate, the large number of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections has allowed for substantial genetic change, leading to a multitude of emerging variants. Using a recently determined mutation rate (per site replication), as well as within-host parameter estimates for symptomatic SARS-CoV-2 infection, we apply a stochastic transmission-bottleneck model to describe the survival probability of de novo SARS-CoV-2 mutations as a function of bottleneck size and selection coefficient. For narrow bottlenecks, we find that mutations affecting per-target-cell attachment rate (with phenotypes associated with fusogenicity and ACE2 binding) have similar transmission probabilities to mutations affecting viral load clearance (with phenotypes associated with humoral evasion). We further find that mutations affecting the eclipse rate (with phenotypes associated with reorganization of cellular metabolic processes and synthesis of viral budding precursor material) are highly favoured relative to all other traits examined. We find that mutations leading to reduced removal rates of infected cells (with phenotypes associated with innate immune evasion) have limited transmission advantage relative to mutations leading to humoral evasion. Predicted transmission probabilities, however, for mutations affecting innate immune evasion are more consistent with the range of clinically estimated household transmission probabilities for de novo mutations. This result suggests that although mutations affecting humoral evasion are more easily transmitted when they occur, mutations affecting innate immune evasion may occur more readily. We examine our predictions in the context of a number of previously characterized mutations in circulating strains of SARS-CoV-2. Our work offers both a null model for SARS-CoV-2 mutation rates and predicts which aspects of viral life history are most likely to successfully evolve, despite low mutation rates and repeated transmission bottlenecks.

Isolation and study of a strain of lumpy skin disease virus of cattle

The identification of new virus strains will prevent the development of outbreaks thanks to the development and use of vaccines. The aim of the work was to isolate and sequence the genome of lumpy skin disease virus from an epizootic in the Kostanay region. The genetic material of virions was identified by polymerase chain reaction (PCR); viral antigens and antibodies to them have been determined by enzyme-linked immunosorbent assay (ELISA) or diffusion test methods; to accumulate the material, the virus has been cultivated in lamb testicle monoclonal cell; new generation sequencing has been performed using MiSeq System and FastQ software; strain affiliation has been established by the BLASTN-alignment method. Specific amplicons of the virus with a length of 347 bp. were detected in skin samples but not found in blood. Antigens in 1:5-1:320 dilutions were identified in skin material and 2-3 passages of monoclonals; less antigenic activity was found in the blood in a 1:2 dilution. In response to the pathogen, specific immunoglobulins were synthesized in the serum of 67% of the studied animals and were detected in dilutions of 1:100-1:400. The viral material was accumulated in monoclonals and isolated in a sucrose gradient. The whole-genome sequence of the obtained material confirmed the isolation of a new strain of nodular dermatitis virus with a percentage of similarity to the closest homologues of 99.66%. The strain was named Dermatitis nodularis bovum/2018/Kostanay/KZ; the sequence has been submitted to GeneBank, and the object has been deposited in the Collection of Microorganisms under accession number M-9-21/D. The obtained information can be used to prevent the spread of foci of cattle infection

Hyperacetylated microtubules assist porcine deltacoronavirus nsp8 to degrade MDA5 via SQSTM1/p62-dependent selective autophagy.

The microtubule (MT) is a highly dynamic polymer that functions in various cellular processes through MT hyperacetylation. Thus, many viruses have evolved mechanisms to hijack the MT network of the cytoskeleton to allow intracellular replication of viral genomic material. Coronavirus non-structural protein 8 (nsp8), a component of the viral replication transcriptional complex, is essential for viral survival. Here, we found that nsp8 of porcine deltacoronavirus (PDCoV), an emerging enteropathogenic coronavirus with a zoonotic potential, inhibits interferon (IFN)-β production by targeting melanoma differentiation gene 5 (MDA5), the main pattern recognition receptor for coronaviruses in the cytoplasm. Mechanistically, PDCoV nsp8 interacted with MDA5 and induced autophagy to degrade MDA5 in wild-type cells, but not in autophagy-related (ATG)5 or ATG7 knockout cells. Further screening for autophagic degradation receptors revealed that nsp8 interacts with sequestosome 1/p62 and promotes p62-mediated selective autophagy to degrade MDA5. Importantly, PDCoV nsp8 induced hyperacetylation of MTs, which in turn triggered selective autophagic degradation of MDA5 and subsequent inhibition of IFN-β production. Overall, our study uncovers a novel mechanism employed by PDCoV nsp8 to evade host innate immune defenses. These findings offer new insights into the interplay among viruses, IFNs, and MTs, providing a promising target to develop anti-viral drugs against PDCoV.IMPORTANCECoronavirus nsp8, a component of the viral replication transcriptional complex, is well conserved and plays a crucial role in viral replication. Exploration of the role mechanism of nsp8 is conducive to the understanding of viral pathogenesis and development of anti-viral strategies against coronavirus. Here, we found that nsp8 of PDCoV, an emerging enteropathogenic coronavirus with a zoonotic potential, is an interferon antagonist. Further studies showed that PDCoV nsp8 interacted with MDA5 and sequestosome 1/p62, promoting p62-mediated selective autophagy to degrade MDA5. We further found that PDCoV nsp8 could induce hyperacetylation of MT, therefore triggering selective autophagic degradation of MDA5 and inhibiting IFN-β production. These findings reveal a novel immune evasion strategy used by PDCoV nsp8 and provide insights into potential therapeutic interventions.

COVID-19 Associated Longitudinal Extensive Transverse Myelitis with Cerebral Spinal Fluid SARS-CoV-2 Detection

There have been multiple reports of COVID-19 associated neurological symp-toms, including several cases of para-infectious myelitis.We report a case of an 84-year-old male patient presenting with acute paraparesis coinciding with active mild COVID-19 pneumonia. Spinal cord magnetic resonance imaging (MRI) showed longitudinally extensive transverse myelitis (LETM) affecting the entire cervical cord to the tenth thoracic level, and cerebral spinal fluid (CSF) was positive for SARS-CoV-2. The patient was evaluated for other causes of LETM, which were convincingly excluded. He was treated with steroid pulses and plasmapheresis, with significant imaging improvement. The patient has since made a good recovery, being now able to walk unassisted. Although several instances of COVID-19 associated myelitis have been reported, positive identification of the virus in CSF is rare. The identification of viral material may support the hypothesis of direct spinal cord infection in co-occurrence with a para-infectious inflammatory reaction.

Targeted genomic sequencing of avian influenza viruses in wetland sediment from wild bird habitats.

Diverse influenza A viruses (IAVs) circulate in wild birds, including highly pathogenic strains that infect poultry and humans. Consequently, surveillance of IAVs in wild birds is a cornerstone of agricultural biosecurity and pandemic preparedness. Surveillance is traditionally done by testing wild birds directly, but obtaining these specimens is labor intensive, detection rates can be low, and sampling is often biased toward certain avian species. As a result, local incursions of dangerous IAVs are rarely detected before outbreaks begin. Testing environmental specimens from wild bird habitats has been proposed as an alternative surveillance strategy. These specimens are thought to contain diverse IAVs deposited by a broad range of avian hosts, including species that are not typically sampled by surveillance programs. To enable this surveillance strategy, we developed a targeted genomic sequencing method for characterizing IAVs in these challenging environmental specimens. It combines custom hybridization probes, unique molecular index-based library construction, and purpose-built bioinformatic tools, allowing IAV genomic material to be enriched and analyzed with single-fragment resolution. We demonstrated our method on 90 sediment specimens from wetlands around Vancouver, Canada. We recovered 2,312 IAV genome fragments originating from all eight IAV genome segments. Eleven hemagglutinin subtypes and nine neuraminidase subtypes were detected, including H5, the current global surveillance priority. Our results demonstrate that targeted genomic sequencing of environmental specimens from wild bird habitats could become a valuable complement to avian influenza surveillance programs.IMPORTANCEIn this study, we developed genome sequencing tools for characterizing avian influenza viruses in sediment from wild bird habitats. These tools enable an environment-based approach to avian influenza surveillance. This could improve early detection of dangerous strains in local wild birds, allowing poultry producers to better protect their flocks and prevent human exposures to potential pandemic threats. Furthermore, we purposefully developed these methods to contend with viral genomic material that is diluted, fragmented, incomplete, and derived from multiple strains and hosts. These challenges are common to many environmental specimens, making these methods broadly applicable for genomic pathogen surveillance in diverse contexts.

Recombinantly expressed virus-like particles (VLPs) of canine circovirus for development of an indirect ELISA.

Canine circovirus (CanineCV) is an emerging pathogen in domestic dogs, detected in multiple countries in association with varying clinical and pathological presentations including diarrhoea, vasculitis, granulomatous inflammation, and respiratory signs. Understanding the pathology of CanineCV is confounded by the fact that it has been detected in asymptomatic dogs as well as in diseased dogs concurrently infected with known pathogens. Recombinantly expressed self-assembling Virus-like particles (VLPs) lack viral genomic material but imitate the capsid surface conformations of wild type virion, allowing arrays of biological applications including subunit vaccine development and immunodiagnostics. In this study, full length CanineCV capsid gene was expressed in Escherichia coli followed by two-step purification process to yield soluble capsid protein in high concentration. Transmission electron microscopy (TEM) confirmed the capsid antigen self-assembled into 17-20nm VLPs in glutathione S-transferase (GST) buffer, later utilised to develop an indirect enzyme-linked immunosorbent assay (iELISA). The respective sensitivity and specificity of the proposed iELISA were 94.10% and 88.40% compared with those obtained from Western blot. The mean OD450 value for western blot positive samples was 1.22 (range 0.12-3.39) and negative samples was 0.21 (range 0.07-0.41). An optimal OD450 cut-off of 0.35 was determined by ROC curve analysis. Median inter-assay and intra-assay validation revealed that the iELISA test results were reproducible with coefficients of variation 7.70 (range 5.6-11.9) and 4.21 (range 1.2-7.4). Our results demonstrated that VLP-based iELISA is a highly sensitive method for serological diagnosis of CanineCV infections in dogs, suitable for large-scale epidemiological studies.

A Paradigm Shift: Tiktok’s Impact on Social Media Marketing Strategies

The efficacy of TikTok as a social media marketing tool is examined in this study which uses a quantitative methodology. The research project ensures an accurate representation of TikTok users through the use of probability sampling, enabling trustworthy and generalizable results. Structured questionnaires that aimed to capture different facets of TikTok’s impact on customer behavior and marketing efficacy were used to gather data. The survey instrument included inquiries about the influence of social media elements on purchasing decisions, user engagement, and opinions of marketing content. A thorough examination of patterns and trends was made possible by the statistical methods used to evaluate the data, which were performed using the Statistical Package for the Social Sciences (SPSS). The study focuses on how characteristics exclusive to TikTok – like its algorithm-driven recommendations and short-form video content affect user behavior and engagement. The impact of influencer endorsements, social proof, and viral material on consumer perceptions and purchase decisions are investigated. The study employs quantitative analysis to provide empirical insights into the ways in which TikTok influences user interactions and marketing outcomes. The results should be extremely beneficial in advancing knowledge of social media marketing tactics and how well they work on TikTok, with particular attention to the useful implications for companies looking to use this platform for customer interaction and brand promotion. Overall, by applying rigorous quantitative methodologies to evaluate TikTok’s efficacy and influence in the field of digital marketing, this research offers a comprehensive analysis of the platform’s marketing potential.

Optimization of conditions for cultivation of pathogens of infectious rhinotracheitis and viral diarrhea

This article reflects the results of research on optimizing the conditions of cultivation of strain “ТК-A(VIEV)-В2” of infectious bovine rhinotracheitis (IBR) and strain “ВК-1” of bovine viral diarrhea (BVDV). It was found that the roller cultivation method is most easily applicable and provides the production of viral material with high infectious activity. A comparative analysis of the results of studying the sensitivity of continuous cell cultures to the pathogens of IBR and BVDV of cattle of the latter made it possible to select the MDBK cell line to obtain the maximum titer of viruses. The optimal dose for infecting cell culture with both pathogens was the content of viral particles from 0.2 to 0.5 TCD50/cm3 per cell. The maximum reproduction of viruses was observed using a culture concentration of cells from 110 to 120 thousand cells/cm3 and the duration of their cultivation for 2-3 days. The results obtained can be applied for the further development and creation of preventive and diagnostic tools for IBR and BVDV of cattle.

Importance of quantifying the number of viral reads in metagenomic sequencing of environmental samples from the Huanan Seafood Market.

In March 2023, the Chinese CDC publicly released raw metagenomic sequencing data for environmental samples collected in early 2020 from the Huanan Seafood Market. Prior to that data release, some scientists had suggested that these samples could be informative for establishing if animals such as raccoon dogs had been infected with severe acute respiratory syndrome virus 2 (SARS-CoV-2). However, no one had analyzed how much SARS-CoV-2 was actually present in the metagenomic sequencing data. After the raw data became available, I fully analyzed the abundance of both viral and animal genetic material in the samples. That analysis, which was published in Virus Evolution, found that the SARS-CoV-2 content of most samples was very low and that the abundance of SARS-CoV-2 was most strongly associated with animals such as largemouth bass that are not plausible candidates for having been infected. Based on these results, I concluded that the metagenomic content of the samples was not informative for determining if any non-human animals in the market had been infected with SARS-CoV-2. One of the authors of an earlier study of these samples, Florence Débarre, recently submitted a response to my paper. Here, I reply in turn to explain why it is important to quantify the abundance of viral material before drawing conclusions from metagenomic sequencing. I also report new analyses of other animal coronaviruses in the samples and show that material from some other animal coronaviruses is much more abundant than SARS-CoV-2 in samples collected on the date when most wildlife stall sampling was performed. I further show that material from some of these animal coronaviruses is associated with the animals they probably infect but that no such association exists for SARS-CoV-2. Overall, these new analyses further emphasize the importance of quantifying the actual amount of viral material in metagenomic samples and underscore why the environmental samples from the Huanan Seafood Market are not informative for determining if any non-human animals were infected with SARS-CoV-2.

Goat-derived cell line (<i>Capra hircus</i>) TCh generated by karyological and morphological transformation of YаDK-04 CCL during subcultivation with lanthanide-treated bovine serum

Ability of the continuous cell lines to evolve enables generation of new transformed cell cultures with unlimited life potential and different from the original prototypes in the process of sequential cultivation. There are practically no universal mechanisms and methods for new cell line generation. But it was noted that cell immortalization is associated with chromosomal rearrangements (chromatid morphology) and changes in the number of chromosomes. The paper presents the results of the generation of a new Testis Capra hircus(TCh) cell line, suitable for effective replication of dermatotropic and other types of animal viruses, in order to scale up viral material used for the manufacture of the means for animal disease specific prevention and diagnosis. The monolayer TCh cell line was transformed from the continuous YаDK-04 cell line as a result of more than 50 passages in the growth medium supplemented with 10% of lanthanide-treated bovine serum. Use of the bovine serum purified and supplemented with lanthanides during the cultivation of the continuous cell line YаDK-04 led to significant chromosomal rearrangements and contributed to the formation of a stable and productive new TCh cell line, which differed in cytomorphological and karyological characteristics and had unlimited potential for passaging without changing the cell karyotype and morphology. The novel continuous cell line proved to be suitable for effective reproduction of such disease pathogens as lumpy skin disease, sheep pox, peste des petits ruminants agents. These are mainly viruses of dermatotropic origin.

Direct antigen presentation is the canonical pathway of cytomegalovirus CD8 T-cell priming regulated by balanced immune evasion ensuring a strong antiviral response.

CD8 T cells are important antiviral effectors in the adaptive immune response to cytomegaloviruses (CMV). Naïve CD8 T cells can be primed by professional antigen-presenting cells (pAPCs) alternatively by "direct antigen presentation" or "antigen cross-presentation". In the case of direct antigen presentation, viral proteins are expressed in infected pAPCs and enter the classical MHC class-I (MHC-I) pathway of antigen processing and presentation of antigenic peptides. In the alternative pathway of antigen cross-presentation, viral antigenic material derived from infected cells of principally any cell type is taken up by uninfected pAPCs and eventually also fed into the MHC class-I pathway. A fundamental difference, which can be used to distinguish between these two mechanisms, is the fact that viral immune evasion proteins that interfere with the cell surface trafficking of peptide-loaded MHC-I (pMHC-I) complexes are absent in cross-presenting uninfected pAPCs. Murine cytomegalovirus (mCMV) models designed to disrupt either of the two presentation pathways revealed that both are possible in principle and can substitute each other. Overall, however, the majority of evidence has led to current opinion favoring cross-presentation as the canonical pathway. To study priming in the normal host genetically competent in both antigen presentation pathways, we took the novel approach of enhancing or inhibiting direct antigen presentation by using recombinant viruses lacking or overexpressing a key mCMV immune evasion protein. Against any prediction, the strongest CD8 T-cell response was elicited under the condition of intermediate direct antigen presentation, as it exists for wild-type virus, whereas the extremes of enhanced or inhibited direct antigen presentation resulted in an identical and weaker response. Our findings are explained by direct antigen presentation combined with a negative feedback regulation exerted by the newly primed antiviral effector CD8 T cells. This insight sheds a completely new light on the acquisition of viral immune evasion genes during virus-host co-evolution.