Common and Potential Emerging Foodborne Viruses: A Comprehensive Review.

Human viruses and viruses from animals can cause illnesses in humans after the consumption of contaminated food or water. Contamination may occur during preparation by infected food handlers, during food production because of unsuitably controlled working conditions, or following the consumption of animal-based foods contaminated by a zoonotic virus. This review discussed the recent information available on the general and clinical characteristics of viruses, viral foodborne outbreaks and control strategies to prevent the viral contamination of food products and water. Viruses are responsible for the greatest number of illnesses from outbreaks caused by food, and risk assessment experts regard them as a high food safety priority. This concern is well founded, since a significant increase in viral foodborne outbreaks has occurred over the past 20 years. Norovirus, hepatitis A and E viruses, rotavirus, astrovirus, adenovirus, and sapovirus are the major common viruses associated with water or foodborne illness outbreaks. It is also suspected that many human viruses including Aichi virus, Nipah virus, tick-borne encephalitis virus, H5N1 avian influenza viruses, and coronaviruses (SARS-CoV-1, SARS-CoV-2 and MERS-CoV) also have the potential to be transmitted via food products. It is evident that the adoption of strict hygienic food processing measures from farm to table is required to prevent viruses from contaminating our food.

Chloride Intracellular Channel Protein 1 (CLIC1) Is a Critical Host Cellular Factor for Influenza A Virus Replication.

(1) Background: Influenza A Virus (IAV) uses host cellular proteins during replication in host cells. IAV infection causes elevated expression of chloride intracellular channel protein 1 (CLIC1) in lung epithelial cells, but the importance of this protein in IAV replication is unknown. (2) In this study, we determined the role of CLIC1 in IAV replication by investigating the effects of CLIC1 knockdown (KD) on IAV viral protein translation, genomic RNA transcription, and host cellular proteome dysregulation. (3) Results: CLIC1 KD in A549 human lung epithelial cells resulted in a significant decrease in progeny supernatant IAV, but virus protein expression was unaffected. However, a significantly larger number of viral RNAs accumulated in CLIC1 KD cells. Treatment with a CLIC1 inhibitor also caused a significant reduction in IAV replication, suggesting that CLIC1 is an important host factor in IAV replication. SomaScan®, which measures 1322 proteins, identified IAV-induced dysregulated proteins in wild-type cells and in CLIC1 KD cells. The expression of 116 and 149 proteins was significantly altered in wild-type and in CLIC1 KD cells, respectively. A large number of the dysregulated proteins in CLIC1 KD cells were associated with cellular transcription and predicted to be inhibited during IAV replication. (4) Conclusions: This study suggests that CLIC1 is involved in later stages of IAV replication. Further investigation should clarify mechanism(s) for the development of anti-IAV drugs targeting CLIC1 protein.

Mammalian Reoviruses: Propagation, Quantification, and Storage.

Mammalian reoviruses are pathogens that cause gastrointestinal and respiratory infections. In humans, the mammalian reoviruses usually cause mild or subclinical disease, and they are ubiquitous, with most people mounting immunity at a young age. Reoviruses are prototypic representations of the Reoviridae family, which contains many highly pathogenic viruses. This article describes techniques for culturing mouse fibroblast L929 cell lines, the preferred cell line in which most mammalian reovirus studies take place. In addition, mammalian reovirus propagation, quantification, purification, and storage are described. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Propagation of mammalian reoviruses in cell culture from virus stocks Alternate Protocol 1: Large-scale propagation (and purification) of mammalian reoviruses in cell culture from virus stocks Basic Protocol 2: Quantification of mammalian reoviruses by plaque assay with neutral red staining Alternate Protocol 2: Quantification of mammalian reoviruses by plaque assay with crystal violet staining Basic Protocol 3: Storage of mammalian reoviruses Support Protocol 1: Growth and maintenance of mouse L929 cells Support Protocol 2: Plating L929 cells.

PSMA2 knockdown impacts expression of proteins involved in immune and cellular stress responses in human lung cells.

Proteasome subunit alpha type-2 (PSMA2) is a critical component of the 20S proteasome, which is the core particle of the 26S proteasome complex and is involved in cellular protein quality control by recognizing and recycling defective proteins. PSMA2 expression dysregulation has been detected in different human diseases and viral infections. No study yet has reported PSMA2 knockdown (KD) effects on the cellular proteome. METHODS: We used SOMAScan, an aptamer-based multiplexed technique, to measure >1300 human proteins to determine the impact of PSMA2 KD on A549 human lung epithelial cells. RESULTS: PSMA2 KD resulted in significant dysregulation of 52 cellular proteins involved in different bio-functions, including cellular movement and development, cell death and survival, and cancer. The immune system and signal transduction were the most affected cellular functions. PSMA2 KD caused dysregulation of several signaling pathways involved in immune response, cytokine signaling, organismal growth and development, cellular stress and injury (including autophagy and unfolded protein response), and cancer responses. CONCLUSIONS: In summary, this study helps us better understand the importance of PSMA2 in different cellular functions, signaling pathways, and human diseases.

Temporal proteomic analyses of human lung cells distinguish high pathogenicity influenza viruses and coronaviruses from low pathogenicity viruses.

Newly re-emerging viruses are of significant global concern. In late 2019, a new coronavirus, SARS-CoV-2, emerged in China and soon spread worldwide, causing the COVID-19 pandemic, which to date has caused >6 M deaths. There has been a wealth of studies on this new virus since its emergence. The coronaviruses consist of many animal and human pathogens, with some of the human coronavirus, such as strain OC43, normally causing only mild cold-like symptoms. Viruses usurp host cellular processes to successfully replicate. We used tandem mass tag mass spectrometry-based proteomic analyses of human lung MRC-5 cells infected with OC43 for various periods of time to delineate virus-induced host cell alterations. Numerous proteins involved in lipid metabolism, molecular transport, small molecule biochemistry, cell death and survival, humoral immune response, and inflammatory response were dysregulated. Comparison of our findings to previous studies that examined a range of differentially pathogenic influenza A viruses (IAV), and to SARS-CoV-2 data, revealed that proteins involved in the cell cycle, cytokine signaling, DNA replication, and anti-inflammatory responses were generally similarly affected by virtually all tested IAV and CoV. However, proteins involved in necrosis, protein metabolism, ECM regulation, and signal transduction were generally different. In addition, the more pathogenic CoV and IAV activated Rb-dependent repression of E2F-mediated transcription, whereas less pathogenic influenza and coronaviruses either inhibited or had no effect on this pathway.

Zika Virus Infection of Sertoli Cells Alters Protein Expression Involved in Activated Immune and Antiviral Response Pathways, Carbohydrate Metabolism and Cardiovascular Disease.

Zika virus (ZIKV), a re-emerging virus, causes congenital brain abnormalities and Guillain-Barré syndrome. It is mainly transmitted by Aedes mosquitoes, but infections are also linked to sexual transmissions. Infectious ZIKV has been isolated, and viral RNA has been detected in semen over a year after the onset of initial symptoms, but the mode of long-term persistence is not yet understood. ZIKV can proliferate in human Sertoli cells (HSerC) for several weeks in vitro, suggesting that it might be a reservoir for persistent ZIKV infection. This study determined proteomic changes in HSerC during ZIKV infections by TMT-mass spectrometry analysis. Levels of 4416 unique Sertoli cell proteins were significantly altered at 3, 5, and 7 days after ZIKV infection. The significantly altered proteins include enzymes, transcription regulators, transporters, kinases, peptidases, transmembrane receptors, cytokines, ion channels, and growth factors. Many of these proteins are involved in pathways associated with antiviral response, antigen presentation, and immune cell activation. Several immune response pathway proteins were significantly activated during infection, e.g., interferon signaling, T cell receptor signaling, IL-8 signaling, and Th1 signaling. The altered protein levels were linked to predicted activation of immune response in HSerC, which was predicted to suppress ZIKV infection. ZIKV infection also affected the levels of critical regulators of gluconeogenesis and glycolysis pathways such as phosphoglycerate mutase, phosphoglycerate kinase, and enolase. Interestingly, many significantly altered proteins were associated with cardiac hypertrophy, which may induce heart failure in infected patients. In summary, our research contributes to a better understanding of ZIKV replication dynamics and infection in Sertoli cells.

Influenza A Virus Uses PSMA2 for Downregulation of the NRF2-Mediated Oxidative Stress Response.

Influenza A virus (IAV), an obligatory intracellular parasite, uses host cellular molecules to complete its replication cycle and suppress immune responses. Proteasome subunit alpha type 2 (PSMA2) is a cellular protein highly expressed in IAV-infected human lung epithelial A549 cells. PSMA2 is part of the 20S proteasome complex that degrades or recycles defective proteins and involves proteolytic modification of many cellular regulatory proteins. However, the role of PSMA2 in IAV replication is not well understood. In this study, PSMA2 knockdown (KD) in A549 cells caused a significant reduction in extracellular progeny IAV, but intracellular viral protein translation and viral RNA transcription were not affected. This indicates that PSMA2 is a critical host factor for IAV maturation. To better understand the interplay between PSMA2 KD and IAV infection at the proteomic level, we used the SomaScan 1.3K version, which measures 1,307 proteins to analyze alterations induced by these treatments. We found seven cellular signaling pathways, including phospholipase C signaling, Pak signaling, and nuclear factor erythroid 2p45-related factor 2 (NRF2)-mediated oxidative stress response signaling, that were inhibited by IAV infection but significantly activated by PSMA2 KD. Further analysis of NRF2-mediated oxidative stress response signaling indicated IAV inhibits accumulation of reactive oxygen species (ROS), but ROS levels significantly increased during IAV infection in PSMA2 KD cells. However, IAV infection caused significantly higher NFR2 nuclear translocation that was inhibited in PSMA2 KD cells. This indicates that PSMA2 is required for NRF2-mediated ROS neutralization and that IAV uses PSMA2 to escape viral clearance via the NRF2-mediated cellular oxidative response. IMPORTANCE Influenza A virus (IAV) remains one of the most significant infectious agents, responsible for 3 million to 5 million illnesses each year and more than 50 million deaths during the 20th century. The cellular processes that promote and inhibit IAV infection and pathogenesis remain only partially understood. PSMA2 is a critical component of the 20S proteasome and ubiquitin-proteasome system, which is important in the replication of numerous viruses. This study examined host protein responses to IAV infection alone, PSMA2 knockdown alone, and IAV infection in the presence of PSMA2 knockdown and determined that interfering with PSMA2 function affected IAV maturation. These results help us better understand the importance of PSMA2 in IAV replication and may pave the way for designing additional IAV antivirals targeting PSMA2 or the host proteasome for the treatment of seasonal flu.

HLA-A, HSPA5, IGFBP5 and PSMA2 Are Restriction Factors for Zika Virus Growth in Astrocytic Cells.

(1) Background: Zika virus (ZIKV), an arbo-flavivirus, is transmitted via Aeges aegyptii mosquitoes Following its major outbreaks in 2013, 2014 and 2016, WHO declared it a Public Health Emergency of International Concern. Symptoms of ZIKV infection include acute fever, conjunctivitis, headache, muscle & joint pain and malaise. Cases of its transmission also have been reported via perinatal, sexual and transfusion transmission. ZIKV pathologies include meningo-encephalitis and myelitis in the central nervous system (CNS) and Guillain-Barré syndrome and acute transient polyneuritis in the peripheral nervous system (PNS). Drugs like azithromycin have been tested as inhibitors of ZIKV infection but no vaccines or treatments are currently available. Astrocytes are the most abundant cells in the CNS and among the first cells in CNS infected by ZIKV; (2) Methods: We previously used SOMAScan proteomics to study ZIKV-infected astrocytic cells. Here, we use mass spectrometric analyses to further explain dysregulations in the cellular expression profile of glioblastoma astrocytoma U251 cells. We also knocked down (KD) some of the U251 cellular proteins using siRNAs and observed the impact on ZIKV replication and infectivity; (3) Results & Conclusions: The top ZIKV dysregulated cellular networks were antimicrobial response, cell death, and energy production while top dysregulated functions were antigen presentation, viral replication and cytopathic impact. Th1 and interferon signaling pathways were among the top dysregulated canonical pathways. siRNA-mediated KD of HLA-A, IGFBP5, PSMA2 and HSPA5 increased ZIKV titers and protein synthesis, indicating they are ZIKV restriction factors. ZIKV infection also restored HLA-A expression in HLA-A KD cells by 48 h post-infection, suggesting interactions between this gene product and ZIKV.

Slow Off-Rate Modified Aptamer (SOMAmer) Proteomic Analysis of Patient-Derived Malignant Glioma Identifies Distinct Cellular Proteomes.

Malignant gliomas derive from brain glial cells and represent >75% of primary brain tumors. This includes anaplastic astrocytoma (grade III; AS), the most common and fatal glioblastoma multiforme (grade IV; GBM), and oligodendroglioma (ODG). We have generated patient-derived AS, GBM, and ODG cell models to study disease mechanisms and test patient-centered therapeutic strategies. We have used an aptamer-based high-throughput SOMAscan® 1.3K assay to determine the proteomic profiles of 1307 different analytes. SOMAscan® proteomes of AS and GBM self-organized into closely adjacent proteomes which were clearly distinct from ODG proteomes. GBM self-organized into four proteomic clusters of which SOMAscan® cluster 4 proteome predicted a highly inter-connected proteomic network. Several up- and down-regulated proteins relevant to glioma were successfully validated in GBM cell isolates across different SOMAscan® clusters and in corresponding GBM tissues. Slow off-rate modified aptamer proteomics is an attractive analytical tool for rapid proteomic stratification of different malignant gliomas and identified cluster-specific SOMAscan® signatures and functionalities in patient GBM cells.

Autophagy, Unfolded Protein Response, and Neuropilin-1 Cross-Talk in SARS-CoV-2 Infection: What Can Be Learned from Other Coronaviruses.

The COVID-19 pandemic is caused by the 2019-nCoV/SARS-CoV-2 virus. This severe acute respiratory syndrome is currently a global health emergency and needs much effort to generate an urgent practical treatment to reduce COVID-19 complications and mortality in humans. Viral infection activates various cellular responses in infected cells, including cellular stress responses such as unfolded protein response (UPR) and autophagy, following the inhibition of mTOR. Both UPR and autophagy mechanisms are involved in cellular and tissue homeostasis, apoptosis, innate immunity modulation, and clearance of pathogens such as viral particles. However, during an evolutionary arms race, viruses gain the ability to subvert autophagy and UPR for their benefit. SARS-CoV-2 can enter host cells through binding to cell surface receptors, including angiotensin-converting enzyme 2 (ACE2) and neuropilin-1 (NRP1). ACE2 blockage increases autophagy through mTOR inhibition, leading to gastrointestinal complications during SARS-CoV-2 virus infection. NRP1 is also regulated by the mTOR pathway. An increased NRP1 can enhance the susceptibility of immune system dendritic cells (DCs) to SARS-CoV-2 and induce cytokine storm, which is related to high COVID-19 mortality. Therefore, signaling pathways such as mTOR, UPR, and autophagy may be potential therapeutic targets for COVID-19. Hence, extensive investigations are required to confirm these potentials. Since there is currently no specific treatment for COVID-19 infection, we sought to review and discuss the important roles of autophagy, UPR, and mTOR mechanisms in the regulation of cellular responses to coronavirus infection to help identify new antiviral modalities against SARS-CoV-2 virus.

A Single Point Mutation, Asn16→Lys, Dictates the Temperature-Sensitivity of the Reovirus tsG453 Mutant.

Studies of conditionally lethal mutants can help delineate the structure-function relationships of biomolecules. Temperature-sensitive (ts) mammalian reovirus (MRV) mutants were isolated and characterized many years ago. Two of the most well-defined MRV ts mutants are tsC447, which contains mutations in the S2 gene encoding viral core protein σ2, and tsG453, which contains mutations in the S4 gene encoding major outer-capsid protein σ3. Because many MRV ts mutants, including both tsC447 and tsG453, encode multiple amino acid substitutions, the specific amino acid substitutions responsible for the ts phenotype are unknown. We used reverse genetics to recover recombinant reoviruses containing the single amino acid polymorphisms present in ts mutants tsC447 and tsG453 and assessed the recombinant viruses for temperature-sensitivity by efficiency-of-plating assays. Of the three amino acid substitutions in the tsG453 S4 gene, Asn16-Lys was solely responsible for the tsG453ts phenotype. Additionally, the mutant tsC447 Ala188-Val mutation did not induce a temperature-sensitive phenotype. This study is the first to employ reverse genetics to identify the dominant amino acid substitutions responsible for the tsC447 and tsG453 mutations and relate these substitutions to respective phenotypes. Further studies of other MRV ts mutants are warranted to define the sequence polymorphisms responsible for temperature sensitivity.

Embryonic Origins of Virus-Induced Hearing Loss: Overview of Molecular Etiology.

Hearing loss, one of the most prevalent chronic health conditions, affects around half a billion people worldwide, including 34 million children. The World Health Organization estimates that the prevalence of disabling hearing loss will increase to over 900 million people by 2050. Many cases of congenital hearing loss are triggered by viral infections during different stages of pregnancy. However, the molecular mechanisms by which viruses induce hearing loss are not sufficiently explored, especially cases that are of embryonic origins. The present review first describes the cellular and molecular characteristics of the auditory system development at early stages of embryogenesis. These developmental hallmarks, which initiate upon axial specification of the otic placode as the primary root of the inner ear morphogenesis, involve the stage-specific regulation of several molecules and pathways, such as retinoic acid signaling, Sonic hedgehog, and Wnt. Different RNA and DNA viruses contributing to congenital and acquired hearing loss are then discussed in terms of their potential effects on the expression of molecules that control the formation of the auditory and vestibular compartments following otic vesicle differentiation. Among these viruses, cytomegalovirus and herpes simplex virus appear to have the most effect upon initial molecular determinants of inner ear development. Moreover, of the molecules governing the inner ear development at initial stages, SOX2, FGFR3, and CDKN1B are more affected by viruses causing either congenital or acquired hearing loss. Abnormalities in the function or expression of these molecules influence processes like cochlear development and production of inner ear hair and supporting cells. Nevertheless, because most of such virus-host interactions were studied in unrelated tissues, further validations are needed to confirm whether these viruses can mediate the same effects in physiologically relevant models simulating otic vesicle specification and growth.

Update on Proteomic approaches to uncovering virus-induced protein alterations and virus -host protein interactions during the progression of viral infection.

INTRODUCTION: Viruses induce profound changes in the cells they infect. Understanding these perturbations will assist in designing better therapeutics to combat viral infection. System-based proteomic assays now provide unprecedented opportunity to monitor large numbers of cellular proteins. AREAS COVERED: This review will describe various quantitative and functional mass spectrometry-based methods, and complementary non-mass spectrometry-based methods, such as aptamer profiling and proximity extension assays, and examples of how each are used to delineate how viruses affect host cells, identify which viral proteins interact with which cellular proteins, and how these change during the course of a viral infection. PubMed was searched multiple times prior to manuscript submissions and revisions, using virus, viral, proteomics; in combination with each keyword. The most recent examples of published works from each search were then analyzed. EXPERT OPINION: There has been exponential growth in numbers and types of proteomic analyses in recent years. Continued development of reagents that allow increased multiplexing and deeper proteomic probing of the cell, at quantitative and functional levels, enhancements that target more important protein modifications, and improved bioinformatics software tools and pathway prediction algorithms will accelerate this growth and usher in a new era of host proteome understanding.

Zika Infection Disrupts Proteins Involved in the Neurosensory System.

Newly re-emerging viruses are of great global concern, especially when there are no therapeutic interventions available during the time of an outbreak. There are still no therapeutic interventions for the prevention of Zika virus (ZIKV) infections despite its resurgence more than a decade ago. Newborns infected with ZIKV suffer from microcephaly and delayed neurodevelopment, but the underlying causes are largely unknown. All viruses hijack the host cellular machinery to undergo successful replication. Our tandem mass tag mass spectrometry-based proteomic monitoring of cells infected with ZIKV revealed that among the thousands of host proteins dysregulated over time, many protein candidates were linked to neurodevelopmental processes, including the development of the auditory and visual/retinal system. The role of these dysregulated neurodevelopmental-associated host proteins for ZIKV propagation in eukaryotic cells remains elusive. For the first time, we present temporal neurodevelopmental proteomic responses in cells undergoing ZIKV infection. The future goal is to identify host proteins whose dysregulation results in neurosensory alterations reported in children born to ZIKV-infected mothers.

ZIKV Infection Induces DNA Damage Response and Alters the Proteome of Gastrointestinal Cells.

The zika virus (ZIKV) is a neurotropic virus that causes congenital abnormalities in babies when they are infected in utero. Some studies have reported these congenital abnormalities result from ZIKV attacking neural progenitor cells within the brain which differentiate into neurons, oligodendrocytes, and astrocytes. Each of these glial cells play important roles during development of the fetal brain. In addition to ZIKV-induced congenital abnormalities, infected patients experience gastrointestinal complications. There are presently no reports investigating the role of this virus at the proteomic level in gastrointestinal associated cells, so we conducted an in vitro proteomic study of ZIKV-induced changes in Caco-2, a colon-derived human cell line which is known to be permissive to ZIKV infection. We used SomaScan, a new aptamer-based proteomic tool to identify host proteins that are dysregulated during ZIKV infection at 12, 24, and 48 h post-infection. Bioinformatic analyses predicted that dysregulation of differentially-regulated host proteins results in various gastrointestinal diseases. Validation of the clinical relevance of these promising protein targets will add to the existing knowledge of ZIKV biology. These potential proteins may be useful targets towards the development of therapeutic interventions.

Zika virus dysregulates human Sertoli cell proteins involved in spermatogenesis with little effect on tight junctions.

Zika virus (ZIKV), a neglected tropical disease until its re-emergence in 2007, causes microcephaly in infants and Guillain-Barré syndrome in adults. Its re-emergence and spread to more than 80 countries led the World Health Organization in 2016 to declare a Public Health Emergency. ZIKV is mainly transmitted by mosquitos, but can persist in infected human male semen for prolonged periods and may be sexually transmitted. Testicular Sertoli cells support ZIKV replication and may be a reservoir for persistent ZIKV infection. Electrical impedance analyses indicated ZIKV infection rapidly disrupted Vero cell monolayers but had little effect upon human Sertoli cells (HSerC). We determined ZIKV-induced proteomic changes in HSerC using an aptamer-based multiplexed technique (SOMAscan) targeting >1300 human proteins. ZIKV infection caused differential expression of 299 proteins during three different time points, including 5 days after infection. Dysregulated proteins are involved in different bio-functions, including cell death and survival, cell cycle, maintenance of cellular function, cell signaling, cellular assembly, morphology, movement, molecular transport, and immune response. Many signaling pathways important for maintenance of HSerC function and spermatogenesis were highly dysregulated. These included IL-6, IGF1, EGF, NF-κB, PPAR, ERK/MAPK, and growth hormone signaling. Down-regulation of the PPAR signaling pathway might impact cellular energy supplies. Upstream molecule analysis also indicated microRNAs involved in germ cell development were downregulated by infection. Overall, this study leads to a better understanding of Sertoli cellular mechanisms used by ZIKV during persistent infection and possible ZIKV impacts on spermatogenesis.

Autophagy Modulators Profoundly Alter the Astrocyte Cellular Proteome.

Autophagy is a key cellular process that involves constituent degradation and recycling during cellular development and homeostasis. Autophagy also plays key roles in antimicrobial host defense and numerous pathogenic organisms have developed strategies to take advantage of and/or modulate cellular autophagy. Several pharmacologic compounds, such as BafilomycinA1, an autophagy inducer, and Rapamycin, an autophagy inhibitor, have been used to modulate autophagy, and their effects upon notable autophagy markers, such as LC3 protein lipidation and Sequestosome-1/p62 alterations are well defined. We sought to understand whether such autophagy modulators have a more global effect upon host cells and used a recently developed aptamer-based proteomic platform (SOMAscan®) to examine 1305 U-251 astrocytic cell proteins after the cells were treated with each compound. These analyses, and complementary cytokine array analyses of culture supernatants after drug treatment, revealed substantial perturbations in the U-251 astrocyte cellular proteome. Several proteins, including cathepsins, which have a role in autophagy, were differentially dysregulated by the two drugs as might be expected. Many proteins, not previously known to be involved in autophagy, were significantly dysregulated by the compounds, and several, including lactadherin and granulins, were up-regulated by both drugs. These data indicate that these two compounds, routinely used to help dissect cellular autophagy, have much more profound effects upon cellular proteins.

Aptamer Profiling of A549 Cells Infected with Low-Pathogenicity and High-Pathogenicity Influenza Viruses.

Influenza A viruses (IAVs) are important animal and human emerging and re-emerging pathogens that are responsible for yearly seasonal epidemics and sporadic pandemics. IAVs cause a wide range of clinical illnesses, from relatively mild infections by seasonal strains, to acute respiratory distress during infections with highly pathogenic avian IAVs (HPAI). For this study, we infected A549 human lung cells with lab prototype A/PR/8/34 (H1N1) (PR8), a seasonal H1N1 (RV733), the 2009 pandemic H1N1 (pdm09), or with two avian strains, an H5N1 HPAI strain or an H7N9 strain that has low pathogenicity in birds but high pathogenicity in humans. We used a newly-developed aptamer-based multiplexed technique (SOMAscan®) to examine >1300 human lung cell proteins affected by the different IAV strains, and identified more than 500 significantly dysregulated cellular proteins. Our analyses indicated that the avian strains induced more profound changes in the A549 global proteome compared to all tested low-pathogenicity H1N1 strains. The PR8 strain induced a general activation, primarily by upregulating many immune molecules, the seasonal RV733 and pdm09 strains had minimal effect upon assayed molecules, and the avian strains induced significant downregulation, primarily in antimicrobial response, cardiovascular and post-translational modification systems.

Influenza a virus-triggered autophagy decreases the pluripotency of human-induced pluripotent stem cells.

Maternal influenza infection during pregnancy was reported multiple times as the possible cause of many defects and congenital anomalies. Apart from several cases of influenza-related miscarriage during various trimesters of pregnancy, some epidemiological data suggest a link between maternal influenza infection and genetic abnormalities in offspring. However, there are no reports yet describing how maternal influenza alters cellular pathways at early stages of development to result in congenital defects in the fetus. In the present study, using proteomic approaches, we utilized human-induced pluripotent stem cells (hiPSCs) for modeling intrablastocyst infection with influenza virus to not only investigate the vulnerability and responses of pluripotent stem cells to this virus but also to determine the possible impacts of influenza on pluripotency and signaling pathways controlling differentiation and embryogenesis. Our data indicated viral protein production in influenza A virus (IAV)-infected hiPSCs. However, viral replication was restricted in these cells, but cell viability and pluripotency were negatively affected. These events occurred simultaneously with an excessive level of IAV-induced autophagy as well as cytopathic effects. Quantitative SOMAscan screening also indicated that changes in the proteome of hiPSCs corresponded to abnormal differentiation in these cells. Taken together, our results showed that IAV-modulated reduction in hiPSC pluripotency is associated with significant activation of autophagy. Further investigations are required to explore the role of IAV-induced autophagy in leading pluripotent stem cells toward abnormal differentiation and impaired development in early stages of embryogenesis.

Zika Virus Infection Disrupts Astrocytic Proteins Involved in Synapse Control and Axon Guidance.

The first human Zika virus (ZIKV) outbreak was reported in Micronesia in 2007, followed by one in Brazil in 2015. Recent studies have reported cases in Europe, Oceania and Latin America. In 2016, ZIKV transmission was also reported in the US and the World Health Organization declared it a Public Health Emergency of International Concern. Because various neurological conditions are associated with ZIKV, such as microcephaly, Guillain-Barré syndrome, and other disorders of both the central and peripheral nervous systems, including encephalopathy, (meningo)encephalitis and myelitis, and because of the lack of reliable patient diagnosis, numerous ongoing studies seek to understand molecular mechanisms underlying ZIKV pathogenesis. Astrocytes are one of the most abundant cells in the CNS. They control axonal guidance, synaptic signaling, neurotransmitter trafficking and maintenance of neurons, and are targeted by ZIKV. In this study, we used a newly developed multiplexed aptamer-based technique (SOMAScan) to examine > 1300 human astrocyte cell proteins. We identified almost 300 astrocyte proteins significantly dysregulated by ZIKV infection that span diverse functions and signaling pathways, including protein translation, synaptic control, cell migration and differentiation.