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Recently identified broadly neutralizing antibodies (bnAbs) show great potential for clinical interventions against HIV-1 infection. However, resistant strains may impose substantial challenges. Here, we report on the identification and characterization of a panel of HIV-1 strains with broad and potent resistance against a large number of bnAbs, particularly those targeting the CD4-binding site (CD4bs). Site-directed mutagenesis revealed that several key epitope mutations facilitate resistance and are located in the inner domain, loop D, and β23/loop V5/β24 of HIV-1 gp120. The resistance is largely correlated with binding affinity of antibodies to the envelope trimers expressed on the cell surface. Our results therefore demonstrate the existence of broadly resistant HIV-1 strains against CD4bs neutralizing antibodies. Treatment strategies based on the CD4bs bnAbs must overcome such resistance to achieve optimal clinical outcomes. |
Pemphigus vulgaris (PV) is an autoimmune bullous disease of the skin and mucous membranes characterized by the presence of circulating and tissue-bound autoantibodies against keratinocyte cell surface antigens, specifically desmoglein (Dsg) 1 and 3. The pathogenic role of anti-Dsg antibodies is well-established, while the mechanism of blister formation is only partly defined. We have applied a previously developed method for the efficient immortalization of IgG+ memory B cells to identify novel target antigens in PV. A human monoclonal antibody reactive with a hitherto unreported non-Dsg antigen was isolated. Immunoprecipitation and immunoblotting studies with keratinocyte extracts indicated α-catenin as the putative antigen, then confirmed by immunoblotting on the recombinant protein. Four of ten PV sera reacted with recombinant α-catenin. Although the isolated human monoclonal antibody was per se unable to dissociate keratinocyte monolayers and also to synergize with a pathogenic antibody in vitro, further studies are warranted to assess its possible in vivo contribution in the multifactorial pathogenesis and heterogeneous manifestations of PV disease. |
BACKGROUND: The majority of infants hospitalized with primary respiratory syncytial virus (RSV) infection have no obvious risk factors for severe disease. OBJECTIVE: The aim of this study (Assessing Predictors of Infant RSV Effects and Severity, AsPIRES) was to identify factors associated with severe disease in full-term healthy infants younger than 10 months with primary RSV infection. METHODS: RSV infected infants were enrolled from 3 cohorts during consecutive winters from August 2012 to April 2016 in Rochester, New York. A birth cohort was prospectively enrolled and followed through their first winter for development of RSV infection. An outpatient supplemental cohort was enrolled in the emergency department or pediatric offices, and a hospital cohort was enrolled on admission with RSV infection. RSV was diagnosed by reverse transcriptase-polymerase chain reaction. Demographic and clinical data were recorded and samples collected for assays: buccal swab (cytomegalovirus polymerase chain reaction, PCR), nasal swab (RSV qualitative PCR, complete viral gene sequence, 16S ribosomal ribonucleic acid [RNA] amplicon microbiota analysis), nasal wash (chemokine and cytokine assays), nasal brush (nasal respiratory epithelial cell gene expression using RNA sequencing [RNAseq]), and 2 to 3 ml of heparinized blood (flow cytometry, RNAseq analysis of purified cluster of differentiation [CD]4+, CD8+, B cells and natural killer cells, and RSV-specific antibody). Cord blood (RSV-specific antibody) was also collected for the birth cohort. Univariate and multivariate logistic regression will be used for analysis of data using a continuous Global Respiratory Severity Score (GRSS) as the outcome variable. Novel statistical methods will be developed for integration of the large complex datasets. RESULTS: A total of 453 infants were enrolled into the 3 cohorts; 226 in the birth cohort, 60 in the supplemental cohort, and 78 in the hospital cohort. A total of 126 birth cohort infants remained in the study and were evaluated for 150 respiratory illnesses. Of the 60 RSV positive infants in the supplemental cohort, 42 completed the study, whereas all 78 of the RSV positive hospital cohort infants completed the study. A GRSS was calculated for each RSV-infected infant and is being used to analyze each of the complex datasets by correlation with disease severity in univariate and multivariate methods. CONCLUSIONS: The AsPIRES study will provide insights into the complex pathogenesis of RSV infection in healthy full-term infants with primary RSV infection. The analysis will allow assessment of multiple factors potentially influencing the severity of RSV infection including the level of RSV specific antibodies, the innate immune response of nasal epithelial cells, the adaptive response by various lymphocyte subsets, the resident airway microbiota, and viral factors. Results of this study will inform disease interventions such as vaccines and antiviral therapies. |
Bats are unusual mammals, with the ability to fly, and long lifespans. In addition, bats have a low incidence of cancer, but the mechanisms underlying this phenomenon remain elusive. Here we discovered that bat cells are more resistant than human and mouse cells to DNA damage induced by genotoxic drugs. We found that bat cells accumulate less chemical than human and mouse cells, and efficient drug efflux mediated by the ABC transporter ABCB1 underlies this improved response to genotoxic reagents. Inhibition of ABCB1 triggers an accumulation of doxorubicin, DNA damage, and cell death. ABCB1 is expressed at higher levels in several cell lines and tissues derived from bats compared to humans. Furthermore, increased drug efflux and high expression of ABCB1 are conserved across multiple bat species. Our findings suggest that enhanced efflux protects bat cells from DNA damage induced by genotoxic compounds, which may contribute to their low cancer incidence. |
INTRODUCTION: Bovine parainfluenza virus-3 (BPIV3) and bovine respiratory syncytial virus (BRSV) are the cause of respiratory disease in cattle worldwide. With other pathogens, they cause bovine respiratory disease complex (BRDC) in ruminants. The aim of the study was the detection and molecular characterisation of BPIV3 and BRSV from nasal swabs and lung samples of cows in and around the Erzurum region of eastern Turkey. MATERIAL AND METHODS: In total, 155 samples were collected. Of animals used in the study 92 were males and 63 females. The age of the animals was between 9 months and 5 years, mean 1.4 years. Most males were in the fattening period and being raised in open sheds; females were in the lactating period and kept in free stall barns. All samples were tested for the presence of viral genes using RT-PCR. Gene-specific primers in a molecular method (RT-PCR) identified BRSV (fusion gene) and BPIV3 (matrix gene) strains at the genus level. RESULTS: RNA from BRSV and BPIV3 was detected in two (1.29%) and three (1.93%) samples, respectively, one of each of which was sequenced and the sequences were aligned with reference virus strains. Phylogenetic analyses clustered the strains in genotype C/BPIV3 and subgroup III/BRSV. CONCLUSION: The results indicate that BRSV and BPIV3 contribute to bovine respiratory disease cases in Turkey. This is the first report on their detection and molecular characterisation in ruminants in Turkey. |
Infections with immunosuppressive pigeon circovirus (PiCV) pose the most severe health problem to the global pigeon breeding. The vaccination with immunogenic PiCV recombinant capsid protein (PiCV rCP) is a potential tool for disease control. Because of the high prevalence of PiCV asymptomatic infections, the subclinically infected pigeons will be vaccinated in practice. The aim of this study was to answer a question if vaccination of asymptomatic, infected with PiCV pigeons induces a similar immune response to PiCV rCP as in uninfected birds. One hundred and twenty 6-week-old carrier pigeons were divided into 4 groups (2 groups of naturally infected and uninfected with PiCV individuals). Birds from groups V and V1 were vaccinated twice with PiCV rCP mixed with an adjuvant, whereas pigeons from groups C and C1 were immunized with an adjuvant only. The expression of genes encoding IFN-γ, CD4, and CD8 T lymphocyte receptors; the number of anti-PiCV rCP IgY-secreting B cells (SBC) and anti-PiCV rCP IgY were evaluated 2, 21, 39 and 46 days post vaccination (dpv). Study results showed that the expression of CD8 and IFN-γ genes was higher in both groups of infected pigeons than in the uninfected birds, irrespective of vaccination. In the uninfected birds, the expression of these genes was insignificantly higher in the vaccinated pigeons. The anti-PiCV rCP IgY-SBC were detected on 2 and 23 dpv and seroconversion was noted on 23 and 39 dpv in V and V1 groups, respectively. In the light of the results obtained, it could be concluded that pigeon circovirus recombinant capsid protein elicits the immune response in both naturally infected and uninfected pigeons, but its rate varies depending on PiCV infectious status. The infection with PiCV masks the potential cellular immune response to the vaccination with PiCV rCP and leads to the suppression of humoral immunity. |
The discovery of the NET network (neutrophil extracellular trap) has revolutionized the perception of defense mechanisms used by neutrophils in infections and non-infectious states, as this mechanism proves the complexity of the ways in which neutrophils can act in the organism. The paper describes the NET network and its participation in bacterial, viral, fungal and parasitic infections, both in a positive and a negative aspect. In addition, attention was paid to the participation of NETs in the course of autoimmune diseases, cancer, as well as its impact on pregnancy and fertility in mammals. |
To probe the effect of 3′,8″-dimerization on antioxidant flavonoids, acacetin and its 3′,8″-dimer isoginkgetin were comparatively analyzed using three antioxidant assays, namely, the ·O(2)(−) scavenging assay, the Cu(2+) reducing assay, and the 2,2′-azino bis(3-ethylbenzothiazolin-6-sulfonic acid) radical scavenging assay. In these assays, acacetin had consistently higher IC(50) values than isoginkgetin. Subsequently, the acacetin was incubated with 4-methoxy-2,2,6,6-tetramethylpiperidine-1-oxy radicals (4-methoxy-TEMPO) and then analyzed by ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC−ESI−Q−TOF−MS) technology. The results of the UHPLC−ESI−Q−TOF−MS analysis suggested the presence of a dimer with m/z 565, 550, 413, 389, 374, 345, 330, and 283 peaks. By comparison, standard isoginkgetin yielded peaks at m/z 565, 533, 518, 489, 401, 389, 374, and 151 in the mass spectra. Based on these experimental data, MS interpretation, and the relevant literature, we concluded that isoginkgetin had higher electron transfer potential than its monomer because of the 3′,8″-dimerization. Additionally, acacetin can produce a dimer during its antioxidant process; however, the dimer is not isoginkgetin. |
Chronic hepatitis B virus (HBV) infection represents a worldwide public health concern with approximately 250 million people chronically infected and at risk of developing liver cirrhosis and hepatocellular carcinoma. Nucleos(t)ide analogues (NUC) are the most widely used therapies for HBV infection, but they often require long-lasting administration to avoid the risk of HBV reactivation at withdrawal. Therefore, there is an urgent need to develop novel treatments to shorten the duration of NUC therapy by accelerating virus control, and to complement the effect of available anti-viral therapies. In chronic HBV infection, virus-specific T cells are functionally defective, and this exhaustion state is a key determinant of virus persistence. Reconstitution of an efficient anti-viral T cell response may thus represent a rational strategy to treat chronic HBV patients. In this perspective, the enhancement of adaptive immune responses by a checkpoint inhibitor blockade, specific T cell vaccines, lymphocyte metabolism targeting, and autologous T cell engineering, including chimeric antigen receptor (CAR) and TCR-redirected T cells, constitutes a promising immune modulatory approach for a therapeutic restoration of protective immunity. The advances of the emerging immune-based therapies in the setting of the HBV research field will be outlined. |
In the USA and other western nations, respiratory syncytial virus is one of the most commonly encountered respiratory viruses among patients who have been diagnosed with a hematologic malignancy or who have undergone a stem cell transplant. Multiple studies have been performed to evaluate the complications associated with respiratory syncytial virus infections. Other studies have evaluated therapeutic agents and strategies in which these agents can be used. There have also been numerous reports of outbreaks in bone marrow transplant units and oncology wards, where infection control measures have been invaluable in controlling the spread of disease. However, despite these novel approaches, respiratory syncytial virus continues to be potentially fatal in immunocompromised populations. In this review, we discuss the incidence of respiratory syncytial viral infections, risk factors associated with progression from upper respiratory tract infection to lower respiratory tract infection, other complications and outcomes (including mortality), management strategies, and prevention strategies in patients with a hematologic malignancy and in hematopoietic cell transplant recipients. |
Since 2015, Zika virus (ZIKV) has caused large epidemics in the Americas. Households are natural targets for control interventions, but quantification of the contribution of household transmission to overall spread is needed to guide policy. We developed a modeling framework to evaluate this contribution and key epidemic features of the ZIKV epidemic in Martinique in 2015–2016 from the joint analysis of a household transmission study (n = 68 households), a study among symptomatic pregnant women (n = 281), and seroprevalence surveys of blood donors (n = 457). We estimated that the probability of mosquito-mediated within-household transmission (from an infected member to a susceptible one) was 21% (95% credible interval (CrI): 5, 51), and the overall probability of infection from outside the household (i.e., in the community) was 39% (95% CrI: 27, 50). Overall, 50% (95% CrI: 43, 58) of the population was infected, with 22% (95% CrI: 5, 46) of infections acquired in households and 40% (95% CrI: 23, 56) being asymptomatic. The probability of presenting with Zika-like symptoms due to another cause was 16% (95% CrI: 10, 23). This study characterized the contribution of household transmission in ZIKV epidemics, demonstrating the benefits of integrating multiple data sets to gain more insight into epidemic dynamics. |
Innate immune interferons (IFNs), particularly type I IFNs, are primary mediators regulating animal antiviral, antitumor, and cell-proliferative activity. These antiviral cytokines have evolved remarkable molecular and functional diversity to confront ever-evolving viral threats and physiological regulation. We have annotated IFN gene families across 110 animal genomes, and showed that IFN genes, after originating in jawed fishes, had several significant evolutionary surges in vertebrate species of amphibians, bats and ungulates, particularly pigs and cattle. For example, pigs have the largest but still expanding type I IFN family consisting of nearly 60 IFN-coding genes that encode seven IFN subtypes including multigene subtypes of IFN-α, -δ, and -ω. Whereas, subtypes such as IFN-α and -β have been widely studied in many species, the unconventional subtypes such as IFN-ω have barely been investigated. We have cross-species defined the IFN evolution, and shown that unconventional IFN subtypes particularly the IFN-ω subtype have evolved several novel features including: (1) being a signature multi-gene subtype expanding primarily in mammals such as bats and ungulates, (2) emerging isoforms that have superior antiviral potency than typical IFN-α, (3) highly cross-species antiviral (but little anti-proliferative) activity exerted in cells of humans and other mammalian species, and (4) demonstrating potential novel molecular and functional properties. This study focused on IFN-ω to investigate the immunogenetic evolution and functional diversity of unconventional IFN subtypes, which may further IFN-based novel antiviral design pertinent to their cross-species high antiviral and novel activities. |
Metagenomics is helping to expand the known diversity of viruses, especially of those with poorly studied hosts in remote areas. The Neotropical region harbors a considerable diversity of avian species that may play a role as both host and short-distance vectors of unknown viruses. Viral metagenomics of cloacal swabs from 50 Neotropical birds collected in French Guiana revealed the presence of four complete astrovirus genomes. They constitute an early diverging novel monophyletic clade within the Avastrovirus phylogeny, representing a putative new astrovirus species (provisionally designated as Avastrovirus 5) according to the International Committee on Taxonomy of Viruses (ICTV) classification criteria. Their genomic organization shares some characteristics with Avastrovirus but also with Mamastrovirus. The pan-astrovirus RT-PCR analysis of the cloacal samples of 406 wild Neotropical birds showed a community-level prevalence of 4.9% (5.1% in passerines, the highest described so far in this order of birds). By screening birds of a remote region, we expanded the known host range of astroviruses to the avian families Cardinalidae, Conopophagidae, Furnariidae, Thamnophilidae, Turdidae and Tyrannidae. Our results provide important first insights into the unexplored viral communities, the ecology, epidemiology and features of host-pathogen interactions that shape the evolution of avastroviruses in a remote Neotropical rainforest. |
The repair of white matter damage is of paramount importance for functional recovery after brain injuries. Here, we report that interleukin-4 (IL-4) promotes oligodendrocyte regeneration and remyelination. IL-4 receptor expression was detected in a variety of glial cells after ischemic brain injury, including oligodendrocyte lineage cells. IL-4 deficiency in knockout mice resulted in greater deterioration of white matter over 14 d after stroke. Consistent with these findings, intranasal delivery of IL-4 nanoparticles after stroke improved white matter integrity and attenuated long-term sensorimotor and cognitive deficits in wild-type mice, as revealed by histological immunostaining, electron microscopy, diffusion tensor imaging, and electrophysiology. The selective effect of IL-4 on remyelination was verified in an ex vivo organotypic model of demyelination. By leveraging primary oligodendrocyte progenitor cells (OPCs), microglia-depleted mice, and conditional OPC-specific peroxisome proliferator-activated receptor gamma (PPARγ) knockout mice, we discovered a direct salutary effect of IL-4 on oligodendrocyte differentiation that was mediated by the PPARγ axis. Our findings reveal a new regenerative role of IL-4 in the central nervous system (CNS), which lies beyond its known immunoregulatory functions on microglia/macrophages or peripheral lymphocytes. Therefore, intranasal IL-4 delivery may represent a novel therapeutic strategy to improve white matter integrity in stroke and other brain injuries. |
Yeast Two-Hybrid (Y2H) and reverse Two-Hybrid (RY2H) are powerful protein–protein interaction screening methods that rely on the interaction of bait and prey proteins fused to DNA binding (DB) and activation domains (AD), respectively. Y2H allows identification of protein interaction partners using screening libraries, while RY2H is used to determine residues critical to a given protein–protein interaction by exploiting site-directed mutagenesis. Currently, both these techniques still rely on sequencing of positive clones using conventional Sanger sequencing. For Y2H, a screen can yield several positives; the identification of such clones is further complicated by the fact that sequencing products usually contain vector sequence. For RY2H, obtaining a complete sequence is required to identify the full range of residues involved in protein–protein interactions. However, with Sanger sequencing limited to 500–800 nucleotides, sequencing is usually carried from both ends for clones greater than this length. Analysis of such RY2H data thus requires assembly of sequencing products combined with trimming of vector sequences and of low-quality bases at the beginning and ends of sequencing products. Further, RY2H analysis requires collation of mutations that abrogate a DB/AD interaction. Here, we present 2HybridTools, a Java program with a user-friendly interface that allows addressing all these issues inherent to both Y2H and RY2H. Specifically, for Y2H, 2HybridTools enables automated identification of positive clones, while for RY2H, 2HybridTools provides detailed mutation reports as a basis for further investigation of given protein–protein interactions. |
Influenza rates for the US Army and West Point cadets showed that seasonal influenza was not necessarily an annual event, and there was little influenzal illness in the decade before 1918 except for 1911 and 1916. Annual records from 1862–1918 also indicated a similar paucity of influenzal illness before 1890. |
Senescence is a cellular phenotype present in health and disease, characterized by a stable cell-cycle arrest and an inflammatory response called senescence-associated secretory phenotype (SASP). The SASP is important in influencing the behavior of neighboring cells and altering the microenvironment; yet, this role has been mainly attributed to soluble factors. Here, we show that both the soluble factors and small extracellular vesicles (sEVs) are capable of transmitting paracrine senescence to nearby cells. Analysis of individual cells internalizing sEVs, using a Cre-reporter system, show a positive correlation between sEV uptake and senescence activation. We find an increase in the number of multivesicular bodies during senescence in vivo. sEV protein characterization by mass spectrometry (MS) followed by a functional siRNA screen identify interferon-induced transmembrane protein 3 (IFITM3) as being partially responsible for transmitting senescence to normal cells. We find that sEVs contribute to paracrine senescence. |
Sepsis involves a disordered host response to systemic infection leading to high morbidity and mortality. Despite intense research, targeted sepsis therapies beyond antibiotics have remained elusive. The cornerstone of sepsis research is the development of animal models to mimic human bacterial infections and test novel pharmacologic targets. Nonhuman primates (NHPs) have served as an attractive, but expensive, animal to model human bacterial infections due to their nearly identical cardiopulmonary anatomy and physiology, as well as host response to infection. Several NHP species have provided substantial insight into sepsis-mediated inflammation, endothelial dysfunction, acute lung injury, and multi-organ failure. The use of NHPs has usually focused on translating therapies from early preclinical models to human clinical trials. However, despite successful sepsis interventions in NHP models, there are still no FDA-approved sepsis therapies. This review highlights major NHP models of bacterial sepsis and their relevance to clinical medicine. |
African swine fever virus (ASFV) is a complex, cytoplasmic double-stranded DNA (dsDNA) virus that is currently expanding throughout the world. Currently, circulating virulent genotype II Armenia/07-like viruses cause fatal disease in pigs and wild boar, whereas attenuated strains induce infections with various levels of chronic illness. Sensing cytosolic dsDNA, mainly by the key DNA sensor cyclic GMP-AMP synthase (cGAS), leads to the synthesis of type I interferon and involves signaling through STING, TBK1, and IRF3. After phosphorylation, STING translocates from the endoplasmic reticulum to the Golgi compartment and to the perinuclear region, acting as an indispensable adaptor connecting the cytosolic detection of DNA to the TBK1-IRF3 signaling pathway. We demonstrate here that attenuated NH/P68, but not virulent Armenia/07, activates the cGAS-STING-IRF3 cascade very early during infection, inducing STING phosphorylation and trafficking through a mechanism involving cGAMP. Both TBK1 and IRF3 are subsequently activated and, in response to this, a high level of beta interferon (IFN-β) was produced during NH/P68 infection; in contrast, Armenia/07 infection generated IFN-β levels below those of uninfected cells. Our results show that virulent Armenia/07 ASFV controls the cGAS-STING pathway, but these mechanisms are not at play when porcine macrophages are infected with attenuated NH/P68 ASFV. These findings show for the first time the involvement of the cGAS-STING-IRF3 route in ASFV infection, where IFN-β production or inhibition was found after infection by attenuated or virulent ASFV strains, respectively, thus reinforcing the idea that ASFV virulence versus attenuation may be a phenomenon grounded in ASFV-mediated innate immune modulation where the cGAS-STING pathway might play an important role. IMPORTANCE African swine fever, a devastating disease for domestic pigs and wild boar, is currently spreading in Europe, Russia, and China, becoming a global threat with huge economic and ecological consequences. One interesting aspect of ASFV biology is the molecular mechanism leading to high virulence of some strains compared to more attenuated strains, which produce subclinical infections. In this work, we show that the presently circulating virulent Armenia/07 virus blocks the synthesis of IFN-β, a key mediator between the innate and adaptive immune response. Armenia/07 inhibits the cGAS-STING pathway by impairing STING activation during infection. In contrast, the cGAS-STING pathway is efficiently activated during NH/P68 attenuated strain infection, leading to the production of large amounts of IFN-β. Our results show for the first time the relationship between the cGAS-STING pathway and ASFV virulence, contributing to uncover the molecular mechanisms of ASFV virulence and to the rational development of ASFV vaccines. |
While impressive clinical responses have been observed using chimeric antigen receptor (CAR) T cells targeting CD19+ hematologic malignancies, limited clinical benefit has been observed using CAR T cells for a variety of solid tumors. Results of clinical studies have highlighted several obstacles which CAR T cells face in the context of solid tumors, including insufficient homing to tumor sites, lack of expansion and persistence, encountering a highly immunosuppressive tumor microenvironment, and heterogeneous antigen expression. In this review, we review clinical outcomes and discuss strategies to improve the antitumor activity of CAR T cells for solid tumors. |
Ribosome-mediated transcription attenuation is a basic posttranscriptional regulation mechanism in bacteria. Liberated attenuator RNAs arising in this process are generally considered nonfunctional. In Sinorhizobium meliloti, the tryptophan (Trp) biosynthesis genes are organized into three operons, trpE(G), ppiD-trpDC-moaC-moeA, and trpFBA-accD-folC, of which only the first one, trpE(G), contains a short ORF (trpL) in the 5′-UTR and is regulated by transcription attenuation. Under conditions of Trp sufficiency, transcription is terminated between trpL and trpE(G), and a small attenuator RNA, rnTrpL, is produced. Here, we show that rnTrpL base-pairs with trpD and destabilizes the polycistronic trpDC mRNA, indicating rnTrpL-mediated downregulation of the trpDC operon in trans. Although all three trp operons are regulated in response to Trp availability, only in the two operons trpE(G) and trpDC the Trp-mediated regulation is controlled by rnTrpL. Together, our data show that the trp attenuator coordinates trpE(G) and trpDC expression posttranscriptionally by two fundamentally different mechanisms: ribosome-mediated transcription attenuation in cis and base-pairing in trans. Also, we present evidence that rnTrpL-mediated regulation of trpDC genes expression in trans is conserved in Agrobacterium and Bradyrhizobium, suggesting that the small attenuator RNAs may have additional conserved functions in the control of bacterial gene expression. |
The political, social, economic, and security implications of health-related issues such as emerging infectious diseases or the epidemic of Non Communicable Diseases offer a rare opportunity for professionals in foreign policy and international relations to engage with the health arena and at the same time for global health experts to enter into and intersect with the domain of diplomacy. The aim of this review is to understand and explore the concepts of global health diplomacy (GHD), health security, and human security. For this narrative review, a literature search was done in PubMed, Scopus, and EBSCO for the “global health diplomacy,” “health security,” and “human security,” and full-texts were reviewed. The recent outbreaks of Ebola in West Africa and Zika in South America are pertinent examples of the nature of the human security crisis and the imminent and severe threat posed to human life across the globe as a result of these epidemics. The Commission on Human Security defines human security as the protection of the vital core of all human lives from critical and pervasive threats. We highlight the ways in which health has now become an issue of national security/global concern and also how GHD can aid in the development of new bilateral or multilateral agreements to safeguard the health and security of people in our globalized world. The paper provides a prospective about, and overview of, health and human security that essentially emphasizes the growing interlinkages between global health, diplomacy, and foreign policy. |
BACKGROUND: Avian infectious bronchitis virus (IBV) is a major respiratory disease-causing agent in birds that leads to significant losses. Dendritic cells (DCs) are specialised cells responsible for sampling antigens and presenting them to T cells, which also play an essential role in recognising and neutralising viruses. Recent studies have suggested that non-coding RNAs may regulate the functional program of DCs. Expression of host non-coding RNAs changes markedly during infectious bronchitis virus infection, but their role in regulating host immune function has not been explored. Here, microarrays of mRNAs, miRNAs, and lncRNAs were globally performed to analyse how avian DCs respond to IBV. RESULTS: First, we found that IBV stimulation did not enhance the maturation ability of avian DCs. Interestingly, inactivated IBV was better able than IBV to induce DC maturation and activate lymphocytes. We identified 1093 up-regulated and 845 down-regulated mRNAs in IBV-infected avian DCs. Gene Ontology analysis suggested that cellular macromolecule and protein location (GO-BP) and transcription factor binding (GO-MF) were abundant in IBV-stimulated avian DCs. Meanwhile, pathway analysis indicated that the oxidative phosphorylation and leukocyte transendothelial migration signalling pathways might be activated in the IBV group. Moreover, alteration of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) was detected in IBV-stimulated avian DCs. In total, 19 significantly altered (7 up and 12 down) miRNAs and 101 (75 up and 26 down) lncRNAs were identified in the IBV-treated group. Further analysis showed that the actin cytoskeleton and MAPK signal pathway were related to the target genes of IBV-stimulated miRNAs. Finally, our study identified 2 TF-microRNA and 53 TF–microRNA–mRNA interactions involving 1 TF, 2 miRNAs, and 53 mRNAs in IBV-stimulated avian DCs. CONCLUSIONS: Our research suggests a new mechanism to explain why IBV actively blocks innate responses needed for inducing immune gene expression and also provides insight into the pathogenic mechanisms of avian IBV. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12864-019-5940-6) contains supplementary material, which is available to authorized users. |
BACKGROUND: T cell immunoglobulin mucin domain-1 (TIM-1) is a phosphatidylserine (PS) receptor, mediating filovirus entry into cells through interactions with PS on virions. TIM-1 expression has been implicated in Ebola virus (EBOV) pathogenesis; however, it remains unclear whether this is due to TIM-1 serving as a filovirus receptor in vivo or, as others have suggested, TIM-1 induces a cytokine storm elicited by T cell/virion interactions. Here, we use a BSL2 model virus that expresses EBOV glycoprotein to demonstrate the importance of TIM-1 as a virus receptor late during in vivo infection. METHODOLOGY/PRINCIPAL FINDINGS: Infectious, GFP-expressing recombinant vesicular stomatitis virus encoding either full length EBOV glycoprotein (EBOV GP/rVSV) or mucin domain deleted EBOV glycoprotein (EBOV GPΔO/rVSV) was used to assess the role of TIM-1 during in vivo infection. GFP-expressing rVSV encoding its native glycoprotein G (G/rVSV) served as a control. TIM-1-sufficient or TIM-1-deficient BALB/c interferon α/β receptor(-/-) mice were challenged with these viruses. While G/rVSV caused profound morbidity and mortality in both mouse strains, TIM-1-deficient mice had significantly better survival than TIM-1-expressing mice following EBOV GP/rVSV or EBOV GPΔO/rVSV challenge. EBOV GP/rVSV or EBOV GPΔO/rVSV in spleen of infected animals was high and unaffected by expression of TIM-1. However, infectious virus in serum, liver, kidney and adrenal gland was reduced late in infection in the TIM-1-deficient mice, suggesting that virus entry via this receptor contributes to virus load. Consistent with higher virus loads, proinflammatory chemokines trended higher in organs from infected TIM-1-sufficient mice compared to the TIM-1-deficient mice, but proinflammatory cytokines were more modestly affected. To assess the role of T cells in EBOV GP/rVSV pathogenesis, T cells were depleted in TIM-1-sufficient and -deficient mice and the mice were challenged with virus. Depletion of T cells did not alter the pathogenic consequences of virus infection. CONCLUSIONS: Our studies provide evidence that at late times during EBOV GP/rVSV infection, TIM-1 increased virus load and associated mortality, consistent with an important role of this receptor in virus entry. This work suggests that inhibitors which block TIM-1/virus interaction may serve as effective antivirals, reducing virus load at late times during EBOV infection. |
Sport training leads to adaptation to physical effort that is reflected by the changes in blood parameters. In equine endurance athletes, blood testing is accepted as a support in training, however, only the changes before versus after exercise in creatine phosphokinase activity (CPK) and basic blood parameters are usually measured. This study is the first longitudinal investigation of the changes in routinely measured blood parameters and, additionally, serum amyloid A (SAA), during seven months, in Arabian horses introduced to endurance training and competing in events for young horses. It has been determined that CPK, aspartate aminotransferase (AST), packed cell volume (PCV), hemoglobin concentration, red blood cell count (RBC), and concentration of total serum protein (TSP) slightly increased after training sessions and competitions in similar manner. The increase in white blood cell (WBC) count was higher after competitions and SAA increased only after competitions. Total protein concentration was the only parameter that increased with training during a 7-month program. SAA indicated only in the case of heavy effort, and, it thus may be helpful in the monitoring of training in young horses. In an optimal program, its concentration should not increase after a training session but only after heavy effort, which should not be repeated too often. |
Differences among hosts, resulting from genetic variation in the immune system or heterogeneity in drug treatment, can impact within-host pathogen evolution. Genetic association studies can potentially identify such interactions. However, extensive and correlated genetic population structure in hosts and pathogens presents a substantial risk of confounding analyses. Moreover, the multiple testing burden of interaction scanning can potentially limit power. We present a Bayesian approach for detecting host influences on pathogen evolution that exploits vast existing data sets of pathogen diversity to improve power and control for stratification. The approach models key processes, including recombination and selection, and identifies regions of the pathogen genome affected by host factors. Our simulations and empirical analysis of drug-induced selection on the HIV-1 genome show that the method recovers known associations and has superior precision-recall characteristics compared to other approaches. We build a high-resolution map of HLA-induced selection in the HIV-1 genome, identifying novel epitope-allele combinations. |
BACKGROUND: Vernal keratoconjunctivitis (VKC) is a common eye disease and can result in permanent decrease or loss of vision. Houttuynia eye drops (HED) is used for the treatment of VKC. However, the clinical evidence of HED has not been well concluded. Herein, we described a proposed systemic review and meta-analysis to evaluate the clinical efficacy of HED for the treatment of VKC. METHODS: Six electronic databases (Medline, Embase, the Cochrane database, Chinese National Knowledge Infrastructure, Wanfang database, and Chinese Biology and Medicine database) will be searched for randomized controlled trials (RCTs) which evaluating the clinical efficacy of HED for the treatment of VKC. Studies meet the eligibility criteria will be included. Data of the included studies will be extracted and the quality will also be evaluated. Data synthesis will be performed using RevMan software. Sensitivity analysis and publication bias will also be investigated. RESULTS: This study will provide high-quality systemic review and synthesis of RCTs on efficacy of HED for the treatment of VKC. CONCLUSION: This systemic review and meta-analysis will conclude the efficacy of HED for the treatment of VKC. REGISTRATION: PROSPERO CRD42019124737 |
BACKGROUND: Garcinia species contain bioactive compounds such as flavonoids, xanthones, triterpernoids, and benzophenones with antibacterial, antifungal, anti-inflammatory, and antioxidant activities. In addition, many of these compounds show interesting biological properties such as anti-human immunodeficiency virus activity. Garcinia parvifolia is used in traditional medicine. Currently, the antiviral activity of G. parvifolia is not known. METHODS: This study was conducted to determine the effects of ethyl acetate (45 L Ea), ethanol (45 L Et), and hexane (45 L H) leaf extracts of G. parvifolia on the infectivity of pseudorabies virus (PrV) in Vero cells. The antiviral effects of the extracts were determined by cytopathic effect (CPE), inhibition, attachment, and virucidal assays. RESULTS: The 50% cytotoxicity concentration (CC(50)) values obtained were 237.5, 555.0, and < 1.25 μg/mL for 45 L Ea, 45 L Et, and 45 L H, respectively. The 45 L Ea showed the greatest viral inhibition potency of 75% at 125 μg/mL. Both 45 L Ea and 45 l Et caused 100% residual viral inhibition at 250 μg/mL. The selectivity index values for 45 L Ea, 45 L Et, and 45 L H were 2.65, 1.75, and 0.10 showing that 45 L Ea had the greatest antiviral activity among the three extracts. CONCLUSION: This study showed that ethyl acetate is the best solvent to be used to obtain extract from G. parvifolia leaves with potent antiviral activities. |
BACKGROUND: The major infectious diseases of hepatitis B has constituted an acute public health challenge in China. An effective and affordable HBV control model is urgently needed. A national project of Community-based Collaborative Innovation HBV (CCI-HBV) demonstration areas has optimized the existing community healthcare resources and obtained initial results in HBV control. METHODS: Based on the existing community healthcare network, CCI-HBV project combined the community health management and health contract signing service for long-staying residents in hepatitis B screening. Moreover, HBV field research strategy was popularized in CCI-HBV areas. After screening, patients with seropositive results were enrolled in corresponding cohorts and received treatment at an early stage. And the uninfected people received medical supports including health education through new media, behavior intervention and HBV vaccinations. In this process, a cloud-based National Information Platform (NIP) was established to collect and store residents’ epidemiological data. In addition, a special quality control team was set up for CCI project. RESULTS: After two rounds of screening, HBsAg positive rate dropped from 5.05% (with 5,173,003 people screened) to 4.57% (with 3,819,675 people screened), while the rate of new HBV infections was 0.28 per 100 person-years in the fixed cohorts of 2,584,322 people. The quality control team completed PPS sampling simultaneously and established the serum sample database with 2,800,000 serum samples for unified testing. CONCLUSIONS: CCI-HBV project has established a large-scale field research to conduct whole-population screening and intervention. We analyzed the HBsAg prevalence and new infection rate of HBV in the fixed population for the epidemic trend and intervention effect. The purpose of CCI-HBV project is to establish and evaluate a practical model of grid management and field strategy, to realize the new goal to control hepatitis B in China. To provide policymakers with a feasible model, our results are directly applicable. TRIAL REGISTRATION: The project was funded by the Major Projects of Science Research for the 11th and 12th five-year plans of China, entitled “The prevention and control of AIDS, viral hepatitis and other major infectious diseases”, Grant Nos. 2009ZX10004901, 2011ZX10004901, 2013ZX10004904, 2014ZX10004007 and 2014ZX10004008. |
Tyrosinase inhibitors improve skin whitening by inhibiting the formation of melanin precursors in the skin. The inhibitory activity of seven phlorotannins (1–7), triphlorethol A (1), eckol (2), 2-phloroeckol (3), phlorofucofuroeckol A (4), 2-O-(2,4,6-trihydroxyphenyl)-6,6′-bieckol (5), 6,8′-bieckol (6), and 8,8′-bieckol (7), from Ecklonia cava was tested against tyrosinase, which converts tyrosine into dihydroxyphenylalanine. Compounds 3 and 5 had IC(50) values of 7.0 ± 0.2 and 8.8 ± 0.1 μM, respectively, in competitive mode, with K(i) values of 8.2 ± 1.1 and 5.8 ± 0.8 μM. Both compounds showed the characteristics of slow-binding inhibitors over the time course of the enzyme reaction. Compound 3 had a single-step binding mechanism and compound 5 a two-step-binding mechanism. With stable AutoDock scores of −6.59 and −6.68 kcal/mol, respectively, compounds 3 and 5 both interacted with His85 and Asn260 at the active site. |
The enzyme kynurenine aminotransferase (KAT) catalyses the conversion of kynurenine (KYN) to kynurenic acid (KYNA). Although the isozymes KAT1–4 have been identified, KYNA is mainly produced by KAT2 in brain tissues. KNYA is an antagonist of N-methyl-D-aspartate and α-7-nicotinic acetylcholine receptors, and accumulation of KYNA in the brain has been associated with the pathology of schizophrenia. Therefore, KAT2 could be exploited as a therapeutic target for the management of schizophrenia. Although currently available KAT2 inhibitors irreversibly bind to pyridoxal 5′-phosphate (PLP), inhibition via this mechanism may cause adverse side effects because of the presence of other PLP-dependent enzymes. Therefore, we identified novel selective KAT2 inhibitors by screening approximately 13,000 molecules. Among these, glycyrrhizic acid (GL) and its analogues, glycyrrhetinic acid (GA) and carbenoxolone (CBX), were identified as KAT2 inhibitors. These compounds were highly selective for KAT2 and competed with its substrate KYN, but had no effects on the other 3 KAT isozymes. Furthermore, we demonstrated that in complex structures that were predicted in docking calculations, GL, GA and CBX were located on the same surface as the aromatic ring of KYN. These results indicate that GL and its analogues are highly selective and competitive inhibitors of KAT2. |
Infectious disease modeling has played a prominent role in recent outbreaks, yet integrating these analyses into public health decision-making has been challenging. We recommend establishing ‘outbreak science’ as an inter-disciplinary field to improve applied epidemic modeling. |
Bats, unique among mammals with powered flight, have many species with the longest size-proportionate lifespan of all mammals. Evolutionary adaptations would have been required to survive the elevated body temperatures during flight. Heat shock protein (HSP), highly conserved master regulators of cell stress, expression was examined across tissues and various cell lines in bats. Basal expression level of major HSPs (HSP70 and HSP90) is significantly higher in two different bat species compared to other mammals. This HSP expression could be a bat-unique, key factor to modulate cellular stress and death. Consequently, bat cells survive prolonged heat treatment, along with other stress stimuli, in a HSP-dependent manner, whereas other mammalian cells succumbed. This suggests HSP expression in bats could be an important adaption to intrinsic metabolic stresses like flight and therefore an important model to study stress resilience and longevity in general. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12192-019-01013-y) contains supplementary material, which is available to authorized users. |
The initial step of retrovirus entry—the interaction between the virus envelope glycoprotein trimer and a cellular receptor—is complex, involving multiple, noncontiguous determinants in both proteins that specify receptor choice, binding affinity and the ability to trigger conformational changes in the viral glycoproteins. Despite the complexity of this interaction, retroviruses have the ability to evolve the structure of their envelope glycoproteins to use a different cellular protein as receptors. The highly homologous subgroup A to E Avian Sarcoma and Leukosis Virus (ASLV) glycoproteins belong to the group of class 1 viral fusion proteins with a two-step triggering mechanism that allows experimental access to intermediate structures during the fusion process. We and others have taken advantage of replication-competent ASLVs and exploited genetic selection strategies to force the ASLVs to naturally evolve and acquire envelope glycoprotein mutations to escape the pressure on virus entry and still yield a functional replicating virus. This approach allows for the simultaneous selection of multiple mutations in multiple functional domains of the envelope glycoprotein that may be required to yield a functional virus. Here, we review the ASLV family and experimental system and the reverse engineering approaches used to understand the evolution of ASLV receptor usage. |
Background: Nebulised medical aerosols are designed to deliver drugs to the lungs to aid in the treatment of respiratory diseases. However, an unintended consequence is the potential for fugitive emissions during patient treatment, which may pose a risk factor in both clinical and homecare settings. Methods: The current study examined the potential for fugitive emissions, using albuterol sulphate as a tracer aerosol during high-flow therapy. A nasal cannula was connected to a head model or alternatively, a interface was connected to a tracheostomy tube in combination with a simulated adult and paediatric breathing profile. Two aerodynamic particle sizers (APS) recorded time-series aerosol concentrations and size distributions at two different distances relative to the simulated patient. Results: The results showed that the quantity and characteristics of the fugitive emissions were influenced by the interface type, patient type and supplemental gas-flow rate. There was a trend in the adult scenarios; as the flow rate increased, the fugitive emissions and the mass median aerodynamic diameter (MMAD) of the aerosol both decreased. The fugitive emissions were comparable when using the adult breathing profiles for the nasal cannula and tracheostomy interfaces; however, there was a noticeable distinction between the two interfaces when compared for the paediatric breathing profiles. The highest recorded aerosol concentration was 0.370 ± 0.046 mg m(−3) from the tracheostomy interface during simulated paediatric breathing with a gas-flow rate of 20 L/min. The averaged MMAD across all combinations ranged from 1.248 to 1.793 µm by the APS at a distance of 0.8 m away from the patient interface. Conclusions: Overall, the results highlight the potential for secondary inhalation of fugitive emissions released during simulated aerosol treatment with concurrent high-flow therapy. The findings will help in developing policy and best practice for risk mitigation from fugitive emissions. |
Picornaviruses are associated with acute and chronic diseases. The clinical manifestations of infections are often mild, but infections may also lead to respiratory symptoms, gastroenteritis, myocarditis, meningitis, hepatitis, and poliomyelitis, with serious impacts on human health and economic losses in animal husbandry. Thus far, research on picornaviruses has mainly focused on structural proteins such as VP1, whereas the non-structural protein 2B, which plays vital roles in the life cycle of the viruses and exhibits a viroporin or viroporin-like activity, has been overlooked. Viroporins are viral proteins containing at least one amphipathic α-helical structure, which oligomerizes to form transmembrane hydrophilic pores. In this review, we mainly summarize recent research data on the viroporin or viroporin-like activity of 2B proteins, which affects the biological function of the membrane, regulates cell death, and affects the host immune response. Considering these mechanisms, the potential application of the 2B protein as a candidate target for antiviral drug development is discussed, along with research challenges and prospects toward realizing a novel treatment strategy for picornavirus infections. |
Adenovirus vectored vaccines are a highly effective strategy to induce cellular immune responses which are particularly effective against intracellular pathogens. Recombinant simian adenovirus vectors were developed to circumvent the limitations imposed by the use of human adenoviruses due to widespread seroprevalence of neutralising antibodies. We have constructed a replication deficient simian adenovirus-vectored vaccine (ChAdOx2) expressing 4 genes from the Mycobacterium avium subspecies paratuberculosis (AhpC, Gsd, p12 and mpa). Safety and T-cell immunogenicity results of the first clinical use of the ChAdOx2 vector are presented here. The trial was conducted using a ‘three-plus-three’ dose escalation study design. We demonstrate the vaccine is safe, well tolerated and immunogenic. |
Influenza D virus (IDV) has first been identified in 2011 in the USA and was shown to mainly circulate in cattle. While IDV is associated with mild respiratory signs, its prevalence is still unknown. In the present study we show that IDV has been circulating throughout France in cattle and small ruminants, with 47.2% and 1.5% seropositivity, respectively. The high prevalence and moderate pathogenicity of IDV in cattle suggest that it may play an initiating role in the bovine respiratory disease complex. |
DNA vaccines present one of the most cost-effective platforms to develop global vaccines, which have been tested for nearly three decades in preclinical and clinical settings with some success in the clinic. However, one of the major challenges for the development of DNA vaccines is their poor immunogenicity in humans, which has led to refinements in DNA delivery, dosage in prime/boost regimens and the inclusion of adjuvants to enhance their immunogenicity. In this review, we focus on adjuvants that can enhance the immunogenicity of DNA encoded antigens and highlight the development of a novel cytolytic DNA platform encoding a truncated mouse perforin. The application of this innovative DNA technology has considerable potential in the development of effective vaccines. |
Seoul virus (SEOV) is a zoonotic orthohantavirus carried by black and brown rats, and can cause hemorrhagic fever with renal syndrome in humans. Human cases of SEOV virus infection have most recently been reported in the USA, United Kingdom, France and the Netherlands and were primarily associated with contact with pet rats and feeder rats. Infection of rats results in an asymptomatic but persistent infection. Little is known about the cell tropism of SEOV in its reservoir and most available data is based on experimental infection studies in which rats were inoculated via a route which does not recapitulate virus transmission in nature. Here we report the histopathological analysis of SEOV cell tropism in key target organs following natural infection of a cohort of feeder rats, comprising 19 adults and 11 juveniles. All adult rats in this study were positive for SEOV specific antibodies and viral RNA in their tissues. One juvenile rat was seropositive, but negative in the rRT-PCR. Of the 19 adult rats of which subsequently additional organs were tested, SEOV RNA was detected in all lungs, followed by kidney (79%) and liver (74%). Histopathologic changes associated with SEOV infection were primarily found in the liver, consistent with a pathological diagnosis of a mild hepatitis. In conclusion, natural SEOV infection results in mild inflammation of the liver in the absence of clinical disease. |
Influenza is a disease that poses a significant health burden worldwide. Vaccination is the best way to prevent influenza virus infections. However, conventional vaccines are only effective for a short period of time due to the propensity of influenza viruses to undergo antigenic drift and antigenic shift. The efficacy of these vaccines is uncertain from year-to-year due to potential mismatch between the circulating viruses and vaccine strains, and mutations arising due to egg adaptation. Subsequently, the inability to store these vaccines long-term and vaccine shortages are challenges that need to be overcome. Conventional vaccines also have variable efficacies for certain populations, including the young, old, and immunocompromised. This warrants for diverse efficacious vaccine developmental approaches, involving both active and passive immunization. As opposed to active immunization platforms (requiring the use of whole or portions of pathogens as vaccines), the rapidly developing passive immunization involves administration of either pathogen-specific or broadly acting antibodies against a kind or class of pathogens as a treatment to corresponding acute infection. Several antibodies with broadly acting capacities have been discovered that may serve as means to suppress influenza viral infection and allow the process of natural immunity to engage opsonized pathogens whilst boosting immune system by antibody-dependent mechanisms that bridge the innate and adaptive arms. By that; passive immunotherapeutics approach assumes a robust tool that could aid control of influenza viruses. In this review, we comment on some improvements in influenza management and promising vaccine development platforms with an emphasis on the protective capacity of passive immunotherapeutics especially when coupled with the use of antivirals in the management of influenza infection. |
The current classification of animal viruses is largely based on the virus molecular world. Less attention is given to why and how virus fitness results from the success of virus transmission. Virus transmission reflects the infection-shedding-transmission dynamics, and with it, the organ system involvement and other, macroscopic dimensions of the host environment. This study describes the transmission ecology of the world main livestock viruses, 36 in total, a mix of RNA, DNA and retroviruses. Following an iterative process, the viruses are virtually ranked in an outer- to inner-body fashion, by organ system, on ecological grounds. Also portrayed are the shifts in virus host tropism and virus genome. The synthesis of the findings reveals a predictive virus evolution framework, based on the outer- to inner-body changes in the interplay of host environment-transmission modes-organ system involvement-host cell infection cycle-virus genome. Outer-body viruses opportunistically respond to the variation in the external environment. For example, respiratory and enteric viruses tend to be associated with poultry and pig mass rearing. Ruminant and equine viruses tend to be more deep-rooted and host-specific, and also establish themselves in the vital inner-body systems. It is concluded that the framework may assist the study of new emerging viruses and pandemic risks. |
The traditional design of effective vaccines for rapidly-evolving pathogens, such as influenza A virus, has failed to provide broad spectrum and long-lasting protection. With low cost whole genome sequencing technology and powerful computing capabilities, novel computational approaches have demonstrated the potential to facilitate the design of a universal influenza vaccine. However, few studies have integrated computational optimization in the design and discovery of new vaccines. Understanding the potential of computational vaccine design is necessary before these approaches can be implemented on a broad scale. This review summarizes some promising computational approaches under current development, including computationally optimized broadly reactive antigens with consensus sequences, phylogenetic model-based ancestral sequence reconstruction, and immunomics to compute conserved cross-reactive T-cell epitopes. Interactions between virus-host-environment determine the evolvability of the influenza population. We propose that with the development of novel technologies that allow the integration of data sources such as protein structural modeling, host antibody repertoire analysis and advanced phylodynamic modeling, computational approaches will be crucial for the development of a long-lasting universal influenza vaccine. Taken together, computational approaches are powerful and promising tools for the development of a universal influenza vaccine with durable and broad protection. |
Picobirnaviruses (PBVs) are mostly found in animal alimentary samples. In this study, among 576 respiratory specimens from 476 mammals and 100 chickens, genogroup I PBVs were detected in three cattle and three monkeys, and a genogroup II PBV-positive sample was collected from one cattle specimen. More than one PBV sequence type was observed in two and one genogroup I PBV-positive samples from cattle and monkeys, respectively. Twenty-four complete/near-complete segments 2 (nine from respiratory and 15 from alimentary samples) from the cattle and monkey genogroup I PBVs and one complete segment 2 from the cattle genogroup II PBV were sequenced. Similar to other studies, the cattle PBVs also showed a high diversity. In contrast, the monkey PBVs observed in this study were clustered into three distinct clades. Within each clade, all the sequences showed >99% amino acid identities. This unique phenomenon is probably due to the fact that monkeys in our locality reside in separated troops with minimal inter-troop contact. |
This review provides a comparison of the theoretical issues and experimental findings for plasmid DNA and mRNA vaccine technologies. While both have been under development since the 1990s, in recent years, significant excitement has turned to mRNA despite the licensure of several veterinary DNA vaccines. Both have required efforts to increase their potency either via manipulating the plasmid DNA and the mRNA directly or through the addition of adjuvants or immunomodulators as well as delivery systems and formulations. The greater inherent inflammatory nature of the mRNA vaccines is discussed for both its potential immunological utility for vaccines and for the potential toxicity. The status of the clinical trials of mRNA vaccines is described along with a comparison to DNA vaccines, specifically the immunogenicity of both licensed veterinary DNA vaccines and select DNA vaccine candidates in human clinical trials. |
Recently, droplet-based microfluidic systems have been widely used in various biochemical and molecular biological assays. Since this platform technique allows manipulation of large amounts of data and also provides absolute accuracy in comparison to conventional bioanalytical approaches, over the last decade a range of basic biochemical and molecular biological operations have been transferred to drop-based microfluidic formats. In this review, we introduce recent advances and examples of droplet-based microfluidic techniques that have been applied in biochemistry and molecular biology research including genomics, proteomics and cellomics. Their advantages and weaknesses in various applications are also comprehensively discussed here. The purpose of this review is to provide a new point of view and current status in droplet-based microfluidics to biochemists and molecular biologists. We hope that this review will accelerate communications between researchers who are working in droplet-based microfluidics, biochemistry and molecular biology. |
Interferon-induced transmembrane proteins (IFITMs) have been shown to strongly affect influenza A virus (IAV) infectivity in tissue culture. Moreover, polymorphisms in IFITM3 have been associated with the severity of the disease in humans. IFITM3 appears to act early in the infection, but its mechanism of action and potential interactions with incoming IAV structures are not yet defined. Here, we visualized endogenous IFITM3 interactions with IAV in the human lung epithelial cell line A549 and in primary human airway epithelial cells employing stimulated emission depletion super-resolution microscopy. By applying an iterative approach for the cluster definition and computational cluster analysis, we found that IFITM3 reorganizes into clusters as IAV infection progresses. IFITM3 cluster formation started at 2-3 h post infection and increased over time to finally coat IAV-containing endosomal vesicles. This IAV-induced phenotype was due to the endosomal recruitment of IFITM3 rather than to an overall increase in the IFITM3 abundance. While the IAV-induced IFITM3 clustering and localization to endosomal vesicles was comparable in primary human airway epithelial cells and the human lung epithelial cell line A549, the endogenous IFITM3 signal was higher in primary cells. Moreover, we observed IFITM3 signals adjacent to IAV-containing recycling endosomes. |
Consisting of nearly 60 functional genes, porcine interferon (IFN)-complex represents an evolutionary surge of IFN evolution in domestic ungulate species. To compare with humans and mice, each of these species contains about 20 IFN functional genes, which are better characterized using the conventional IFN-α/β subtypes as examples. Porcine IFN-complex thus represents an optimal model for studying IFN evolution that resulted from increasing viral pressure during domestication and industrialization. We hypothesize and justify that porcine IFN-complex may extend its functionality in antiviral and immunomodulatory activity due to its superior molecular diversity. Furthermore, these unconventional IFNs could even confer some functional and signaling novelty beyond that of the well-studied IFN-α/β subtypes. Investigations into porcine IFN-complex will further our understanding of IFN biology and promote IFN-based therapeutic designs to confront swine viral diseases. |
Although the role of exogenous small interfering RNA (siRNA) and P-element induced wimpy testis (PIWI)-interacting RNA (piRNA) pathways in mosquito antiviral immunity is increasingly better understood, there is still little knowledge regarding the role of mosquito cellular microRNA (miRNA). Identifying direct interactions between the mosquito miRNAs and the RNA genome of arboviruses and choosing the relevant miRNA candidates to explore resulting antiviral mechanisms are critical. Here, we carried out genomic analyses to identify Aedes aegypti miRNAs that potentially interact with various lineages and genotypes of chikungunya, dengue, and Zika viruses. By using prediction tools with distinct algorithms, several miRNA binding sites were commonly found within different genotypes/and or lineages of each arbovirus. We further analyzed those miRNAs that could target more than one arbovirus, required a low energy threshold to form miRNA-viralRNA (vRNA) complexes, and predicted potential RNA structures using RNAhybrid software. We predicted miRNA candidates that might participate in regulating arboviral replication in Ae. aegypti. Even without any experimental validation, which should be done as a next step, this study can shed further light on the role of miRNA in mosquito innate immunity and targets for future studies. |
Interest in bat-related viruses has increased considerably during the last decade, leading to the discovery of a rising number of new viruses in several bat species. Poxviridae are a large, diverse family of DNA viruses that can infect a wide range of vertebrates and invertebrates. To date, only a few documented detections of poxviruses have been described in bat populations on three different continents (America, Africa, and Australia). These viruses are phylogenetically dissimilar and have diverse clinical impacts on their hosts. Herein, we report the isolation, nearly complete genome sequencing, and annotation of a novel poxvirus detected from an insectivorous bat (Hypsugo savii) in Northern Italy. The virus is tentatively named Hypsugopoxvirus (HYPV) after the bat species from which it was isolated. The nearly complete genome size is 166,600 nt and it encodes 161 genes. Genome analyses suggest that HYPV belongs to the Chordopoxvirinae subfamily, with the highest nucleotide identity (85%) to Eptesipoxvirus (EPTV) detected from a microbat Eptesicus fuscus in WA, USA, in 2011. To date, HYPV represents the first poxvirus detected in bats in Europe; thus, its viral ecology and disease associations should be investigated further. |
Infectious diseases are linked to 15%–20% of cancers worldwide. Among them, Epstein–Barr virus (EBV) is an oncogenic herpesvirus that chronically infects over 90% of the adult population, with over 200,000 cases of cancer and 150,000 cancer-related deaths attributed to it yearly. Acute EBV infection can present as infectious mononucleosis, and lead to the future onset of multiple cancers, including Burkitt lymphoma, Hodgkin lymphoma, nasopharyngeal carcinoma, and gastric carcinoma. Many of these cancers express latent viral genes, including Epstein–Barr virus nuclear antigen 1 (EBNA1) and latent membrane proteins 1 and 2 (LMP1 and LMP2). Previous attempts to create potent immunogens against EBV have been reported but generated mixed success. We designed novel Synthetic Consensus (SynCon) DNA vaccines against EBNA1, LMP1 and LMP2 to improve on the immune potency targeting important antigens expressed in latently infected cells. These EBV tumor antigens are hypothesized to be useful targets for potential immunotherapy of EBV-driven cancers. We optimized the genetic sequences for these three antigens, studied them for expression, and examined their immune profiles in vivo. We observed that these immunogens generated unique profiles based on which antigen was delivered as the vaccine target. EBNA1vax and LMP2Avax generated the most robust T cell immunity. Interestingly, LMP1vax was a very weak immunogen, generating very low levels of CD8 T cell immunity both as a standalone vaccine and as part of a trivalent vaccine cocktail. LMP2Avax was able to drive immunity that impacted EBV-antigen-positive tumor growth. These studies suggest that engineered EBV latent protein vaccines deserve additional study as potential agents for immunotherapy of EBV-driven cancers. |
Symbiotic microbes impact the function and development of the central nervous system (CNS); however, little is known about the contribution of the microbiota during viral-induced neurologic damage. We identify that commensals aid in host defense following infection with a neurotropic virus through enhancing microglia function. Germfree mice or animals that receive antibiotics are unable to control viral replication within the brain leading to increased paralysis. Microglia derived from germfree or antibiotic-treated animals cannot stimulate viral-specific immunity and microglia depletion leads to worsened demyelination. Oral administration of toll-like receptor (TLR) ligands to virally infected germfree mice limits neurologic damage. Homeostatic activation of microglia is dependent on intrinsic signaling through TLR4, as disruption of TLR4 within microglia, but not the entire CNS (excluding microglia), leads to increased viral-induced clinical disease. This work demonstrates that gut immune-stimulatory products can influence microglia function to prevent CNS damage following viral infection. |
The hypogravity motor syndrome (HMS) is one of the deleterious impacts of weightlessness on the human body in orbital space missions. There is a hypothesis that disorders of musculoskeletal system as part of HMS arise in consequence of changes in spinal motor neurons. The study was aimed at bioinformatic analysis of transcriptome changes in lumbar spinal cords of mice after a 30-day spaceflight aboard biosatellite Bion-M1 (space group, S) and subsequent 7-day readaptation to the Earth’s gravity (recovery group, R) when compared with control mice (C group) housed in simulated biosatellite conditions on the Earth. Gene ontology and human phenotype ontology databases were used to detect biological processes, molecular functions, cellular components, and human phenotypes associated with HMS. Our results suggest resemblance of molecular changes developing in space orbit and during the postflight recovery to terrestrial neuromuscular disorders. Remarkably, more prominent transcriptome changes were revealed in R vs. S and R vs. C comparisons that are possibly related to the 7-day recovery period in the Earth’s gravity condition. These data may assist with establishment of HMS pathogenesis and proposing effective preventive and therapeutic options. |
A key step in the Ebola virus (EBOV) replication cycle involves conformational changes in viral glycoprotein 2 (GP2) which facilitate host-viral membrane fusion and subsequent release of the viral genome. Ebola GP2 plays a critical role in virus entry and has similarities in mechanism and structure to the HIV gp41 protein for which inhibitors have been successfully developed. In this work, a putative binding pocket for the C-terminal heptad repeat in the N-terminal heptad repeat trimer was targeted for identification of small molecules that arrest EBOV-host membrane fusion. Two computational structure-based virtual screens of ∼1.7 M compounds were performed (DOCK program) against a GP2 five-helix bundle, resulting in 165 commercially available compounds purchased for experimental testing. Based on assessment of inhibitory activity, cytotoxicity, and target specificity, four promising candidates emerged with 50% inhibitory concentration values in the 3 to 26 μM range. Molecular dynamics simulations of the two most potent candidates in their DOCK-predicted binding poses indicate that the majority of favorable interactions involve seven highly conserved residues that can be used to guide further inhibitor development and refinement targeting EBOV. IMPORTANCE The most recent Ebola virus disease outbreak, from 2014 to 2016, resulted in approximately 28,000 individuals becoming infected, which led to over 12,000 causalities worldwide. The particularly high pathogenicity of the virus makes paramount the identification and development of promising lead compounds to serve as inhibitors of Ebola infection. To limit viral load, the virus-host membrane fusion event can be targeted through the inhibition of the class I fusion glycoprotein of Ebolavirus. In the current work, several promising small-molecule inhibitors that target the glycoprotein GP2 were identified through systematic application of structure-based computational and experimental drug design procedures. |
The use of Internet-based systems for infectious disease surveillance has been increasingly explored in recent years. However, few studies have used Internet search query or social media data to monitor spatial and temporal trends of avian influenza in China. This study investigated the potential of using search query and social media data in detecting and monitoring avian influenza A (H7N9) cases in humans in China. We collected weekly data on laboratory-confirmed H7N9 cases in humans, as well as H7N9-related Baidu Search Index (BSI) and Weibo Posting Index (WPI) data in China from 2013 to 2017, to explore the spatial and temporal trends of H7N9 cases and H7N9-related Internet search queries. Our findings showed a positive relationship of H7N9 cases with BSI and WPI search queries spatially and temporally. The outbreak threshold time and peak time of H7N9-related BSI and WPI searches preceded H7N9 cases in most years. Seasonal autoregressive integrated moving average (SARIMA) models with BSI (β = 0.008, p < 0.001) and WPI (β = 0.002, p = 0.036) were used to predict the number of H7N9 cases. Regression tree model analysis showed that the average H7N9 cases increased by over 2.4-fold (26.8/11) when BSI for H7N9 was > = 11524. Both BSI and WPI data could be used as indicators to develop an early warning system for H7N9 outbreaks in the future. |
BACKGROUND: Angiotensin‐converting enzyme 2 (ACE2) is a homologue of angiotensin‐converting enzyme (ACE) and produces angiotensin peptides (APs), such as angiotensin 1‐9 and 1‐7 that are vasodilatory and natriuretic, and act to counterbalance angiotensin II. HYPOTHESIS: Evidence of ACE2 can be found in tissues and plasma of dogs. Equilibrium concentrations of renin angiotensin aldosterone system (RAAS) APs differ in dogs with heart disease compared to healthy dogs and recombinant human ACE2 (rhACE2) alters relative concentrations of APs. ANIMALS: Forty‐nine dogs with and 34 dogs without heart disease. METHODS: Immunohistochemistry and assays for tissue and plasma ACE2 activity and equilibrium concentrations of plasma RAAS APs were performed. RESULTS: Immunolabeling for ACE2 was present in kidney and myocardial tissue. Median plasma ACE2 activity was significantly increased in dogs with congestive heart failure (CHF; 6.9 mU/mg; interquartile range [IQR], 5.1‐12.1) as compared to control (2.2 mU/mg; IQR, 1.8‐3.0; P = .0003). Plasma equilibrium analysis of RAAS APs identified significant increases in the median concentrations of beneficial APs, such as angiotensin 1‐7, in dogs with CHF (486.7 pg/mL; IQR, 214.2‐1168) as compared to those with preclinical disease (41.0 pg/mL; IQR, 27.4‐45.1; P < .0001) or control (11.4 pg/mL; IQR, 7.1‐25.3; P = .01). Incubation of plasma samples from dogs with CHF with rhACE2 increased beneficial APs, such as angiotensin 1‐9 (preincubation, 10.3 pg/mL; IQR, 4.4‐37.2; postincubation, 2431 pg/mL; IQR, 1355‐3037; P = .02), while simultaneously decreasing maladaptive APs, such as angiotensin II (preincubation, 53.4 pg/mL; IQR, 28.6‐226.4; postincubation, 2.4 pg/mL; IQR, 0.50‐5.8; P = .02). CONCLUSIONS AND CLINICAL IMPORTANCE: Recognition of the ACE2 system expands the conventional view of the RAAS in the dog and represents an important potential therapeutic target. |
BACKGROUND: Salmonellosis is a major cause of morbidity and mortality in neonatal calves, often occurring before preventative vaccines can be administered. HYPOTHESIS/OBJECTIVE: To evaluate the protective effect on calves of colostrum from cows vaccinated with a commercially available Salmonella Newport bacterin against a Salmonella Typhimurium challenge. ANIMALS: Twenty Holstein bull calves from a university dairy farm. METHODS: Nonrandomized placebo‐controlled trial in which colostrum was harvested from 30 cows that received 2 doses of either Salmonella bacterin or saline before calving. Colostrum collected from each group was pooled and fed to 2 groups of 10 calves at birth. At approximately 2 weeks of age, calves were challenged with Salmonella Typhimurium. Clinical, hematologic, microbiological, and postmortem findings were compared between the 2 groups. RESULTS: No differences in mortality, clinical findings, hematology results, blood and fecal cultures, or necropsy findings between the 2 groups were observed. Vaccinated cows had higher colostral titers, and calves fed this colostrum had higher serum titers (mean difference, 0.429; mean [SE], 0.852 [0.02] for vaccinated versus 0.423 [0.02] for control calves). CONCLUSIONS AND CLINICAL IMPORTANCE: Transfer of colostral immunoglobulins from Salmonella enterica serotype Newport bacterin to neonatal calves was not sufficient to decrease mortality, clinical signs, sepsis, intestinal damage, or fecal shedding when exposed to a highly pathogenic Salmonella isolate. A large‐scale randomized controlled clinical trial is needed to evaluate the efficacy of this bacterin when administered in the dry period for prevention of salmonellosis in neonatal calves. |
Interferons orchestrate host antiviral responses in jawed vertebrates. They are categorized into three classes; IFN1 and IFN3 are the primary antiviral cytokine lineages, while IFN2 responds to a broader variety of pathogens. The evolutionary relationships within and between these three classes have proven difficult to resolve. Here, we reassess interferon evolution, considering key phylogenetic pitfalls including taxon sampling, alignment quality, model adequacy, and outgroup choice. We reveal that cartilaginous fishes, and hence the jawed vertebrate ancestor, possess(ed) orthologs of all three interferon classes. We show that IFN3 groups sister to IFN1, resolve the origins of the human IFN3 lineages, and find that intronless IFN3s emerged at least three times. IFN2 genes are highly conserved, except for IFN-γ-rel, which we confirm resulted from a teleost-specific duplication. Our analyses show that IFN1 phylogeny is highly sensitive to phylogenetic error. By accounting for this, we describe a new backbone IFN1 phylogeny that implies several IFN1 genes existed in the jawed vertebrate ancestor. One of these is represented by the intronless IFN1s of tetrapods, including mammalian-like repertoires of reptile IFN1s and a subset of amphibian IFN1s, in addition to newly-identified intron-containing shark IFN1 genes. IFN-f, previously only found in teleosts, likely represents another ancestral jawed vertebrate IFN1 family member, suggesting the current classification of fish IFN1s into two groups based on the number of cysteines may need revision. The providence of the remaining fish IFN1s and the coelacanth IFN1s proved difficult to resolve, but they may also be ancestral jawed vertebrate IFN1 lineages. Finally, a large group of amphibian-specific IFN1s falls sister to all other IFN1s and was likely also present in the jawed vertebrate ancestor. Our results verify that intronless IFN1s have evolved multiple times in amphibians and indicate that no one-to-one orthology exists between mammal and reptile IFN1s. Our data also imply that diversification of the multiple IFN1s present in the jawed vertebrate ancestor has occurred through a rapid birth-death process, consistent with functional maintenance over a 450-million-year host-pathogen arms race. In summary, this study reveals a new model of interferon evolution important to our understanding of jawed vertebrate antiviral immunity. |
Sepsis is a life-threatening condition caused by an immune response triggered by infection, and highly elevated cytokine/chemokine levels in the blood play crucial roles in the progression of sepsis. Serum exosomes are nanovesicles that have multiple biological functions, playing roles in antigen presentation, intercellular signal communication, inflammatory response and immune surveillance. However, the biological functions and related molecular bases remain to be elucidated. In this study, we investigated the profiles of cytokines/chemokines harbored in the exosomes of septic mice and explored the mechanisms of immunomodulation on T cells treated with exosomes harvested from septic mice. Blood cytokines/chemokines existed in both the soluble form and in the insoluble exosomal form; the profiles of the cytokines/chemokines in these two forms displayed different dynamics in the blood of mice challenged with LPS. Exosomes from septic mice induced the differentiation of Th1/Th2 cells, which was blocked by specific antibodies targeting IL-12 and IL-4. In addition, these exosomes significantly augmented the proliferation and migration of T lymphocytes. Furthermore, preadministration of exosomes by intravenous injection restrained the inflammatory response, attenuated lung and liver tissue damage, and prolonged the survival of cecal ligation and puncture (CLP) mice. Our results indicate that exosomes enriched with cytokines/chemokines play critical roles in T cell differentiation, proliferation and chemotaxis during the sepsis process and have a protective effect on cecal ligation and puncture (CLP) mice. Thus, these findings not only strengthen our understanding of the role of sepsis via exosomes but also provide potential targets for therapeutic applications. |
Duchenne muscular dystrophy (DMD) is an X-linked disorder characterized by progressive muscle degeneration, caused by the absence of dystrophin. Exon skipping by antisense oligonucleotides (ASOs) has recently gained recognition as therapeutic approach in DMD. Conjugation of a peptide to the phosphorodiamidate morpholino backbone (PMO) of ASOs generated the peptide-conjugated PMOs (PPMOs) that exhibit a dramatically improved pharmacokinetic profile. When tested in animal models, PPMOs demonstrate effective exon skipping in target muscles and prolonged duration of dystrophin restoration after a treatment regime. Herein we summarize the main pathophysiological features of DMD and the emergence of PPMOs as promising exon skipping agents aiming to rescue defective gene expression in DMD and other neuromuscular diseases. The listed PPMO laboratory findings correspond to latest trends in the field and highlight the obstacles that must be overcome prior to translating the animal-based research into clinical trials tailored to the needs of patients suffering from neuromuscular diseases. |
[Image: see text] Ecofriendly N-heterocyclic carbene (NHC) organocatalysis can control the N(1)-functionalization (aza-Michael addition) and C(3)-functionalization (Morita–Baylis–Hillman reaction, MBH) of isatins in the absence of (1) a protecting group, (2) a stoichiometric reagent, and (3) heat energy. The challengeable N(1)-functionalization of N-unsubstituted isatins into N-substituted (NS) isatins was realized through 10 mol % NHC and 10 mol % 1,8-diazabicyclo[5.4.0]undec-7-ene catalysts within 10 min with up to 98% isolation yield. The subsequent MBH adducts of as-synthesized NS-isatins (N(1)/C(3)-functionalization) was perfectly acquired in 10 mol % NHC and 10 mol % 1,4-diazabicyclo[2.2.2]octane catalysis within 30 min with superiority to C(3)/N(1)-functionalization (MBH/aza-Michael). For guiding the application to a versatile druggable isatin library, the NHC catalysis was compared with reported functionalization of isatins in view of green chemistry principles including solvent scoring of ACS GCI pharmaceutical roundtable, E-factor, atom economy, and so on. |
Viral pathogens account for a significant proportion of the burden of emerging infectious diseases in humans. The Wellcome Trust-Vietnamese Initiative on Zoonotic Infections (WT-VIZIONS) is aiming to understand the circulation of viral zoonotic pathogens in animals that pose a potential risk to human health. Evidence suggests that human exposure and infections with hepatitis E virus (HEV) genotypes (GT) 3 and 4 results from zoonotic transmission. Hypothesising that HEV GT3 and GT4 are circulating in the Vietnamese pig population and can be transmitted to humans, we aimed to estimate the seroprevalence of HEV exposure in a population of farmers and the general population. We additionally performed sequence analysis of HEV in pig populations in the same region to address knowledge gaps regarding HEV circulation and to evaluate if pigs were a potential source of HEV exposure. We found a high prevalence of HEV GT3 viral RNA in pigs (19.1% in faecal samples and 8.2% in rectal swabs) and a high HEV seroprevalence in pig farmers (16.0%) and a hospital-attending population (31.7%) in southern Vietnam. The hospital population was recruited as a general-population proxy even though this particular population subgroup may introduce bias. The detection of HEV RNA in pigs indicates that HEV may be a zoonotic disease risk in this location, although a larger sample size is required to infer an association between HEV positivity in pigs and seroprevalence in humans. |
We present our experience in haploidentical stem cell transplantation (haplo SCT) in children with benign disorders. We performed a retrospective study where children aged up to 18 years diagnosed to have benign disorders and underwent haplo SCT from 2002 to September 2017 were included. Of the 54 children, the most common indications were Fanconi anaemia 12 (22%), severe aplastic anaemia 8 (14%) and primary immune deficiency disorders (PID) 25 (46%). Post-transplant cyclophosphamide (PTCy) was used in 41 (75.9%) and ex vivo T depletion in 13 (24.1%). Engraftment rates were 70% with acute graft versus host disease in 36% and cytomegalovirus reactivation in 55% children. There was a statistically significant difference found between survival with siblings as donors as compared to parents (p value 0.018). Overall survival was 60% which is the 1-year survival, with 68% survival among those with PIDs. Cytokine release syndrome was noted in 12/41 (29%) of children who received T replete graft and PTCy. In children over 6 months of age, PTCy at a cost of INR 1200 provides cost effective T cell depletion comparable with TCR α/β depletion priced at INR 1200,000. Haplo SCT is feasible option for cure in children with benign disorder. |
DOCK2 is a guanine-nucleotide-exchange factor for Rac proteins. Activated Rac serves various cellular functions including the reorganization of the actin cytoskeleton in lymphocytes and neutrophils and production of reactive oxygen species in neutrophils. Since 2015, six unrelated patients with combined immunodeficiency and early-onset severe viral infections caused by bi-allelic loss-of-function mutations in DOCK2 have been described. Until now, the function of phagocytes, specifically neutrophils, has not been assessed in human DOCK2 deficiency. Here, we describe a new kindred with four affected siblings harboring a homozygous splice-site mutation (c.2704-2 A > C) in DOCK2. The mutation results in alternative splicing and a complete loss of DOCK2 protein expression. The patients presented with leaky severe combined immunodeficiency or Omenn syndrome. The novel mutation affects EBV-B cell migration and results in NK cell dysfunction similar to previous observations. Moreover, both cytoskeletal rearrangement and reactive oxygen species production are partially impaired in DOCK2-deficient neutrophils. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10875-019-00603-w) contains supplementary material, which is available to authorized users. |
ABSTRACT: Excessive activities of cysteinyl cathepsins (CysCts) contribute to the progress of many diseases; however, therapeutic inhibition has been problematic. Zn(2+) is a natural inhibitor of proteases with CysHis dyads or CysHis(Xaa) triads. Biguanide forms bidentate metal complexes through the two imino nitrogens. Here, it is discussed that phenformin (phenylethyl biguanide) is a model for recruitment of endogenous Zn(2+) to inhibit CysHis/CysHis(X) peptidolysis. Phenformin is a Zn(2+)-interactive, anti-proteolytic agent in bioassay of living tissue. Benzoyl-l-arginine amide (BAA) is a classical substrate of papain-like proteases; the amide bond is scissile. In this review, the structures of BAA and the phenformin-Zn(2+) complex were compared in silico. Their chemistry and dimensions are discussed in light of the active sites of papain-like proteases. The phenyl moieties of both structures bind to the “S2” substrate-binding site that is typical of many proteases. When the phenyl moiety of BAA binds to S2, then the scissile amide bond is directed to the position of the thiolate-imidazolium ion pair, and is then hydrolyzed. However, when the phenyl moiety of phenformin binds to S2, then the coordinated Zn(2+) is directed to the identical position; and catalysis is inhibited. Phenformin stabilizes a “Zn(2+) sandwich” between the drug and protease active site. Hundreds of biguanide derivatives have been synthesized at the 1 and 5 nitrogen positions; many more are conceivable. Various substituent moieties can register with various arrays of substrate-binding sites so as to align coordinated Zn(2+) with catalytic partners of diverse proteases. Biguanide is identified here as a modifiable pharmacophore for synthesis of therapeutic CysCt inhibitors with a wide range of potencies and specificities. GRAPHICAL ABSTRACT: Phenformin-Zn(2+) Complex [Image: see text] |
[Image: see text] The 3D structures of biomolecules determine their biological function. Established methods in biomolecule structure determination typically require purification, crystallization, or modification of target molecules, which limits their applications for analyzing trace amounts of biomolecules in complex matrices. Here, we developed instruments and methods of mobility capillary electrophoresis (MCE) and its coupling with MS for the 3D structural analysis of biomolecules in the liquid phase. Biomolecules in complex matrices could be separated by MCE and sequentially detected by MS. The effective radius and the aspect ratio of each separated biomolecule were simultaneously determined through the separation by MCE, which were then used as restraints in determining biomolecule conformations through modeling. Feasibility of this method was verified by analyzing a mixture of somatostatin and bradykinin, two peptides with known liquid-phase structures. Proteins could also be structurally analyzed using this method, which was demonstrated for lysozyme. The combination of MCE and MS for complex sample analysis was also demonstrated. MCE and MCE–MS would allow us to analyze trace amounts of biomolecules in complex matrices, which has the potential to be an alternative and powerful biomolecule structure analysis technique. |
We characterized natural vertical transmission of Zika virus in pools of Aedes aegypti larvae hatched from eggs collected in Jojutla, Morelos, Mexico. Of the 151 pools analyzed, 17 tested positive for Zika virus RNA; infectious Zika virus was successfully isolated from 1 of the larvae pools (31N) in C6/36 cells. Real-time quantitative PCR and indirect immunofluorescence assays confirmed the identity of the isolate, named Zika virus isolate 31N; plaque assays in Vero cells demonstrated the isolate’s infectivity in a mammalian cell line. We obtained the complete genome of Zika virus isolate 31N by next-generation sequencing and identified 3 single-nucleotide variants specific to Zika virus isolate 31N using the meta-CATS tool. These results demonstrate the occurrence of natural vertical transmission of Zika virus in wild Ae. aegypti mosquitoes and suggest that this transmission mode could aid in the spread and maintenance of Zika virus in nature. |
Climate change and anthropogenic activity are currently driving large changes in nutritional availability across ecosystems, with consequences for infectious disease. An increase in host nutrition could lead to more resources for hosts to expend on the immune system or for pathogens to exploit. In this paper, we report a meta-analysis of studies on host–pathogen systems across the tree of life, to examine the impact of host nutritional quality and quantity on pathogen virulence. We did not find broad support across studies for a one-way effect of nutrient availability on pathogen virulence. We thus discuss a hypothesis that there is a balance between the effect of host nutrition on the immune system and on pathogen resources, with the pivot point of the balance differing for vertebrate and invertebrate hosts. Our results suggest that variation in nutrition, caused by natural or anthropogenic factors, can have diverse effects on infectious disease outcomes across species. |
Guanine (G)-quadruplexes (G4s) are unique nucleic acid structures that are formed by stacked G-tetrads in G-rich DNA or RNA sequences. G4s have been reported to play significant roles in various cellular events in both macro- and micro-organisms. The identification and characterization of G4s can help to understand their different biological roles and potential applications in diagnosis and therapy. In addition to biophysical and biochemical methods to interrogate G4 formation, G4 fluorescent turn-on ligands can be used to target and visualize G4 formation both in vitro and in cells. Here, we review several representative classes of G4 fluorescent turn-on ligands in terms of their interaction mechanism and application perspectives. Interestingly, G4 structures are commonly identified in DNA and RNA aptamers against targets that include proteins and small molecules, which can be utilized as G4 tools for diverse applications. We therefore also summarize the recent development of G4-containing aptamers and highlight their applications in biosensing, bioimaging, and therapy. Moreover, we discuss the current challenges and future perspectives of G4 fluorescent turn-on ligands and G4-containing aptamers. |
Longstanding discordances and enigmas persist as to the specificities and other properties of antibodies (Abs) most effective in preventing or limiting many viral infections in mammals; in turn, failure to decipher key complexities has added to headwinds for both Ab-based therapeutic approaches and rational vaccine design. More recently, experimental approaches have emerged—and continue to emerge—for discerning the functional role of Ab structure, especially the Fc portion of antibody, in combating viral infections in vivo. A wide range of in vitro measures of antibody activity, from neutralization to antibody-dependent cell mediated cytotoxicity (ADCC)—each of these terms representing only an operational notion defined by the particulars of a given assay—are poised for assignment of both relevance and reliability in forecasting outcomes of infection. Of the several emergent technical opportunities for clarity, attention here is drawn to three realms: the increasing array of known modifications that can be engineered into Abs to affect their in vivo activities; the improvement of murine models involving knockouts and knock-ins of host genes including Fc receptors; and the development of additional virological design tools to differentiate Abs that act primarily by inhibiting viral entry from antibodies that mainly target viral antigens (Ags) on cell surfaces. To illustrate some of the opportunities with either zoonotic (emerging, spillover) or ancient human-adapted viruses, we draw examples from a wide range of viruses that affect humans. |
Manipulating cell cycle is one of the common strategies used by viruses to generate favorable cellular environment to facilitate viral replication. Coxsackievirus B (CVB) is one of the major viral pathogens of human myocarditis and cardiomyopathy. Because of its small genome, CVB depends on cellular machineries for productive replication. However, how the structural and non-structural components of CVB would manipulate cell cycle is not clearly understood. In this study, we demonstrated that the capsid protein VP1 of CVB type 3 (CVB3) induced cell cycle arrest at G1 phase. G1 arrest was the result of the decrease level of cyclin E and the accumulation of p27(Kip1). Study on the gene expression profile of the cells expressing VP1 showed that the expression of both heat shock protein 70-1 (Hsp70-1) and Hsp70-2 was significantly up-regulated. Knockdown of Hsp70 resulted in the increased level of cyclin E and the reduction of p27(Kip1). We further demonstrated that the phosphorylation of the heat shock factor 1, which directly promotes the expression of Hsp70, was also increased in the cell expressing VP1. Moreover, we show that CVB3 infection also induced G1 arrest, likely due to dysregulating Hsp70, cyclin E, and p27, while knockdown of Hsp70 dramatically inhibited viral replication. Cell cycle arrest at G1 phase facilitated CVB3 infection, since viral replication in the cells synchronized at G1 phase dramatically increased. Taken together, this study demonstrates that the VP1 of CVB3 induces cell cycle arrest at G1 phase through up-regulating Hsp70. Our findings suggest that the capsid protein VP1 of CVB is capable of manipulating cellular activities during viral infection. |
Extracellular vesicles (EVs) are thought to be important in cell-cell communication and have elicited extraordinary interest as potential biomarkers of disease. However, quantitative methods to enable elucidation of mechanisms underlying release are few. Here, we describe a cell-based assay for monitoring EV release using the EV-enriched tetraspanin CD63 fused to the small, ATP-independent reporter enzyme, Nanoluciferase. Release of CD63-containing EVs from stably expressing cell lines was monitored by comparing luciferase activity in culture media to that remaining in cells. HEK293, U2OS, U87 and SKMel28 cells released 0.3%-0.6% of total cellular CD63 in the form of EVs over 5 hrs, varying by cell line. To identify cellular machinery important for secretion of CD63-containing EVs, we performed a screen of biologically active chemicals in HEK293 cells. While a majority of compounds did not significantly affect EV release, treating cells with the plecomacrolides bafilomycin or concanamycin, known to inhibit the V-ATPase, dramatically increased EV release. Interestingly, alkalization of the endosomal lumen using weak bases had no effect, suggesting a pH-independent enhancement of EV release by V-ATPase inhibitors. The ability to quantify EVs in small samples will enable future detailed studies of release kinetics as well as further chemical and genetic screening to define pathways involved in EV secretion. |
BACKGROUND: Information on the clinical, epidemiological and molecular characterization of human metapneumovirus in critically ill adult patients with severe community-acquired pneumonia (CAP) and the role of biomarkers identifying bacterial coinfection is scarce. METHODS: This is a retrospective epidemiological study of adult patients with hMPV severe CAP admitted to ICU during a ten-year period with admission PSI score ≥ 3. RESULTS: The 92.8% of the 28 patients with severe CAP due to human metapneumovirus were detected during the first half of the year. Median age was 62 years and 60.7% were male. The genotyping of isolated human metapneumovirus showed group B predominance (60.7%). All patients had acute respiratory failure. Median APACHE II and SOFA score were 13 and 6.55, respectively. The 25% were coinfected with Streptococcus pneumoniae. 60.7% of the patients had shock at admission and 50% underwent mechanical ventilation. Seven patients developed ARDS, three of them younger than 60 years and without comorbidities. Mortality in ICU was 14.3%. Among survivors, ICU and hospital stay were 6.5 and 14 days, respectively. Plasma levels of procalcitonin were higher in patients with bacterial coinfection (18.2 vs 0.54; p < 0.05). The levels of C-reactive protein, however, were similar. CONCLUSION: Human metapneumovirus was associated with severe CAP requiring ICU admission among elderly patients or patients with comorbidities, but also in healthy young subjects. These patients often underwent mechanical ventilation with elevated health resource consumption. While one out of four patients showed pneumococcal coinfection, plasma procalcitonin helped to implement antimicrobial stewardship. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s13613-019-0559-y) contains supplementary material, which is available to authorized users. |
BACKGROUND: There is a need to better distinguish viral infections from antibiotic-requiring bacterial infections in children presenting with clinical community-acquired pneumonia (CAP) to assist health care workers in decision making and to improve the rational use of antibiotics. OBJECTIVE: The overall aim of the Trial of Respiratory infections in children for ENhanced Diagnostics (TREND) study is to improve the differential diagnosis of bacterial and viral etiologies in children aged below 5 years with clinical CAP, by evaluating myxovirus resistance protein A (MxA) as a biomarker for viral CAP and by evaluating an existing (multianalyte point-of-care antigen detection test system [mariPOC respi] ArcDia International Oy Ltd.) and a potential future point-of-care test for respiratory pathogens. METHODS: Children aged 1 to 59 months with clinical CAP as well as healthy, hospital-based, asymptomatic controls will be included at a pediatric emergency hospital in Stockholm, Sweden. Blood (analyzed for MxA and C-reactive protein) and nasopharyngeal samples (analyzed with real-time polymerase chain reaction as the gold standard and antigen-based mariPOC respi test as well as saved for future analyses of a novel recombinase polymerase amplification–based point-of-care test for respiratory pathogens) will be collected. A newly developed algorithm for the classification of CAP etiology will be used as the reference standard. RESULTS: A pilot study was performed from June to August 2017. The enrollment of study subjects started in November 2017. Results are expected by the end of 2019. CONCLUSIONS: The findings from the TREND study can be an important step to improve the management of children with clinical CAP. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/12705 |
BACKGROUND: In Southeast Asian countries, including Myanmar, poultry farming is a major industry. In order to manage and maintain stable productivity, it is important to establish policies for biosecurity. Infectious respiratory diseases are a major threat to poultry farming. Avian influenza and Newcastle disease have been reported in Myanmar, but no scientific information is available for other respiratory pathogens, such as mycoplasmas and infectious bronchitis virus (IBV). Identifying the genotypes and serotypes of IBVs is especially important to inform vaccination programs. In this study, we detected Mycoplasma gallisepticum (MG), M. synoviae (MS), and IBV in several poultry farms in Myanmar. RESULTS: Samples were collected from 20 farms in three major poultry farming areas in Myanmar, and MG, MS, and IBV were detected on two, four, and eight farms, respectively, by polymerase chain reaction. Phylogenetic analysis revealed that the observed MG and MS isolates were not identical to vaccine strains. Three different genotypes of IBV were detected, but none was an unknown variant. CONCLUSIONS: Mycoplasmas and IBV were detected on poultry farms in Myanmar. Periodic surveillance is required to establish the distribution of each pathogen, and to institute better vaccine protocols. |
BACKGROUND: Acute respiratory distress syndrome (ARDS) is a common acute and severe disease in clinic. Recent studies indicated that Cxc chemokine ligand 5 (CXCL5), an inflammatory chemokine, was associated with tumorigenesis. The present study investigated the role of the CXCL5/Cxc chemokine receptor 2 (CXCR2) bio-axis in ARDS, and explored the underlying molecular mechanism. MATERIAL/METHODS: The pathological morphology of lung tissue and degree of pulmonary edema were assessed by hematoxylin-eosin staining and pulmonary edema score, respectively. Real-time PCR and Western blot analysis were performed to detect the expression levels of CXCL5, CXCR2, Matrix metalloproteinases 2 (MMP2), and Matrix metalloproteinases 9 (MMP9) in lung tissues. Enzyme-linked immunosorbent assay (ELISA) was performed to determine the expression levels of CXCL5 and inflammatory factors (IL-1β, IL-6, TNF-α, and IL-10) in serum. RESULTS: The results demonstrated that diffuse alveolar damage and pulmonary edema appeared in lipopolysaccharide (LPS)-induced ARDS and were positively correlated with the severity of ARDS. In addition, CXCL5 and its receptor CXCR2 were overexpressed by upregulation of MMP2 and MMP9 in lung tissues of ARDS. In addition, CXCL5 neutralizing antibody effectively alleviated inflammatory response, diffuse alveolar damage, and pulmonary edema, and decreased the expression levels of MMP2 and MMP9 compared to LPS-induced ARDS. CONCLUSIONS: We found that CXCL5/CXCR2 accelerated the progression of ARDS, partly by upregulation of MMP2 and MMP9 in lung tissues with the release of inflammatory factors. |
Human leukocyte antigen (HLA) alleles have a high degree of polymorphism, which determines their peptide-binding motifs and subsequent T-cell receptor recognition. The simplest way to understand the cross-presentation of peptides by different alleles is to classify these alleles into supertypes. A1 and A3 HLA supertypes are widely distributed in humans. However, direct structural and functional evidence for peptide presentation features of key alleles (e.g., HLA-A(*)30:01 and -A(*)30:03) are lacking. Herein, the molecular basis of peptide presentation of HLA-A(*)30:01 and -A(*)30:03 was demonstrated by crystal structure determination and thermostability measurements of complexes with T-cell epitopes from influenza virus (NP44), human immunodeficiency virus (RT313), and Mycobacterium tuberculosis (MTB). When binding to the HIV peptide, RT313, the PΩ-Lys anchoring modes of HLA-A(*)30:01, and -A(*)30:03 were similar to those of HLA-A(*)11:01 in the A3 supertype. However, HLA-A(*)30:03, but not -A(*)30:01, also showed binding with the HLA(*)01:01-favored peptide, NP44, but with a specific structural conformation. Thus, different from our previous understanding, HLA-A(*)30:01 and -A(*)30:03 have specific peptide-binding characteristics that may lead to their distinct supertype-featured binding peptide motifs. Moreover, we also found that residue 77 in the F pocket was one of the key residues for the divergent peptide presentation characteristics of HLA-A(*)30:01 and -A(*)30:03. Interchanging residue 77 between HLA-A(*)30:01 and HLA-A(*)30:03 switched their presented peptide profiles. Our results provide important recommendations for screening virus and tumor-specific peptides among the population with prevalent HLA supertypes for vaccine development and immune interventions. |
BACKGROUD: Organizing pneumonia (OP) is a rare complication of influenza infection that has substantial morbidity. We report the first case of OP associated with avian influenza H7N9 infection that had significant improvement with corticosteroid treatment. CASE PRESENTATION: A 35-year-old male admitted to intensive care unit because of respiratory failure. He was diagnosed as severe pneumonia caused by avian influenza H7N9 viral infection. After initial clinical improvement supported by extracorporeal membrane oxygenation (ECMO), the patient’s condition worsened with persistent fever, refractory hypoxemia. Chest x-rays and computed tomographies showed areas of consolidation and ground glass opacification. Although OP was suspected and 1 mg/kg methylprednisolone was used, the patient’s condition didn’t improved considerably. An open lung biopsy was performed, and histopathological examination of the specimen was compatible with OP. The patient was treated with methylprednisolone 1.5 mg/kg for 5 days. ECMO was weaned on day 15, and he was discharged on day 71 with good lung recovery. CONCLUSIONS: To the best of our knowledge, this was the first case of successful management of refractory severe respiratory failure caused by avian influenza H7N9 infection complicated with OP. Refractory hypoxia with clinical manifestation and radiological findings compatible with OP, a differential diagnosis should be considered among patients at the second or third week of influenza H7N9 infection, especially in patients with clinical condition deteriorated after the primary influenza pneumonia was controlled. And a steroid dose of methylprednisolone 1.5 mg/kg may be suggested for treatment of OP associated with avian influenza H7N9 infection. |
BACKGROUND: A severe seasonal influenza epidemic was observed during 2017–2018 in China, prompting questions on clinical characteristics and outcomes of severe cases with influenza. METHODS: We retrospectively collected clinical data and outcomes of laboratory-confirmed hospitalized patients (severe to critical) during Jan-2011 to Feb-2018 from five hospitals, followed by a systematic analysis of cases from 2017 to 2018 (n = 289) and all previous epidemics during 2011–2017 (n = 169). RESULTS: In-hospital fatality was over 5-folds higher during the 2017–2018 (p < 0.01) in which 19 patients died (6.6%), whereas only 2 mortalities (1.2%) were observed during 2011–2017. Of the 289 hospitalized in 2017–2018, 153 were confirmed with influenza B virus, 110 with A/H1N1pdm09, and 26 A/H3N2, whereas A/H1N1pdm09 was the predominant cause of hospitalization in previous seasons combined (45%). Fatal cases in 2017–2018 were exclusively associated with either influenza B or A/H1N1pdm09. Our results show that a significant lower proportion of patients aged 14 or greater were treated with oseltamivir, during the 2017–2018 epidemic, and exhibited higher levels of clinical severity. CONCLUSIONS: In-hospital fatality rate might be significantly higher in the 2017–2018 season in China. A sufficient supply of oseltamivir and antiviral therapy within 48 h from onset could reduce fatality rates. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12879-019-4181-2) contains supplementary material, which is available to authorized users. |
Antibodies against foreign antigens are a critical component of the overall immune response and can facilitate pathogen clearance during a primary infection and also protect against subsequent infections. Dysregulation of the antibody response can lead to an autoimmune disease, malignancy, or enhanced infection. Since the experimental delineation of a distinct B cell lineage in 1965, various methods have been developed to understand antigen-specific B cell responses in the context of autoimmune diseases, primary immunodeficiencies, infection, and vaccination. In this review, we summarize the established techniques and discuss new and emerging technologies for probing the B cell response in vitro and in vivo by taking advantage of the specificity of B cell receptor (BCR)-associated and secreted antibodies. These include ELISPOT, flow cytometry, mass cytometry, and fluorescence microscopy to identify and/or isolate primary antigen-specific B cells. We also present our approach to identify rare antigen-specific B cells using magnetic enrichment followed by flow cytometry. Once these cells are isolated, in vitro proliferation assays and adoptive transfer experiments in mice can be used to further characterize antigen-specific B cell activation, function, and fate. Transgenic mouse models of B cells targeting model antigens and of B cell signaling have also significantly advanced our understanding of antigen-specific B cell responses in vivo. |
Replicating recombinant vector vaccines consist of a fully competent viral vector backbone engineered to express an antigen from a foreign transgene. From the perspective of viral replication, the transgene is not only dispensable but may even be detrimental. Thus vaccine revertants that delete or inactivate the transgene may evolve to dominate the vaccine virus population both during the process of manufacture of the vaccine as well as during the course of host infection. A particular concern is that this vaccine evolution could reduce its antigenicity—the immunity elicited to the transgene. We use mathematical and computational models to study vaccine evolution and immunity. These models include evolution arising during the process of manufacture, the dynamics of vaccine and revertant growth, plus innate and adaptive immunity elicited during the course of infection. Although the selective basis of vaccine evolution is easy to comprehend, the immunological consequences are not. One complication is that the opportunity for vaccine evolution is limited by the short period of within-host growth before the viral population is cleared. Even less obvious, revertant growth may only weakly interfere with vaccine growth in the host and thus have a limited effect on immunity to vaccine. Overall, we find that within-host vaccine evolution can sometimes compromise vaccine immunity, but only when the extent of evolution during vaccine manufacture is severe, and this evolution can be easily avoided or mitigated. |
Ranavirus cross-species infections have been documented, but the viral proteins involved in the interaction with cell receptors have not yet been identified. Here, viral cell-binding proteins and their cognate cellular receptors were investigated using two ranaviruses, Andrias davidianus ranavirus (ADRV) and Rana grylio virus (RGV), and two different cell lines, Chinese giant salamander thymus cells (GSTC) and Epithelioma papulosum cyprinid (EPC) cells. The heparan sulfate (HS) analog heparin inhibited plaque formation of ADRV and RGV in the two cell lines by more than 80% at a concentration of 5 μg/mL. In addition, enzymatic removal of cell surface HS by heparinase I markedly reduced plaque formation by both viruses and competition with heparin reduced virus-cell binding. These results indicate that cell surface HS is involved in ADRV and RGV cell binding and infection. Furthermore, recombinant viral envelope proteins ADRV-58L and RGV-53R bound heparin-Sepharose beads implying the potential that cell surface HS is involved in the initial interaction between ranaviruses and susceptible host cells. To our knowledge, this is the first report identifying cell surface HS as ranavirus binding factor and furthers understanding of interactions between ranaviruses and host cells. |
Heparan sulfate proteoglycans (HSPG) are composed of unbranched, negatively charged heparan sulfate (HS) polysaccharides attached to a variety of cell surface or extracellular matrix proteins. Widely expressed, they mediate many biological activities, including angiogenesis, blood coagulation, developmental processes, and cell homeostasis. HSPG are highly sulfated and broadly used by a range of pathogens, especially viruses, to attach to the cell surface. In this review, we summarize the current knowledge on HSPG–virus interactions and distinguish viruses with established HS binding, viruses that bind HS only after intra-host or cell culture adaptation, and finally, viruses whose dependence on HS for infection is debated. We also provide an overview of the antiviral compounds designed to interfere with HS binding. Many questions remain about the true importance of these receptors in vivo, knowledge that is critical for the design of future antiviral therapies. |
Infectious laryngotracheitis (ILT) is an acute respiratory disease of poultry caused by infectious laryngotracheitis virus (ILTV). Control of the disease with live attenuated vaccines administered via eye drop build upon immune responses generated by the eye-associated lymphoid tissues. The aim of this study was to assess cytokine and lymphocyte changes in the conjunctiva-associated lymphoid tissues (CALT) and Harderian gland (HG) stimulated by the ocular inoculation of the ILTV chicken embryo origin (CEO) vaccine strain and virulent strain 63140. This study offers strong evidence to support the roles that the CALT and HG play in the development of protective ILTV immune responses. It supports the premise that ILTV-mediated immunomodulation favors the B cell response over those of T cells. Further, it provides evidence that expansions of CD8α(+) cells, with the concomitant expression of the Granzyme A gene, are key to reducing viral genomes in the CALT and halting ILTV cytolytic replication in the conjunctiva. Ultimately, this study revealed that the early upregulation of interleukin (IL)-12p40 and Interferon (IFN)-γ cytokine genes, which shape the antigen-specific cell-mediated immune responses, retarded the decline of virus replication, and enhanced the development of lesions in the conjunctiva epithelium. |
The herbs Plantago asiatica and Clerodendrum trichotomum have been commonly used for centuries in indigenous and folk medicine in tropical and subtropical regions of the world. In this study, we show that extracts from these herbs have antiviral effects against the respiratory syncytial virus (RSV) in vitro cell cultures and an in vivo mouse model. Treatment of HEp2 cells and A549 cells with a non-cytotoxic concentration of Plantago asiatica or Clerodendrum trichotomum extract significantly reduced RSV replication, RSV-induced cell death, RSV gene transcription, RSV protein synthesis, and also blocked syncytia formation. Interestingly, oral inoculation with each herb extract significantly improved viral clearance in the lungs of BALB/c mice. Based on reported information and a high-performance liquid chromatography (HPLC) analysis, the phenolic glycoside acteoside was identified as an active chemical component of both herb extracts. An effective dose of acteoside exhibited similar antiviral effects as each herb extract against RSV in vitro and in vivo. Collectively, these results suggest that extracts of Plantago asiatica and Clerodendrum trichotomum could provide a potent natural source of an antiviral drug candidate against RSV infection. |
The −2/−1 programmed ribosomal frameshifting (−2/−1 PRF) mechanism in porcine reproductive and respiratory syndrome virus (PRRSV) leads to the translation of two additional viral proteins, nonstructural protein 2TF (nsp2TF) and nsp2N. This −2/−1 PRF mechanism is transactivated by a viral protein, nsp1β, and cellular poly(rC) binding proteins (PCBPs). Critical elements for −2/−1 PRF, including a slippery sequence and a downstream C-rich motif, were also identified in 11 simarteriviruses. However, the slippery sequences (XXXUCUCU instead of XXXUUUUU) in seven simarteriviruses can only facilitate −2 PRF to generate nsp2TF. The nsp1β of simian hemorrhagic fever virus (SHFV) was identified as a key factor that transactivates both −2 and −1 PRF, and the universally conserved Tyr111 and Arg114 in nsp1β are essential for this activity. In vitro translation experiments demonstrated the involvement of PCBPs in simarterivirus −2/−1 PRF. Using SHFV reverse genetics, we confirmed critical roles of nsp1β, slippery sequence, and C-rich motif in −2/−1 PRF in SHFV-infected cells. Attenuated virus growth ability was observed in SHFV mutants with impaired expression of nsp2TF and nsp2N. Comparative genomic sequence analysis showed that key elements of −2/−1 PRF are highly conserved in all known arteriviruses except equine arteritis virus (EAV) and wobbly possum disease virus (WPDV). Furthermore, −2/−1 PRF with SHFV PRF signal RNA can be stimulated by heterotypic nsp1βs of all non-EAV arteriviruses tested. Taken together, these data suggest that −2/−1 PRF is an evolutionarily conserved mechanism employed in non-EAV/-WPDV arteriviruses for the expression of additional viral proteins that are important for viral replication. IMPORTANCE Simarteriviruses are a group of arteriviruses infecting nonhuman primates, and a number of new species have been established in recent years. Although these arteriviruses are widely distributed among African nonhuman primates of different species, and some of them cause lethal hemorrhagic fever disease, this group of viruses has been undercharacterized. Since wild nonhuman primates are historically important sources or reservoirs of human pathogens, there is concern that simarteriviruses may be preemergent zoonotic pathogens. Thus, molecular characterization of simarteriviruses is becoming a priority in arterivirology. In this study, we demonstrated that an evolutionarily conserved ribosomal frameshifting mechanism is used by simarteriviruses and other distantly related arteriviruses for the expression of additional viral proteins. This mechanism is unprecedented in eukaryotic systems. Given the crucial role of ribosome function in all living systems, the potential impact of the in-depth characterization of this novel mechanism reaches beyond the field of virology. |
Infectious diseases remain as the major causes of human and animal morbidity and mortality leading to significant healthcare expenditure in India. The country has experienced the outbreaks and epidemics of many infectious diseases. However, enormous successes have been obtained against the control of major epidemic diseases, such as malaria, plague, leprosy and cholera, in the past. The country's vast terrains of extreme geo-climatic differences and uneven population distribution present unique patterns of distribution of viral diseases. Dynamic interplays of biological, socio-cultural and ecological factors, together with novel aspects of human-animal interphase, pose additional challenges with respect to the emergence of infectious diseases. The important challenges faced in the control and prevention of emerging and re-emerging infectious diseases range from understanding the impact of factors that are necessary for the emergence, to development of strengthened surveillance systems that can mitigate human suffering and death. In this article, the major emerging and re-emerging viral infections of public health importance have been reviewed that have already been included in the Integrated Disease Surveillance Programme. |
Influenza is a highly contagious virus that causes seasonal epidemics and unpredictable pandemics. Four influenza virus types have been identified to date: A, B, C and D, with only A–C known to infect humans. Influenza A and B viruses are responsible for seasonal influenza epidemics in humans and are responsible for up to a billion flu infections annually. The M2 protein is present in all influenza types and belongs to the class of viroporins, i.e., small proteins that form ion channels that increase membrane permeability in virus-infected cells. In influenza A and B, AM2 and BM2 are predominantly proton channels, although they also show some permeability to monovalent cations. By contrast, M2 proteins in influenza C and D, CM2 and DM2, appear to be especially selective for chloride ions, with possibly some permeability to protons. These differences point to different biological roles for M2 in types A and B versus C and D, which is also reflected in their sequences. AM2 is by far the best characterized viroporin, where mechanistic details and rationale of its acid activation, proton selectivity, unidirectionality, and relative low conductance are beginning to be understood. The present review summarizes the biochemical and structural aspects of influenza viroporins and discusses the most relevant aspects of function, inhibition, and interaction with the host. |
BACKGROUND: More than 90% of all infections in the head and neck region can be traced back to an odontogenic origin. In rare cases they can lead to sepsis, which may pose a vital threat to the patient. The purpose of this study was to analyse characteristics concerning etiology and progress of severe odontogenic infections with a fulminant development. METHODS: All patients with odontogenic infections requiring hospital admission were included in a retrospective analysis conducted from 02/2012 to 09/2017. Of 483 patients 16 patients (13 male, 3 female) showed severe exacerbation with septic progress. The average age was 52.8 years. All patients underwent at least one surgical procedure that involved an extraoral incision and drainage as well as high volume irrigation intraoperatively. At least one revision was required for four of the patients. Three patients showed an exceedingly severe disease progression with multiorgan dysfunction syndrome (MODS) and circulatory arrest. Antibiotic treatment was adjusted according to the results of an antibiogram and resistogram. Irrigation with saline was done several times a day. RESULTS: Sixteen patients showed odontogenic infections that spread over multiple maxillo-facial and cervical regions accompanied by septic laboratory signs. All these patients needed intensive care and a tracheostomy. The hospitalization period was 27.8 days on average. In 16 cases risk factors for the development of odontogenic abscesses like diabetes mellitus, obesity, chronic alcohol and nicotine abuse, rheumatism and poor oral hygiene were present. Intraoperative swabs showed a typical polymicrobial aerobic and anaerobic spectrum of oral bacteria, especially anaerobes and streptococci, mainly Streptocococcus viridans. CONCLUSION: Odontogenic infections with fulminant progression should be treated based on clinical and imaging data with immediate surgical incision and drainage including elimination of odontogenic foci as well as intensified intra- and postoperative irrigation. If needed, repeat imaging followed by further incisions should be performed. Immediate antibiotic treatment adapted to the antibiogram is of utmost importance. A combination of tazobactam and piperacillin has proven to be a good first choice and can be recommended for abscesses that spread over multiple levels with initial signs of severe infections. |
Astroviruses (AstV) are single-stranded, positive-sense RNA viruses, best known for causing diarrhea in humans and are also found in many other mammals; in those, the relevance in gastroenteritis remains unclear. Recently described neurotropic AstV showed associations with encephalitis in humans as well as in other mammals. In Switzerland, two different neurotropic AstV were identified in cattle, as well as one in a sheep. The high genetic similarity between the ovine and one of the bovine AstV strengthens the hypothesis of an interspecies transmission. In humans, AstV associated with encephalitis were found also in human stool samples, suggesting that in these patients the infection spreads from the gastrointestinal tract to the brain under certain conditions, such as immunosuppression. Whether a similar pathogenesis occurs in ruminants remains unknown. The aims of this study were (1) the investigation of the potential occurrence of neurotropic AstV in feces samples, (2) the discovery and analysis of so far unknown AstV in small ruminants and other ruminant species’ fecal samples and (3) the examination of a potential interspecies transmission of AstV. To achieve these aims, RNA extraction out of 164 fecal samples from different ruminant species was performed and all samples were screened for known neurotropic AstV occurring in Switzerland, as well as for various AstV using RT-PCR. Positive tested samples were submitted to next generation sequencing. The generated sequences were compared to nucleotide- and amino acid databases, virus properties were identified, and phylogenetic analyses as well as recombination analysis were performed. The excretion of neurotropic AstV in small ruminants’ feces could not be demonstrated, but this work suggests the first identification of AstV in goats as well as the discovery of multiple and highly diverse new genetic variants in small ruminants, which lead to a classification into novel genotype-species. Additionally, the prediction of multiple recombination events in four of five newly discovered full or almost full-length genome sequences suggests a plausible interspecies transmission. The findings point out the occurrence and fecal shedding of previously unknown AstV in sheep and goats and pave the way towards a better understanding of the diversity and transmission of AstV in small ruminants. |
Poultry in commercial settings are exposed to a range of stressors. A growing body of information clearly indicates that excess ROS/RNS production and oxidative stress are major detrimental consequences of the most common commercial stressors in poultry production. During evolution, antioxidant defence systems were developed in poultry to survive in an oxygenated atmosphere. They include a complex network of internally synthesised (e.g., antioxidant enzymes, (glutathione) GSH, (coenzyme Q) CoQ) and externally supplied (vitamin E, carotenoids, etc.) antioxidants. In fact, all antioxidants in the body work cooperatively as a team to maintain optimal redox balance in the cell/body. This balance is a key element in providing the necessary conditions for cell signalling, a vital process for regulation of the expression of various genes, stress adaptation and homeostasis maintenance in the body. Since ROS/RNS are considered to be important signalling molecules, their concentration is strictly regulated by the antioxidant defence network in conjunction with various transcription factors and vitagenes. In fact, activation of vitagenes via such transcription factors as Nrf2 leads to an additional synthesis of an array of protective molecules which can deal with increased ROS/RNS production. Therefore, it is a challenging task to develop a system of optimal antioxidant supplementation to help growing/productive birds maintain effective antioxidant defences and redox balance in the body. On the one hand, antioxidants, such as vitamin E, or minerals (e.g., Se, Mn, Cu and Zn) are a compulsory part of the commercial pre-mixes for poultry, and, in most cases, are adequate to meet the physiological requirements in these elements. On the other hand, due to the aforementioned commercially relevant stressors, there is a need for additional support for the antioxidant system in poultry. This new direction in improving antioxidant defences for poultry in stress conditions is related to an opportunity to activate a range of vitagenes (via Nrf2-related mechanisms: superoxide dismutase, SOD; heme oxygenase-1, HO-1; GSH and thioredoxin, or other mechanisms: Heat shock protein (HSP)/heat shock factor (HSP), sirtuins, etc.) to maximise internal AO protection and redox balance maintenance. Therefore, the development of vitagene-regulating nutritional supplements is on the agenda of many commercial companies worldwide. |
Proteolytic cleavage regulates numerous processes in health and disease. One key player is the ubiquitously expressed serine protease furin, which cleaves a plethora of proteins at polybasic recognition motifs. Mammalian substrates of furin include cytokines, hormones, growth factors and receptors. Thus, it is not surprising that aberrant furin activity is associated with a variety of disorders including cancer. Furthermore, the enzymatic activity of furin is exploited by numerous viral and bacterial pathogens, thereby enhancing their virulence and spread. In this review, we describe the physiological and pathophysiological substrates of furin and discuss how dysregulation of a simple proteolytic cleavage event may promote infectious diseases and cancer. One major focus is the role of furin in viral glycoprotein maturation and pathogenicity. We also outline cellular mechanisms regulating the expression and activation of furin and summarise current approaches that target this protease for therapeutic intervention. |
BACKGROUND: Most current state-of-the-art models for searching the International Classification of Diseases, Tenth Revision Clinical Modification (ICD-10-CM) codes use word embedding technology to capture useful semantic properties. However, they are limited by the quality of initial word embeddings. Word embedding trained by electronic health records (EHRs) is considered the best, but the vocabulary diversity is limited by previous medical records. Thus, we require a word embedding model that maintains the vocabulary diversity of open internet databases and the medical terminology understanding of EHRs. Moreover, we need to consider the particularity of the disease classification, wherein discharge notes present only positive disease descriptions. OBJECTIVE: We aimed to propose a projection word2vec model and a hybrid sampling method. In addition, we aimed to conduct a series of experiments to validate the effectiveness of these methods. METHODS: We compared the projection word2vec model and traditional word2vec model using two corpora sources: English Wikipedia and PubMed journal abstracts. We used seven published datasets to measure the medical semantic understanding of the word2vec models and used these embeddings to identify the three–character-level ICD-10-CM diagnostic codes in a set of discharge notes. On the basis of embedding technology improvement, we also tried to apply the hybrid sampling method to improve accuracy. The 94,483 labeled discharge notes from the Tri-Service General Hospital of Taipei, Taiwan, from June 1, 2015, to June 30, 2017, were used. To evaluate the model performance, 24,762 discharge notes from July 1, 2017, to December 31, 2017, from the same hospital were used. Moreover, 74,324 additional discharge notes collected from seven other hospitals were tested. The F-measure, which is the major global measure of effectiveness, was adopted. RESULTS: In medical semantic understanding, the original EHR embeddings and PubMed embeddings exhibited superior performance to the original Wikipedia embeddings. After projection training technology was applied, the projection Wikipedia embeddings exhibited an obvious improvement but did not reach the level of original EHR embeddings or PubMed embeddings. In the subsequent ICD-10-CM coding experiment, the model that used both projection PubMed and Wikipedia embeddings had the highest testing mean F-measure (0.7362 and 0.6693 in Tri-Service General Hospital and the seven other hospitals, respectively). Moreover, the hybrid sampling method was found to improve the model performance (F-measure=0.7371/0.6698). CONCLUSIONS: The word embeddings trained using EHR and PubMed could understand medical semantics better, and the proposed projection word2vec model improved the ability of medical semantics extraction in Wikipedia embeddings. Although the improvement from the projection word2vec model in the real ICD-10-CM coding task was not substantial, the models could effectively handle emerging diseases. The proposed hybrid sampling method enables the model to behave like a human expert. |
Fulminant viral hepatitis (FVH) is a devastating and unexplained condition that strikes otherwise healthy individuals during primary infection with common liver-tropic viruses. We report a child who died of FVH upon infection with hepatitis A virus (HAV) at age 11 yr and who was homozygous for a private 40-nucleotide deletion in IL18BP, which encodes the IL-18 binding protein (IL-18BP). This mutation is loss-of-function, unlike the variants found in a homozygous state in public databases. We show that human IL-18 and IL-18BP are both secreted mostly by hepatocytes and macrophages in the liver. Moreover, in the absence of IL-18BP, excessive NK cell activation by IL-18 results in uncontrolled killing of human hepatocytes in vitro. Inherited human IL-18BP deficiency thus underlies fulminant HAV hepatitis by unleashing IL-18. These findings provide proof-of-principle that FVH can be caused by single-gene inborn errors that selectively disrupt liver-specific immunity. They also show that human IL-18 is toxic to the liver and that IL-18BP is its antidote. |
BACKGROUND: Researchers increasingly use social contact data to inform models for infectious disease spread with the aim of guiding effective policies about disease prevention and control. In this article, we undertake a systematic review of the study design, statistical analyses, and outcomes of the many social contact surveys that have been published. METHODS: We systematically searched PubMed and Web of Science for articles regarding social contact surveys. We followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines as closely as possible. RESULTS: In total, we identified 64 social contact surveys, with more than 80% of the surveys conducted in high-income countries. Study settings included general population (58%), schools or universities (37%), and health care/conference/research institutes (5%). The largest number of studies did not focus on a specific age group (38%), whereas others focused on adults (32%) or children (19%). Retrospective (45%) and prospective (41%) designs were used most often with 6% using both for comparison purposes. The definition of a contact varied among surveys, e.g., a nonphysical contact may require conversation, close proximity, or both. We identified age, time schedule (e.g., weekday/weekend), and household size as relevant determinants of contact patterns across a large number of studies. CONCLUSIONS: We found that the overall features of the contact patterns were remarkably robust across several countries, and irrespective of the study details. By considering the most common approach in each aspect of design (e.g., sampling schemes, data collection, definition of contact), we could identify recommendations for future contact data surveys that may be used to facilitate comparison between studies. |
Viral infections cause substantial human morbidity and mortality, and are a significant health burden worldwide. Following a viral infection, the host may initiate complex antiviral immune responses to antagonize viral invasion and replication. However, proinflammatory antiviral immune responses pose a great threat to the host if not properly held in check. Interleukin (IL)-17 is a pleiotropic cytokine participating in a variety of physiological and pathophysiological conditions, including tissue integrity maintenance, cancer progression, autoimmune disease development and, more intriguingly, infectious diseases. Abundant evidence suggests that while IL-17 plays a crucial role in enhancing effective antiviral immune responses, it may also promote and exacerbate virus-induced illnesses. Accumulated experimental and clinical evidence has broadened our understanding of the seemingly paradoxical role of IL-17 in viral infections and suggests that IL-17-targeted immunotherapy may be a promising therapeutic option. Herein, we summarize current knowledge regarding the protective and pathogenic roles of IL-17 in viral infections, with emphasis on underlying mechanisms. The various and critical roles of IL-17 in viral infections necessitate the development of therapeutic strategies that are uniquely tailored to both the infectious agent and the infection environment. |
We hypothesized that fever in children with viral bronchiolitis indicates the need for consideration of superimposed bacterial pneumonia. We conducted a retrospective study of 349 children aged 2 years and younger with diagnoses of respiratory syncytial virus (RSV) and viral upper respiratory infection. Data were analyzed using Pearson χ(2) test. One hundred seventy-eight children had RSV with no other identified virus. The majority of children (56%) who had only RSV were afebrile. Febrile children with RSV were over twice as likely to be diagnosed with bacterial pneumonia as those who were afebrile (60% vs 27%, P < .001). In the 171 children who had bronchiolitis caused by a virus other than RSV, 51% were afebrile. These children were 8 times more likely to be diagnosed with pneumonia than those who were afebrile (65% vs 8%, P < .001). Evaluation of febrile children with viral bronchiolitis may allow early diagnosis and treatment of secondary bacterial pneumonia. |
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