RESUMEN
Avian influenza outbreaks, including ones caused by highly pathogenic A(H5N1) clade 2.3.4.4b viruses, have devastated animal populations and remain a threat to humans. Risk elements assessed for emerging influenza viruses include their susceptibility to approved antivirals. Here, we screened >20,000 neuraminidase (NA) or polymerase acidic (PA) protein sequences of potentially pandemic A(H5Nx), A(H7Nx), and A(H9N2) viruses that circulated globally in 2010-2023. The frequencies of NA or PA substitutions associated with reduced inhibition (RI) or highly reduced inhibition (HRI) by NA inhibitors (NAIs) (oseltamivir, zanamivir) or a cap-dependent endonuclease inhibitor (baloxavir) were low: 0.60% (137/22,713) and 0.62% (126/20,347), respectively. All tested subtypes were susceptible to NAIs and baloxavir at sub-nanomolar concentrations. A(H9N2) viruses were the most susceptible to oseltamivir, with IC50s 3- to 4-fold lower than for other subtypes (median IC50: 0.18 nM; n = 22). NA-I222M conferred RI of A(H5N1) viruses by oseltamivir (with a 26-fold IC50 increase), but NA-S246N did not reduce inhibition. PA-E23G, PA-K34R, PA-I38M/T, and the previously unreported PA-A36T caused RI by baloxavir in all subtypes tested. Avian A(H9N2) viruses endemic in Egyptian poultry predominantly acquired PA-I38V, which causes only a <3-fold decrease in the baloxavir EC50 and fails to meet the RI criteria. PA-E199A/D in A(H7Nx) and A(H9N2) viruses caused a 2- to 4-fold decrease in EC50 (close to the borderline for RI) and should be closely monitored. Our data indicate antiviral susceptibility is high among avian influenza A viruses with pandemic potential and present novel markers of resistance to existing antiviral interventions.
Asunto(s)
Antivirales , Aves , Dibenzotiepinas , Farmacorresistencia Viral , Inhibidores Enzimáticos , Genotipo , Virus de la Influenza A , Gripe Aviar , Neuraminidasa , Oseltamivir , Piridonas , Triazinas , Neuraminidasa/antagonistas & inhibidores , Neuraminidasa/genética , Antivirales/farmacología , Gripe Aviar/virología , Animales , Inhibidores Enzimáticos/farmacología , Dibenzotiepinas/farmacología , Farmacorresistencia Viral/genética , Piridonas/farmacología , Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza A/genética , Virus de la Influenza A/enzimología , Triazinas/farmacología , Oseltamivir/farmacología , Aves/virología , Morfolinas/farmacología , Endonucleasas/antagonistas & inhibidores , Endonucleasas/genética , Endonucleasas/metabolismo , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/genética , Proteínas Virales/genética , Proteínas Virales/antagonistas & inhibidores , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N1 del Virus de la Influenza A/genética , Subtipo H5N1 del Virus de la Influenza A/enzimología , Zanamivir/farmacología , Fenotipo , Humanos , Concentración 50 InhibidoraRESUMEN
Avian influenza, particularly the H9N2 subtype, presents significant challenges to poultry health, underscoring the need for effective antiviral interventions. This study explores the antiviral capabilities of Belamcanda extract, a traditional Chinese medicinal herb, against H9N2 Avian influenza virus (AIV) in specific pathogen-free (SPF) chicks. Through a comprehensive approach, we evaluated the impact of the extract on cytokine modulation and crucial immunological signaling pathways, essential for understanding the host-virus interaction. Our findings demonstrate that Belamcanda extract significantly modulates the expression of key inflammatory cytokines, including tumor necrosis factor alpha (TNF-α), interleukin-1 (IL-1), interleukin-2 (IL-2), and interleukin-6 (IL-6), which are pivotal to the host's response to H9N2 AIV infection. Western blot analysis further revealed that the extract markedly reduces the expression of critical immune signaling molecules such as toll-like receptor 3 (TLR3), TIR-domain-containing adapter-inducing interferon-ß (TRIF), and nuclear factor kappa B (NF-κB). These insights into the mechanisms by which Belamcanda extract influences host immune responses and hinders viral replication highlight its potential as an innovative antiviral agent for poultry health management. The study advances our comprehension of natural compounds' antiviral mechanisms and lays the groundwork for developing strategies to manage viral infections in poultry. The demonstrated ability of Belamcanda extract to modulate immune responses and inhibit viral replication establishes it as a promising candidate for future antiviral therapy development, especially in light of the need for effective treatments against evolving influenza virus strains and the critical demand for enhanced poultry health management strategies.
Asunto(s)
Antivirales , Pollos , Subtipo H9N2 del Virus de la Influenza A , Gripe Aviar , Enfermedades de las Aves de Corral , Animales , Subtipo H9N2 del Virus de la Influenza A/fisiología , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Gripe Aviar/virología , Gripe Aviar/tratamiento farmacológico , Antivirales/farmacología , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/tratamiento farmacológico , Enfermedades de las Aves de Corral/inmunología , Organismos Libres de Patógenos Específicos , Inflamación/tratamiento farmacológico , Inflamación/veterinaria , Inflamación/virología , Citocinas/metabolismo , Citocinas/genética , Extractos Vegetales/farmacologíaRESUMEN
Avian influenza virus subtype H9N2 has the ability to infect birds and humans, further causing significant losses to the poultry industry and even posing a great threat to human health. Oral vaccine received particular interest for preventing majority infection due to its ability to elicit both mucosal and systemic immune responses, but their development is limited by the bad gastrointestinal (GI) environment, compact epithelium and mucus barrier, and the lack of effective mucosal adjuvants. Herein, we developed the dendritic fibrous nano-silica (DFNS) grafted with Cistanche deserticola polysaccharide (CDP) nanoparticles (CDP-DFNS) as an adjuvant for H9N2 vaccine. Encouragingly, CDP-DFNS facilitated the proliferation of T and B cells, and further induced the activation of T lymphocytes in vitro. Moreover, CDP-DFNS/H9N2 significantly promoted the antigen-specific antibodies levels in serum and intestinal mucosal of chickens, indicating the good ability to elicit both systemic and mucosal immunity. Additional, CDP-DFNS facilitate the activation of CD4 + and CD8 + T cells both in spleen and intestinal mucosal, and the indexes of immune organs. This study suggested that CDP-DFNS may be a new avenue for development of oral vaccine against pathogens that are transmitted via mucosal route.
Asunto(s)
Adyuvantes Inmunológicos , Pollos , Inmunidad Mucosa , Subtipo H9N2 del Virus de la Influenza A , Vacunas contra la Influenza , Gripe Aviar , Nanopartículas , Polisacáridos , Dióxido de Silicio , Animales , Subtipo H9N2 del Virus de la Influenza A/inmunología , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Polisacáridos/administración & dosificación , Polisacáridos/farmacología , Polisacáridos/química , Polisacáridos/inmunología , Dióxido de Silicio/administración & dosificación , Dióxido de Silicio/química , Nanopartículas/administración & dosificación , Vacunas contra la Influenza/administración & dosificación , Vacunas contra la Influenza/inmunología , Inmunidad Mucosa/efectos de los fármacos , Gripe Aviar/prevención & control , Gripe Aviar/inmunología , Adyuvantes Inmunológicos/administración & dosificación , Adyuvantes Inmunológicos/farmacología , Administración Oral , Mucosa Intestinal/inmunología , Mucosa Intestinal/efectos de los fármacos , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunologíaRESUMEN
In our research, we explored a natural substance called Oxymatrine, found in a traditional Chinese medicinal plant, to fight against a common bird flu virus known as H9N2. This virus not only affects birds but can also pose a threat to human health. We focused on how this natural compound can help in stopping the virus from spreading in cells that line the lungs of birds and potentially humans. Our findings show that Oxymatrine can both directly block the virus and boost the body's immune response against it. This dual-action mechanism is particularly interesting because it indicates that Oxymatrine might be a useful tool in developing new ways to prevent and treat this type of bird flu. Understanding how Oxymatrine works against the H9N2 virus could lead to safer and more natural ways to combat viral infections in animals and humans, contributing to the health and well-being of society. The H9N2 Avian Influenza Virus (AIV) is a persistent health threat because of its rapid mutation rate and the limited efficacy of vaccines, underscoring the urgent need for innovative therapies. This study investigated the H9N2 AIV antiviral properties of Oxymatrine (OMT), a compound derived from traditional Chinese medicine, particularly focusing on its interaction with pulmonary microvascular endothelial cells (PMVECs). Employing an array of in vitro assays, including 50% tissue culture infectious dose, Cell Counting Kit-8, reverse transcription-quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blot, we systematically elucidated the multifaceted effects of OMT. OMT dose-dependently inhibited critical antiviral proteins (PKR and Mx1) and modulated the expression of type I interferons and key cytokines (IFN-α, IFN-ß, IL-6, and TNF-α), thereby affecting TLR3 signaling and its downstream elements (NF-κB and IRF-3). OMT's antiviral efficacy extended beyond TLR3-mediated responses, suggesting its potential as a versatile antiviral agent. This study not only contributes to the growing body of research on the use of natural compounds as antiviral agents but also underscores the importance of further investigating the broader application of OMT for combating viral infections.
Asunto(s)
Antivirales , Subtipo H9N2 del Virus de la Influenza A , Gripe Aviar , Matrinas , Transducción de Señal , Receptor Toll-Like 3 , Animales , Perros , Humanos , Antivirales/farmacología , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Gripe Aviar/tratamiento farmacológico , Gripe Aviar/inmunología , Células de Riñón Canino Madin Darby , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 3/metabolismoRESUMEN
The circulation of avian influenza A viruses in poultry is a public health concern due to the potential transmissibility and severity of these viral infections. Monitoring the susceptibility of these viruses to antivirals is important for developing measures to strengthen the level of preparedness against influenza pandemics. However, drug susceptibility information on these viruses is limited. Here, we determined the susceptibilities of avian influenza A(H5N1), A(H5N2), A(H5N8), A(H7N7), A(H7N9), A(H9N1), and A(H9N2) viruses isolated in Japan to the antivirals approved for use there: an M2 inhibitor (amantadine), neuraminidase inhibitors (oseltamivir, peramivir, zanamivir, and laninamivir) and RNA polymerase inhibitors (baloxavir and favipiravir). Genotypic methods that detect amino acid substitutions associated with antiviral resistance and phenotypic methods that assess phenotypic viral susceptibility to drugs have revealed that these avian influenza A viruses are susceptible to neuraminidase and RNA polymerase inhibitors. These results suggest that neuraminidase and RNA polymerase inhibitors currently approved in Japan could be a treatment option against influenza A virus infections in humans.
Asunto(s)
Farmacorresistencia Viral , Gripe Aviar , Gripe Humana , Animales , Antivirales/farmacología , Antivirales/uso terapéutico , ARN Polimerasas Dirigidas por ADN , Farmacorresistencia Viral/genética , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Humanos , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H7N7 del Virus de la Influenza A/efectos de los fármacos , Subtipo H7N9 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Gripe Aviar/epidemiología , Gripe Aviar/virología , Gripe Humana/epidemiología , Gripe Humana/virología , Japón/epidemiología , Neuraminidasa/genética , Neuraminidasa/metabolismo , Oseltamivir/farmacología , Oseltamivir/uso terapéutico , Aves de CorralRESUMEN
BACKGROUND: H9N2 avian influenza viruses (AIVs) infect avian and mammalian hosts and provide internal genes for new emerging highly pathogenic avian viruses that cause severe pneumonia with high mortality, for which few medications are available. Arctiin, a bioactive lignan glycoside, has been reported to possess multiple pharmacological properties. However, the effect of arctiin on H9N2 virus infection is unclear. In the current study, we analyzed the effect of arctiin on H9N2 virus infection and the underlying molecular mechanism in vitro. METHODS: The antiviral effect against H9N2 virus was determined by plaque reduction assay (PRA) and progeny virus reduction assay. We employed MTT assay, qRT-PCR, ELISA, immunofluorescence and Western blotting to better understand the anti-inflammatory effect and corresponding mechanism of arctiin on H9N2 virus-infected cells. RESULTS: The results showed that arctiin had antiviral activity against H9N2 virus. Arctiin treatment reduced H9N2 virus-triggered proinflammatory cytokines, such as IL-6, and TNF-α. Moreover, arctiin significantly suppressed H9N2 virus-mediated expression of COX-2 and PGE2. Furthermore, we found that arctiin inhibited H9N2 virus-mediated activation of RIG-I/JNK MAPK signaling. Interestingly, arctiin treatment obviously reversed H9N2 virus-induced reduction of Nrf2, increased the nuclear translocation of Nrf2, and upregulated Nrf2 signaling target genes (HO-1 and SOD2). Zinc protoporphyrin (Znpp)-an HO-1 inhibitor-weakened the inhibitory effect of arctiin on H9N2 virus-induced RIG-I/JNK MAPK and proinflammatory mediators. CONCLUSION: Taken together, these results suggested that the anti-inflammatory effects of arctiin on H9N2 virus infection may be due to the activation of Nrf2/HO-1 and blocked RIG-I/JNK MAPK signaling; thus, arctiin may be a promising agent for prevention and treatment of H9N2 virus infections.
Asunto(s)
Antiinflamatorios/farmacología , Furanos/farmacología , Glucósidos/farmacología , Hemo-Oxigenasa 1/metabolismo , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Extractos Vegetales/farmacología , Células A549 , Animales , Antiinflamatorios/química , China , Perros , Furanos/química , Glucósidos/química , Humanos , Células de Riñón Canino Madin Darby , Estructura MolecularRESUMEN
The recent advances in our understanding of the host factors in orchestrating qualitatively different immune responses against influenza Type A virus (IAV) have changed the perception of conventional approaches for controlling avian influenza virus (AIV) infection in chickens. Given that infection-induced pathogenicity and replication of influenza virus largely rely on regulating host immune responses, immunoregulatory cytokine profiles often determine the disease outcomes. However, in contrast to the function of other inflammatory cytokines, interleukin-17A (IL-17A) has been described as a 'double-edged sword', indicating that in addition to antiviral host responses, IL-17A has a distinct role in promoting viral infection. Therefore, in the present study, we investigated the chicken IL-17A mediated antiviral immune effects on IAVs infection in primary chicken embryo fibroblasts cells (CEFs). To this end, we first bioengineered a food-grade Lactic Acid Producing Bacteria (LAB), Lactococcus lactis (L. lactis), secreting bioactive recombinant chicken IL-17A (sChIL-17A). Next, the functionality of sChIL-17A was confirmed by transcriptional upregulation of several genes associated with antiviral host responses, including granulocyte-monocyte colony-stimulating factor (GM-CSF) (CSF3 in the chickens), interleukin-6 (IL-6), interferon-α (IFN-α), -ß and -γ genes in primary CEFs cells. Consistent with our hypothesis that such a pro-inflammatory state may translate to immunoprotection against IAVs infection, we observed that sChIL-17A pre-treatment could significantly limit the viral replication and protect the primary CEFs cells against two heterotypic IAVs such as A/turkey/Wisconsin/1/1966(H9N2) and A/PR/8/1934(H1N1). Together, the data presented in this work suggest that exogenous application of sChIL-17A secreted by modified LAB vector may represent an alternative strategy for improving antiviral immunity against avian influenza virus infection in chickens.
Asunto(s)
Bioingeniería , Citoprotección , Fibroblastos/virología , Vectores Genéticos/metabolismo , Subtipo H1N1 del Virus de la Influenza A/fisiología , Subtipo H9N2 del Virus de la Influenza A/fisiología , Interleucina-17/farmacología , Lactobacillales/genética , Animales , Muerte Celular/efectos de los fármacos , Células Cultivadas , Embrión de Pollo , Pollos/virología , Efecto Citopatogénico Viral/efectos de los fármacos , Citoprotección/efectos de los fármacos , Perros , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Perfilación de la Expresión Génica , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/genética , Gripe Aviar/inmunología , Gripe Aviar/virología , Interleucina-17/genética , Células de Riñón Canino Madin Darby , Nisina/farmacología , Fenotipo , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/farmacología , Transcripción Genética/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Proteínas Virales/metabolismo , Replicación Viral/efectos de los fármacosRESUMEN
Curcumin is a potent antimicrobial herb used traditionally as a spice in culinary. This study was designed to evaluate the antiviral effects of curcuma longa extract against H9 influenza virus. A total of 60 embryonated eggs were equally divided into 5 groups with 12 eggs in each group. Group 1 (G1) served as uninfected negative control. Whereas Group 2 (G2) was kept as positive control infected with known virus @ 0.2 ml with 10-9.3 EID50. Group 3 (G3) was offered Curcuma longa @ 0.2 mg/0.2 ml and H9N2 virus (@ 0.2 ml with 10-9.3 EID50. Similarly, Group 4 (G4) was inoculated with extract of Curcuma longa @ 0.2 mg/0.2 ml per egg, whereas Group 5 (G5) was given Ribazole @ 0.2 ml/ egg. The crude extract and virus were administered on the 15th day of incubation and were checked after every 24 hours up to 96th hour post inoculation by random selection of 3 eggs. Death and survival rate were noted in all groups. Gross and histopathological lesions were also observed. Results revealed that Curcuma longa extract had significantly (p<0.05) reduced the mortality rate of embryos infected with H9N2 virus. In G3, increased lymphocytes and mild fatty changes were seen in liver. Whereas, mature RBCs, plasma cells and some lymphoblast's were observed in Spleen. Similarly, the bursa follicles were with lymphocytic aggregation. The G4 (Curcuma longa) and G5 (Ribazole) were with maximum embryo survival after 48 and 72 h post inoculation. This study revealed potential antiviral activity of Curcuma longa against H9N2 influenza viruses and can be opted as alternative to antibiotics and antiviral drugs to minimize the antimicrobial resistance in human and animal population.
Asunto(s)
Antivirales , Curcuma , Subtipo H9N2 del Virus de la Influenza A , Extractos Vegetales/farmacología , Animales , Antivirales/farmacología , Embrión de Pollo , Curcuma/química , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacosRESUMEN
BACKGROUND: Diallyl trisulfide (DATS) is a garlic-derived organosulfur compound. As it has been shown to have anti-viral activity, we hypothesized that it may alleviate infections caused by H9N2 avian influenza virus (AIV), which is prevalent in poultry with pandemic potential. METHODS: Human lung A549 epithelial cells were treated with three different concentrations of DATS 24 h before (pre-treatment) or one hour after (post-treatment) H9N2 AIV infection. Culture supernatants were collected 24 h and 48 h post-infection and analyzed for viral titers and levels of inflammatory and anti-viral immune responses. For in vivo experiments, BABL/c mice were administered daily by intraperitoneal injection with DATS (30 mg/kg) for 2 weeks starting 1 day after H9N2 AIV infection. Clinical signs, lung pathology, and inflammatory and anti-viral immune responses were assessed 2, 4, and 6 days after infection. RESULTS: Both pre-treatment and post-treatment of A549 cells with DATS resulted in reduced viral loads, increased expression of anti-viral genes (RIG-I, IRF-3, and interferon-ß), and decreased expression of inflammatory cytokines (TNF-α and IL-6). These effects were also observed in H9N2 AIV-infected mice treated with DATS. Such treatment also reduced lung edema and inflammation in mice. CONCLUSIONS: Results suggest that DATS has anti-viral activity against H9N2 AIV and may be used as an alternative treatment for influenza virus infection.
Asunto(s)
Compuestos Alílicos/farmacología , Antivirales/farmacología , Subtipo H9N2 del Virus de la Influenza A , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Sulfuros/farmacología , Células A549 , Animales , Pollos , Humanos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Gripe Aviar/tratamiento farmacológico , RatonesRESUMEN
Influenza viruses cause significant morbidity and mortality worldwide. Long-term or frequent use of approved anti-influenza agents has resulted in drug-resistant strains, thereby necessitating the discovery of new drugs. In this study, we found aprotinin, a serine protease inhibitor, as an anti-influenza candidate through screening of compound libraries. Aprotinin has been previously reported to show inhibitory effects on a few influenza A virus (IAV) subtypes (e.g., seasonal H1N1 and H3N2). However, because there were no reports of its inhibitory effects on the other types of influenza viruses, we investigated the inhibitory effects of aprotinin in vitro on a wide range of influenza viruses, including avian and oseltamivir-resistant influenza virus strains. Our cell-based assay showed that aprotinin had inhibitory effects on seasonal human IAVs (H1N1 and H3N2 subtypes), avian IAVs (H5N2, H6N5, and H9N2 subtypes), an oseltamivir-resistant IAV, and a currently circulating influenza B virus. We have also confirmed its activity in mice infected with a lethal dose of influenza virus, showing a significant increase in survival rate. Our findings suggest that aprotinin has the capacity to inhibit a wide range of influenza virus subtypes and should be considered for development as a therapeutic agent against influenza.
Asunto(s)
Antivirales/farmacología , Aprotinina/farmacología , Evaluación Preclínica de Medicamentos , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Inhibidores de Serina Proteinasa/farmacología , Animales , Línea Celular , Perros , Humanos , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H5N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N2 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/crecimiento & desarrollo , Virus de la Influenza B/efectos de los fármacos , Virus de la Influenza B/crecimiento & desarrollo , Células de Riñón Canino Madin Darby , Ratones , Ratones Endogámicos C57BLRESUMEN
Janus kinase (JAK) inhibitors have been developed as novel immunomodulatory drugs and primarily used for treating rheumatoid arthritis and other inflammatory diseases. Recent studies have suggested that this category of anti-inflammatory drugs could be potentially useful for the control of inflammation "storms" in respiratory virus infections. In addition to their role in regulating immune cell functions, JAK1 and JAK2 have been recently identified as crucial cellular factors involved in influenza A virus (IAV) replication and could be potentially targeted for antiviral therapy. Gingerenone A (Gin A) is a compound derived from ginger roots and a dual inhibitor of JAK2 and p70 S6 kinase (S6K1). Our present study aimed to determine the antiviral activity of Gin A on influenza A virus (IAV) and to understand its mechanisms of action. Here, we reported that Gin A suppressed the replication of three IAV subtypes (H1N1, H5N1, H9N2) in four cell lines. IAV replication was also inhibited by Ruxolitinib (Rux), a JAK inhibitor, but not by PF-4708671, an S6K1 inhibitor. JAK2 overexpression enhanced H5N1 virus replication and attenuated Gin A-mediated antiviral activity. In vivo experiments revealed that Gin A treatment suppressed IAV replication in the lungs of H5N1 virus-infected mice, alleviated their body weight loss, and prolonged their survival. Our study suggests that Gin A restricts IAV replication by inhibiting JAK2 activity; Gin A could be potentially useful for the control of influenza virus infections.
Asunto(s)
Antivirales/farmacología , Diarilheptanoides/farmacología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Janus Quinasa 2/antagonistas & inhibidores , Células A549 , Animales , Línea Celular , Perros , Femenino , Células HEK293 , Humanos , Imidazoles/farmacología , Subtipo H1N1 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H5N1 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H9N2 del Virus de la Influenza A/crecimiento & desarrollo , Células de Riñón Canino Madin Darby , Ratones , Ratones Endogámicos C57BL , Nitrilos , Piperazinas/farmacología , Pirazoles/farmacología , Pirimidinas , Proteínas Quinasas S6 Ribosómicas 70-kDa/antagonistas & inhibidores , Replicación Viral/efectos de los fármacosRESUMEN
The aim of this study was to test in vitro the ability of a mixture of citrus extract, maltodextrin, sodium chloride, lactic acid and citric acid (AuraShield L) to inhibit the virulence of infectious bronchitis, Newcastle disease, avian influenza, porcine reproductive and respiratory syndrome (PRRS) and bovine coronavirus viruses. Secondly, in vivo, we have investigated its efficacy against infectious bronchitis using a broiler infection model. In vitro, these antimicrobials had expressed antiviral activity against all five viruses through all phases of the infection process of the host cells. In vivo, the antimicrobial mixture reduced the virus load in the tracheal and lung tissue and significantly reduced the clinical signs of infection and the mortality rate in the experimental group E2 receiving AuraShield L. All these effects were accompanied by a significant reduction in the levels of pro-inflammatory cytokines and an increase in IgA levels and short chain fatty acids (SCFAs) in both trachea and lungs. Our study demonstrated that mixtures of natural antimicrobials, such AuraShield L, can prevent in vitro viral infection of cell cultures. Secondly, in vivo, the efficiency of vaccination was improved by preventing secondary viral infections through a mechanism involving significant increases in SCFA production and increased IgA levels. As a consequence the clinical signs of secondary infections were significantly reduced resulting in recovered production performance and lower mortality rates in the experimental group E2.
Asunto(s)
Antivirales/farmacología , Infecciones por Coronavirus/tratamiento farmacológico , Coronavirus Bovino/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Virus de la Bronquitis Infecciosa/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Virus de la Enfermedad de Newcastle/efectos de los fármacos , Virus del Síndrome Respiratorio y Reproductivo Porcino/efectos de los fármacos , Enfermedades de las Aves de Corral/tratamiento farmacológico , Animales , Línea Celular , Embrión de Pollo , Pollos , Infecciones por Coronavirus/virología , Modelos Animales de Enfermedad , Células Epiteliales/virología , Humanos , Gripe Aviar/metabolismo , Gripe Aviar/virología , Gripe Humana/metabolismo , Gripe Humana/virología , Enfermedad de Newcastle/metabolismo , Enfermedad de Newcastle/virología , Síndrome Respiratorio y de la Reproducción Porcina/metabolismo , Síndrome Respiratorio y de la Reproducción Porcina/virología , Enfermedades de las Aves de Corral/virología , PorcinosRESUMEN
Influenza virus neuraminidase (NA) contains a universally conserved epitope (NAe, NA222-230). However, no studies have reported vaccines targeting this NA conserved epitope and inducing antibodies recognizing NAe. The extracellular domain of M2 (M2e) is considered as an attractive target for a universal influenza vaccine. We generated recombinant influenza H1N1 viruses expressing conserved epitopes in hemagglutinin (HA) molecules: NAe (NAe-HA) or M2e (M2e-HA) within the HA head domain. Inactivated recombinant NAe-HA and M2e-HA viruses were more effective in inducing IgG antibodies specific for an inserted conserved epitope than live recombinant virus. Recombinant inactivated M2e-HA virus vaccination induced cross protection against H3N2 virus with less weight loss compared to NAe-HA and was more effective in inducing humoral and cellular M2e immune responses. This study provides insight into developing recombinant influenza virus vaccines compatible with current platforms to induce antibody responses to conserved poorly immunogenic epitopes.
Asunto(s)
Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Subtipo H1N1 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/biosíntesis , Neuraminidasa/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Virus Reordenados/inmunología , Proteínas de la Matriz Viral/inmunología , Animales , Anticuerpos Antivirales/biosíntesis , Protección Cruzada , Epítopos/química , Epítopos/inmunología , Femenino , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Humanos , Inmunidad Celular/efectos de los fármacos , Inmunidad Humoral/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/inmunología , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N1 del Virus de la Influenza A/genética , Subtipo H5N1 del Virus de la Influenza A/inmunología , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/genética , Subtipo H9N2 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/administración & dosificación , Vacunas contra la Influenza/genética , Ratones , Ratones Endogámicos BALB C , Neuraminidasa/genética , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/virología , Virus Reordenados/efectos de los fármacos , Virus Reordenados/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Vacunación/métodos , Vacunas de Productos Inactivados , Proteínas de la Matriz Viral/genéticaRESUMEN
The H9N2 subtype avian influenza virus (AIV) is one of the most prevalent AIV subtypes that can be found throughout most countries. Currently, due to the neglect of low pathogenic avian influenza virus (LPAIV) and monotonous control technique, an expanding H9N2 virus epizootic have been arisen and causes great economic losses in the poultry industry. Therefore, novel anti-influenza drugs are necessary for the prevention and control of H9N2 AIV. Our previous studies have found that Taishan Pinus massoniana pollen polysaccharides (TPPPS) have antiviral effects, but whether they can inhibit the H9N2 AIV remains unclear. Here, we further investigated the effects of TPPPS on the H9N2 virus and its underlying mechanisms of action. We found that TPPPS significantly inhibited the replication of the H9N2 virus in a dose-dependent manner, especially during the period of virus adsorption in vitro. Transmission electron microscopy demonstrated that TPPPS reduce infection by interfering with virus entry into host cells rather than by interacting with the H9N2 virus particles. A fluorescence quantitative PCR (qPCR) assay and an animal experiment were performed to evaluate the anti-viral effect of TPPPS in vivo. As expected, the lungs of chickens treated with TPPPS had fewer lesions and lower virus contents compared with the PBS group. In addition, pre-treatment with TPPPS clearly enhanced host disease resistance and delayed infection by the H9N2 virus. Taken together, our results reveal that TPPPS suppress H9N2 virus replication both in vitro and in vivo and therefore shows promising as an anti-AIV agent.
Asunto(s)
Antivirales/uso terapéutico , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Pinus/química , Polen/química , Polisacáridos/uso terapéutico , Administración Oral , Animales , Anticuerpos Antivirales/sangre , Pollos/virología , Perros , Gripe Aviar/tratamiento farmacológico , Gripe Aviar/prevención & control , Células de Riñón Canino Madin Darby , Enfermedades de las Aves de Corral/tratamiento farmacológico , Enfermedades de las Aves de Corral/prevención & control , Enfermedades de las Aves de Corral/virología , Carga Viral , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Angelica dahurica (Hoffm.) Benth. & Hook.f. ex Franch. & Sav. (Umbelliferae family) is an herbaceous, perennial plant native to northern and eastern Asia. The root of A. dahurica has traditionally been used under the name "Bai Zhi" as a medicinal plant for colds, dizziness, ulcers, and rheumatism. Moreover, it is also an important ingredient of various prescriptions, such as Gumiganghwal-Tang, for the common cold and influenza. AIM OF THE STUDY: Even though various biological activities of the root of A. dahurica have been reported along with its chemical components, the detailed mechanism of how it exerts anti-influenza activity at the compound level has not been studied. Therefore, we investigated the anti-influenza properties of furanocoumarins purified by bioactivity-guided isolation. MATERIALS AND METHODS: Bioactivity-guided isolation from a 70% EtOH extract of the root of A. dahurica was performed to produce four active furanocoumarins. The inhibition of cytopathic effects (CPEs) was evaluated to ascertain the antiviral activity of these compounds against influenza A (H1N1 and H9N2) viruses. The most potent compound was subjected to detailed mechanistic studies such as the inhibition of viral protein synthesis, CPE inhibition in different phases of the viral replication cycle, neuraminidase (NA) inhibition, antiapoptotic activity using flow cytometry, and immunofluorescence. RESULTS: The bioactivity-guided isolation produced four active furanocoumarins, isoimperatorin (1), oxypeucedanin (2), oxypeucedanin hydrate (3) and imperatorin (4) from the n-BuOH fraction. Among them, compound 2 (followed by compounds 1, 4 and 3) showed a significant CPE inhibition effect, which was stronger than that of the positive control ribavirin, against both H1N1 and H9N2 with an EC50 (µM) of 5.98 ± 0.71 and 4.52 ± 0.39, respectively. Compound 2 inhibited the synthesis of NA and nucleoprotein (NP) in a dose-dependent manner. In the time course assays, the cytopathic effects of influenza A-infected MDCK cells were reduced by 80-90% when treated with compound 2 for 1 and 2 h after infection and declined drastically 3 h after infection. The level of viral NA and NP production was markedly reduced to less than 20% for both proteins in compound 2 (20 µM)-treated cells compared to untreated cells at 2 h after infection. In the molecular docking analysis, compound 2 showed a stronger binding affinity for the C-terminus of polymerase acidic protein (PAC; -36.28 kcal/mol) than the other two polymerase subunits. Compound 2 also exerted an antiapoptotic effect on virus infected cells and significantly inhibited the mRNA expression of caspase-3 and Bax. CONCLUSION: Our results suggest that compound 2 might exert anti-influenza A activity via the inhibition of the early phase of the viral replication cycle, not direct neutralization of surface proteins, such as hemagglutinin and NA, and abnormal apoptosis induced by virus infection. Taken together, these findings suggest that furanocoumarins predominant in A. dahurica play a pivotal role in its antiviral activity. These findings can also explain the reasons for the ethnopharmacological uses of this plant as an important ingredient in many antiviral prescriptions in traditional Chinese medicine (TCM).
Asunto(s)
Angelica , Antivirales/farmacología , Células Epiteliales/efectos de los fármacos , Furocumarinas/farmacología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Extractos Vegetales/farmacología , Angelica/química , Animales , Antivirales/aislamiento & purificación , Apoptosis/efectos de los fármacos , Efecto Citopatogénico Viral/efectos de los fármacos , Perros , Células Epiteliales/metabolismo , Células Epiteliales/patología , Células Epiteliales/virología , Furocumarinas/aislamiento & purificación , Interacciones Microbiota-Huesped , Subtipo H1N1 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H1N1 del Virus de la Influenza A/metabolismo , Subtipo H9N2 del Virus de la Influenza A/crecimiento & desarrollo , Subtipo H9N2 del Virus de la Influenza A/metabolismo , Células de Riñón Canino Madin Darby , Simulación del Acoplamiento Molecular , Neuraminidasa/antagonistas & inhibidores , Neuraminidasa/metabolismo , Infecciones por Orthomyxoviridae/metabolismo , Infecciones por Orthomyxoviridae/patología , Infecciones por Orthomyxoviridae/virología , Extractos Vegetales/aislamiento & purificación , Raíces de Plantas , Replicación Viral/efectos de los fármacosRESUMEN
Low-pathogenicity avian influenza H9N2 remains an endemic disease worldwide despite continuous vaccination, indicating the need for an improved vaccine strategy. Bacillus subtilis (B. subtilis), a gram-positive and endospore-forming bacterium, is a non-pathogenic species that has been used in probiotic formulations for both animals and humans. The objective of the present study was to elucidate the effect of B. subtilis spores as adjuvants in chickens administered inactivated avian influenza virus H9N2. Herein, the adjuvanticity of B. subtilis spores in chickens was demonstrated by enhancement of H9N2 virus-specific IgG responses. B. subtilis spores enhanced the proportion of B cells and the innate cell population in splenocytes from chickens administered both inactivated H9N2 and B. subtilis spores (Spore + H9N2). Furthermore, the H9N2 and spore administration induced significantly increased expression of the pro-inflammatory cytokines IL-1ß and IL-6 compared to that in the H9N2 only group. Additionally, total splenocytes from chickens immunized with inactivated H9N2 in the presence or absence of B. subtilis spores were re-stimulated with inactivated H9N2. The subsequent results showed that the extent of antigen-specific CD4+ and CD8+ T cell proliferation was higher in the Spore + H9N2 group than in the group administered only H9N2. Taken together, these data demonstrate that B. subtilis spores, as adjuvants, enhance not only H9N2 virus-specific IgG but also CD4+ and CD8+ T cell responses, with an increase in pro-inflammatory cytokine production. This approach to vaccination with inactivated H9N2 together with a B. subtilis spore adjuvant in chickens produces a significant effect on antigen-specific antibody and T cell responses against avian influenza virus.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Linfocitos B/inmunología , Bacillus subtilis/química , Pollos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Gripe Aviar/inmunología , Linfocitos T/inmunología , Adyuvantes Inmunológicos/química , Animales , Anticuerpos Antivirales/inmunología , Antivirales/química , Antivirales/farmacología , Subtipo H9N2 del Virus de la Influenza A/inmunología , Enfermedades de las Aves de Corral/inmunología , Esporas Bacterianas/químicaRESUMEN
Biodegradable nanomaterials can protect antigens from degradation, promote cellular absorption, and enhance immune responses. We constructed a eukaryotic plasmid [pCAGGS-opti441-hemagglutinin (HA)] by inserting the optimized HA gene fragment of H9N2 AIV into the pCAGGS vector. The pCAGGS-opti441-HA/DGL was developed through packaging the pCAGGS-opti441-HA with dendrigraft poly-l-lysines (DGLs). DGL not only protected the pCAGGS-opti441-HA from degradation, but also exhibited high transfection efficiency. Strong cellular immune responses were induced in chickens immunized with the pCAGGS-opti441-HA/DGL. The levels of IFN-γ and IL-2, and lymphocyte transformation rate of the vaccinated chickens increased at the third week post the immunization. For the vaccinated chickens, T lymphocytes were activated and proliferated, the numbers of CD3+CD4+ and CD4+/CD8+ increased, and the chickens were protected completely against H9N2 AIV challenge. This study provides a method for the development of novel AIV vaccines, and a theoretical basis for the development of safe and efficient gene delivery carriers.
Asunto(s)
Anticuerpos Antivirales/inmunología , Vacunas contra la Influenza/farmacología , Gripe Aviar/tratamiento farmacológico , Vacunas de ADN/farmacología , Animales , Anticuerpos Antivirales/farmacología , Formación de Anticuerpos/efectos de los fármacos , Formación de Anticuerpos/inmunología , Pollos/inmunología , Pollos/virología , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/inmunología , Subtipo H9N2 del Virus de la Influenza A/patogenicidad , Vacunas contra la Influenza/química , Vacunas contra la Influenza/genética , Vacunas contra la Influenza/inmunología , Gripe Aviar/inmunología , Gripe Aviar/virología , Polilisina/química , Polilisina/farmacología , Vacunas de ADN/química , Vacunas de ADN/inmunologíaRESUMEN
BACKGROUND: Influenza viruses (IVs) have become increasingly resistant to antiviral drugs that target neuraminidase and matrix protein 2 due to gene mutations that alter their drug-binding target protein regions. Consequently, almost all recent IV pandemics have exhibited resistance to commercial antiviral vaccines. To overcome this challenge, an antiviral target is needed that is effective regardless of genetic mutations. MAIN BODY: In particular, hemagglutinin (HA), a highly conserved surface protein across many IV strains, could be an effective antiviral target as it mediates binding of IVs with host cell receptors, which is crucial for membrane fusion. HA has 6 disulfide bonds that can easily bind with the surfaces of gold nanoparticles. Herein, we fabricated porous gold nanoparticles (PoGNPs) via a surfactant-free emulsion method that exhibited strong affinity for disulfide bonds due to gold-thiol interactions, and provided extensive surface area for these interactions. A remarkable decrease in viral infectivity was demonstrated by increased cell viability results after exposing MDCK cells to various IV strains (H1N1, H3N2, and H9N2) treated with PoGNP. Most of all, the viability of MDCK cells infected with all IV strains increased to 96.8% after PoGNP treatment of the viruses compared to 33.9% cell viability with non-treated viruses. Intracellular viral RNA quantification by real-time RT-PCR also confirmed that PoGNP successfully inhibited viral membrane fusion by blocking the viral entry process through conformational deformation of HA. CONCLUSION: We believe that the technique described herein can be further developed for PoGNP-utilized antiviral protection as well as metal nanoparticle-based therapy to treat viral infection. Additionally, facile detection of IAV can be achieved by developing PoGNP as a multiplatform for detection of the virus.
Asunto(s)
Antivirales/farmacología , Oro/farmacología , Virus de la Influenza A/efectos de los fármacos , Nanopartículas del Metal/química , Animales , Perros , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/genética , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/genética , Virus de la Influenza A/genética , Células de Riñón Canino Madin Darby , Fusión de Membrana , Porosidad , ARN Viral/análisis , ARN Viral/genética , Internalización del VirusRESUMEN
Influenza A viruses (IAV) have been a major public health threat worldwide, and options for antiviral therapy become increasingly limited with the emergence of drug-resisting virus strains. New and effective anti-IAV drugs, especially for highly pathogenic influenza, with different modes of action, are urgently needed. The influenza virus glycoprotein hemagglutinin (HA) plays critical roles in the early stage of virus infection, including receptor binding and membrane fusion, making it a potential target for the development of anti-influenza drugs. In this study, we show that OA-10, a newly synthesized triterpene out of 11 oleanane-type derivatives, exhibited significant antiviral activity against four different subtypes of IAV (H1N1, H5N1, H9N2 and H3N2) replications in A549 cell cultures with EC50 ranging from 6.7 to 19.6 µM and a negligible cytotoxicity (CC50 > 640 µM). It inhibited acid-induced hemolysis in a dose-dependent manner, with an IC50 of 26 µM, and had a weak inhibition on the adsorption of H5 HA to chicken erythrocytes at higher concentrations (≥40 µM). Surface plasmon resonance (SPR) analysis showed that OA-10 interacted with HA in a dose-dependent manner with the equilibrium dissociation constants (KD) of the interaction of 2.98 × 10-12 M. Computer-aided molecular docking analysis suggested that OA-10 might bind to the cavity in HA stem region which is known to undergo significant rearrangement during membrane fusion. Our results demonstrate that OA-10 inhibits H5N1 IAV replication mainly by blocking the conformational changes of HA2 subunit required for virus fusion with endosomal membrane. These findings suggest that OA-10 could serve as a lead for further development of novel virus entry inhibitors to prevent and treat IAV infections.
Asunto(s)
Antivirales/farmacología , Glicoproteínas Hemaglutininas del Virus de la Influenza/metabolismo , Virus de la Influenza A/efectos de los fármacos , Ácido Oleanólico/química , Ácido Oleanólico/farmacología , Internalización del Virus/efectos de los fármacos , Células A549 , Animales , Antivirales/química , Pollos , Eritrocitos/efectos de los fármacos , Hemólisis/efectos de los fármacos , Humanos , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H5N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H9N2 del Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza A/fisiología , Concentración 50 Inhibidora , Simulación del Acoplamiento MolecularRESUMEN
Herbal medicines are becoming more popular and acceptable day by day due to their effectiveness, limited side effects, and cost-effectiveness. Cholistani plants are reported as a rich source of antibacterial, antifungal, antiprotozoal, antioxidant, and anticancer agents. The current study has evaluated antiviral potential of selected Cholistani plants. The whole plants were collected, ground and used in extract formation with n-hexane, ethyl acetate and n-butanol. All the extracts were concentrated by using a rotary evaporator and concentrate was finally dissolved in an appropriate vol of the same solvent. All of the extracts were tested for their antiviral potential by using 9-11 days old chick embryonated eggs. Each extract was tested against the Avian Influenza virus H9N2 strain (AIV), New Castle Disease virus Lasoota strain (NDV), Infectious bronchitis virus (IBV) and an Infectious bursal disease virus (IBDV). Hemagglutination test (HA) and Indirect Hemagglutination (IHA) tests were performed for different viruses. The overall order of the antiviral potential of Cholistani plants against viruses was NDV>IBV>IBDV>AIV. In terms of antiviral activity from extracts, the order of activity was n-butanol>ethyl acetate>n-hexane. The medicinal plants Achyranthes aspera, Neuroda procumbens, Panicum antidotale, Ochthochloa compressa and Suaeda fruticose were very effective against all four poultry viruses through their extracts. The low IC50 values of these extracts confirm the high antiviral potential against these viruses. It is worth to mention that Achyranthes aspera was found positive against IBDV through all its extracts which overcome the problem of unavailability of any known drug against IBDV. In short, the study proved that Cholistani plants are rich source of antiviral agent and their extracts can be used as good source of antiviral drugs both in crude and in purified form.