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1.
Zhonghua Yu Fang Yi Xue Za Zhi ; 53(9): 929-933, 2019 Sep 06.
Artigo em Chinês | MEDLINE | ID: mdl-31474076

RESUMO

Objective: Analyze the genetic characteristic of Hemagglutinin(H) gene of measles viruses isolated in Henan Province in 2017. Methods: Swab samples collected from 7 lab confirmed measles cases, and we got the measles virus by Vero/Slam inoculation. Fragment of H genes were amplified by reverse transcription polymerase chain reaction(RT-PCR), then the PCR products were sequenced and analyzed. Results: The age of the 7 measles confirmed cases were between 1 and 50 years old, and all of them were males. All the 7 measles viruses were identified as H1a genotype, and the average distance of the nucleotides and the amino acids was 0.005, respectively. Compared with the Shanghai-191/China-vaccine, there were some changes in isolated virus, such as 240(th), 397(th) and 381(st) sites in the amino acid sequence. Conclusion: The measles genotype which isolated in Henan Province in 2017 was H1a. There were some difference from Shanghai-191/China-vaccine in the nucleotides sequence of H gene, which suggested that it's necessary to strengthen the monitor the variation of measles virus.


Assuntos
Hemaglutininas , Vírus do Sarampo , Sarampo/virologia , Adolescente , Adulto , Criança , Pré-Escolar , China , Genótipo , Hemaglutininas/genética , Humanos , Lactente , Masculino , Vacina contra Sarampo/genética , Vírus do Sarampo/genética , Pessoa de Meia-Idade , Filogenia , Adulto Jovem
2.
Nanoscale ; 11(29): 13878-13884, 2019 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-31304500

RESUMO

Stimulation of dendritic cells (DCs) by antigens (Ags) promotes an Ag-specific immune response that kills Ag-expressing pathogens. These biologically inspired nanocarriers have received much attention as tools to deliver cancer Ags to DCs. A polymer-templated protein nanoball having hemagglutinin (H1-NB) that mimics the influenza virus can be used as a cancer Ag delivery vehicle, as DCs show effective phagocytic activities against H1-NB without any adjuvant. In the present study, H1-NB containing ovalbumin (OVA), a model Ag (H1-OVA-NB), was prepared as an anti-cancer agent and evaluated for its effect on anticancer immunity. H1-OVA-NB treatments in C57BL/6 mice enhanced OVA-specific immune activation and efficiently inhibited B16-OVA tumor growth compared to control groups. Our results indicate that H1-NB is an effective carrier for Ag delivery to DCs and promotes immunotherapy to fight cancer.


Assuntos
Antígenos/imunologia , Células Dendríticas/imunologia , Hemaglutininas/química , Imunoterapia , Nanopartículas/química , Polímeros/química , Animais , Antígenos/química , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Portadores de Fármacos/química , Hemaglutininas/genética , Hemaglutininas/metabolismo , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Orthomyxoviridae/metabolismo , Ovalbumina/genética , Ovalbumina/imunologia , Ovalbumina/metabolismo
3.
Western Pac Surveill Response J ; 10(1): 32-38, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31110840

RESUMO

Introduction: There are two methods of reverse transcription polymerase chain reaction (RT-PCR) that have been the common methods to detect influenza infections: conventional and real-time RT-PCR. From December 2017 to March 2018, several missed diagnoses of influenza A(H1)pdm09 using real-time RT-PCR were reported in northern Viet Nam. This study investigated how these missed detections occurred to determine their effect on the surveillance of influenza. Methods: The haemagglutinin (HA) segments of A(H1N1)pdm09 from both real-time RT-PCR positive and negative samples were isolated and sequenced. The primer and probe sets in the HA gene were checked for mismatches, and phylogenetic analyses were performed to determine the molecular epidemiology of these viruses. Results: There were 86 positive influenza A samples; 32 were A(H1)pdm09 positive by conventional RT-PCR but were negative by real-time RT-PCR. Sequencing was conducted on 23 influenza (H1N1)pdm09 isolates that were recovered from positive samples. Eight of these were negative for A(H1)pdm09 by real-time RT-PCR. There were two different mismatches in the probe target sites of the HA gene sequences of all isolates (n = 23) with additional mismatches only at position 7 (template binding site) identified for all eight negative real-time RT-PCR isolates. The prime target sites had no mismatches. Phylogenetic analysis of the HA gene showed that both the positive and negative real-time RT-PCR isolates were grouped in clade 6B.1; however, the real-time RT-PCR negative viruses were located in a subgroup that referred to substitution I295V. Conclusion: Constant monitoring of genetic changes in the circulating influenza A(H1N1)pdm09 viruses is important for maintaining the sensitivity of molecular detection assays.


Assuntos
Diagnóstico Tardio/tendências , Influenza Humana/diagnóstico , Análise de Sequência de DNA/normas , Testes de Hemaglutinação/métodos , Hemaglutininas/análise , Hemaglutininas/genética , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/patogenicidade , Influenza Humana/epidemiologia , Influenza Humana/mortalidade , Mutação/genética , Filogenia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Análise de Sequência de DNA/métodos , Análise de Sequência de DNA/tendências , Vietnã
4.
J Gen Virol ; 100(6): 958-967, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31140969

RESUMO

Newcastle disease (ND), which is caused by Newcastle disease virus (NDV), is a highly contagious disease in chickens and is a great threat to the poultry industry. Fusion of the viral and target cell membranes is a prerequisite for NDV's entry into host cells. This process is directly mediated by the fusion (F) protein. Although several domains of F are known to regulate membrane fusion activity, the roles of the DI-DII linker (residues 376-381) of the NDV F protein in membrane fusion still remain unclear. To investigate the roles of this linker in NDV F-induced cell-cell fusion, mutations were engineered into this linker by site-directed mutagenesis. These mutants were analysed with respect to cell surface expression and membrane fusion activity. Each of the mutated F proteins in this linker was expressed at the cell surface at a similar level to wild-type (WT) F. However, most of them resulted in significant alterations in fusion activity. In particular, the mutants G377S, A378D, L379A and T380P were able to independently mediate cell fusion in the absence of HN protein in BHK-21 cells. Taken together, the results indicated that the DI-DII linker region has an important effect on the fusion activity of NDV F and mutants in this region could alter the requirement for HN for the promotion of membrane fusion.


Assuntos
Hemaglutininas/genética , Proteínas de Fusão de Membrana/genética , Mutação/genética , Neuraminidase/genética , Vírus da Doença de Newcastle/genética , Animais , Fusão Celular/métodos , Linhagem Celular , Membrana Celular/genética , Cricetinae , Doença de Newcastle/virologia , Células Vero
5.
Arch Virol ; 164(5): 1309-1321, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30877453

RESUMO

Full-length infectious cDNA clones for recombinant HY12 bovine enteroviruses designated as rHY12-3A-2-HA, rHY12-3A-3-HA, and rHY12-3A-9-HA were constructed by the insertion of an epitope from influenza virus hemagglutinin (HA) at the N-terminus of the HY12-encoded 3A protein at amino acid positions 2, 3, and 9. The recombinant HY12 viruses expressing the HA epitope were rescued and characterized using immunoperoxidase monolayer assay, western blotting, and electron microscopy. The three rescued recombinant marker viruses showed similar characteristics, such as TCID50 titer, plaque size, and growth properties, to those of parental rHY12 virus. Comparative analysis of the nucleotide sequences demonstrated the three recombinant marker viruses remained stable for 15 passages with no genetic changes. The recombinant viruses remained viable in various permissive cell lines, including BHK-21, Vero, and PK15 cells, suggesting that the insertion of the HA epitope tag had no effect on virus infectivity. Mice infected with the recombinant marker viruses and the parental virus produced anti-HY12-virus antibodies, while the recombinant marker viruses also produced anti-HA-epitope-tag antibodies. Taken together, these results demonstrate that HY12 viruses containing genetic markers may be useful tools for future investigations of the mechanisms of viral pathogenesis and virus replication, as well as for vaccine development.


Assuntos
Anticorpos Antivirais/imunologia , Enterovirus Bovino/genética , Enterovirus Bovino/imunologia , Epitopos/imunologia , Hemaglutininas/imunologia , Proteínas Virais/imunologia , Animais , Linhagem Celular , Cricetinae , Cães , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Hemaglutininas/genética , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos ICR , Suínos , Células Vero , Proteínas Virais/genética
6.
Mol Biol Evol ; 36(6): 1172-1186, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30851115

RESUMO

Seasonal influenza viruses undergo frequent mutations on their surface hemagglutinin (HA) proteins to escape the host immune response. In these mutations, a few key amino acid sites are associated with significant antigenic cluster transitions. To recognize the cluster-transition determining sites of seasonal influenza A/H3N2 and A/H1N1 viruses systematically and quickly, we developed a computational model named RECDS (recognition of cluster-transition determining sites) to evaluate the contribution of a specific amino acid site on the HA protein in the whole history of antigenic evolution. In RECDS, we ranked all of the HA sites by calculating the contribution scores derived from the forest of gradient boosting classifiers trained by various sequence- and structure-based features. With the RECDS model, we found out that the sites determining influenza antigenicity were mostly around the receptor-binding domain both for the influenza A/H3N2 and A/H1N1 viruses. Specifically, half of the cluster-transition determining sites of the influenza A/H1N1 virus were located in the vestigial esterase domain and basic path area on the HA, which indicated that the differential driving force of the antigenic evolution of the A/H1N1 virus refers to the A/H3N2 virus. Beyond that, the footprints of substitutions responsible for antigenic evolution were inferred according to the phylogenetic trees for the cluster-transition determining sites. The monitoring of genetic variation occurring at these cluster-transition determining sites in circulating influenza viruses on a large scale will potentially reduce current assay workloads in influenza surveillance and the selection of new influenza vaccine strains.


Assuntos
Antígenos Virais/genética , Evolução Molecular , Hemaglutininas/genética , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Algoritmos , Técnicas Genéticas , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H3N2/imunologia , Software
7.
Transbound Emerg Dis ; 66(3): 1252-1267, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30725534

RESUMO

Canine distemper (CD) is one of the highly contagious and invariably fatal viral diseases of dogs and other carnivores. Despite the widespread use of modified live vaccines to control CD, the prevalence of disease has increased at an alarming rate in recent years. Although a number of factors may be ascribed for vaccine failure, antigenic differences among the vaccine and wild-type strains have gained the interest of researchers. Considering the high genetic variability of haemagglutinin gene (H gene) and its role in eliciting the immune response to canine distemper virus (CDV), we have generated nine full-length CDV H gene sequences from infected dogs including three vaccinated cases. Bayesian analysis was performed using 102 full-length H gene nucleotide sequences over a time frame of 76 years (1940-2016) from 18 countries. The time to the most recent common ancestor (tMRCA) of CDV was estimated to be 1696 AD. Phylogenetic reconstruction clustered Indian wild-type viruses into a distinct monophyletic group clearly separated from the previously established CDV lineages. This signifies the presence of a novel genetic variant (proposed as "Lineage India-1/Asia-5") circulating among dog population in India. To investigate the importance of substitutions at amino acid residues 530 and 549 of CDV H protein in determining the host switches from canid to non-canid hosts, we analysed 125 H gene sequences including nine sequences generated in this study. Selection pressure analysis and analysis of amino acid sequences revealed a trend towards adaptation of 549H variants in non-canid hosts although no role of G/E530R/D/N substitution could be identified. This is the first comprehensive study about the nature and ecology of CDV circulating among dog population in India. Outbreaks in vaccinated animals as observed in this study have raised a concern towards the effectiveness of current vaccine strains warranting detailed investigation.


Assuntos
Vírus da Cinomose Canina/genética , Cinomose/virologia , Variação Genética , Hemaglutininas/genética , Sequência de Aminoácidos , Animais , Teorema de Bayes , Carnívoros , Cinomose/epidemiologia , Cães , Índia/epidemiologia , Filogenia
8.
Nano Lett ; 19(3): 1875-1882, 2019 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-30719917

RESUMO

Viruses, such as influenza A, typically bind to the plasma membrane of their host by engaging multiple membrane receptors in parallel, thereby forming so-called multivalent interactions that are created by the collective action of multiple weak ligand-receptor bonds. The overall interaction strength can be modulated by changing the number of engaged receptors. This feature is used by viruses to achieve a sufficiently firm attachment to the host's plasma membrane but also allows progeny viruses to leave the plasma membrane after completing the virus replication cycle. Design of strategies to prevent infection, for example, by disturbing these attachment and detachment processes upon application of antivirals, requires quantification of the underlying multivalent interaction in absence and presence of antivirals. This is still an unresolved problem, as there is currently no approach available that allows for determining the valency (i.e., of the number of receptors bound to a particular virus) on the level of single viruses under equilibrium conditions. Herein, we track the motion of single influenza A/X31 viruses (IAVs; interacting with the ganglioside GD1a incorporated in a supported lipid bilayer) using total internal reflection fluorescence microscopy and show that IAV residence time distributions can be deconvoluted from valency effects by taking the IAV mobility into account. The so-derived off-rate distributions, expressed in dependence of an average, apparent valency, show the expected decrease in off-rate with increasing valency but also show an unexpected peak structure, which can be linked to a competition in the opposing functionalities of the two influenza A virus spike proteins, hemagglutinin (HA), and neuraminidase (NA). By application of the antiviral zanamivir that inhibits the activity of NA, we provide direct evidence, how the HA/NA balance modulates this virus-receptor interaction, allowing us to assess the inhibition concentration of zanamivir based on its effect on the multivalent interaction.


Assuntos
Hemaglutininas/química , Influenza Humana/virologia , Neuraminidase/química , Receptores Virais/química , Membrana Celular/química , Gangliosídeo G(M1)/análogos & derivados , Gangliosídeo G(M1)/química , Hemaglutininas/genética , Humanos , Vírus da Influenza A/química , Vírus da Influenza A/genética , Vírus da Influenza A/patogenicidade , Influenza Humana/genética , Bicamadas Lipídicas/química , Neuraminidase/antagonistas & inibidores , Neuraminidase/genética , Receptores Virais/genética , Zanamivir
9.
Transbound Emerg Dis ; 66(1): 186-194, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30126057

RESUMO

Swine influenza is a worldwide disease, which causes damage to the respiratory system of pigs. The H1N1 and H3N2 subtypes circulate mainly in the swine population of Mexico. There is evidence that new subtypes of influenza virus have evolved genetically and have been rearranged with human viruses and from other species; therefore, the aim of our study was to identify and characterize the genetic changes that have been generated in the different subtypes of the swine influenza virus in Mexican pigs. By sequencing and analyzing phylogenetically the eight segments that form the virus genome, the following subtypes were identified: H1N1, H3N2, H1N2 and H5N2; of which, a H1N1 subtype had a high genetic relationship with the human influenza virus. In addition, a H1N2 subtype related to the porcine H1N2 virus reported in the United States was identified, as well as, two other viruses of avian origin from the H5N2 subtype. Particularly for the H5N2 subtype, this is the first time that its presence has been reported in Mexican pigs. The analysis of these sequences demonstrates that in the swine population of Mexico, circulate viruses that have suffered punctual-specific mutations and rearrangements of their proteins with different subtypes, which have successfully adapted to the Mexican swine population.


Assuntos
Genoma Viral , Vírus da Influenza A/genética , Infecções por Orthomyxoviridae/virologia , Doenças dos Suínos/virologia , Proteínas Virais/genética , Animais , Hemaglutininas/genética , Vírus da Influenza A/classificação , Vírus da Influenza A/enzimologia , Vírus da Influenza A/isolamento & purificação , México , Neuraminidase/genética , Filogenia , Análise de Sequência de RNA/veterinária , Sus scrofa , Suínos
10.
Avian Pathol ; 48(2): 91-97, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30465608

RESUMO

Intensive vaccination strategies against Newcastle disease (ND) have been implemented in many countries for a long time, but ND outbreaks still occur frequently, with most isolates belonging to genotype VII of Newcastle disease virus (NDV). Many researchers have revealed that vaccines closely matched to epidemic viruses provide better protection. Therefore, using a previously established reverse genetics system, we generated a recombinant NDV vaccine strain (rLa Sota-HN) based on the La Sota vaccine strain expressing the haemagglutinin-neuraminidase (HN) protein of genotype VII NDV. The pathogenicity of the recombinant virus was confirmed by the mean death time in 9-day-old specific-pathogen-free embryonated chicken eggs and the intracerebral pathogenicity index in 1-day-old specific-pathogen-free chickens. Subsequently, 1-day-old chickens were immunized with commercial vaccine La Sota and recombinant virus rLa Sota-HN and then challenged with virulent genotype VII NDV strain. The results indicated that recombinant virus rLa Sota-HN provided increased protection of vaccinated chickens from morbidity and mortality, and inhibited the shedding of virulent virus after challenging with genotype VII virus, compared with the conventional vaccine La Sota. Our findings indicated that rLa Sota-HN is a promising vaccine candidate to improve the protection efficiency against ND in chickens, thereby preventing frequent outbreaks of this disease.


Assuntos
Neuraminidase/imunologia , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinação/veterinária , Vacinas Virais/imunologia , Animais , Embrião de Galinha , Galinhas , Feminino , Genótipo , Hemaglutininas/genética , Hemaglutininas/imunologia , Neuraminidase/genética , Doença de Newcastle/virologia , Vírus da Doença de Newcastle/enzimologia , Vírus da Doença de Newcastle/genética , Vírus da Doença de Newcastle/patogenicidade , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Vacinas Sintéticas
11.
Pol J Vet Sci ; 21(3): 623-629, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30468340

RESUMO

Canine distemper virus (CDV) infects wild and domestic Canidae worldwide. The hemag- glutinin (H) gene has the highest genetic variation in the genome of this virus. Thus, the H gene is commonly used for lineage identification and genetic analyses. In order to study the genetic characteristics and pathogenicity of CDV strains prevalent in China, 132 samples were collected from domestic dogs with suspected CDV infection, 58 samples were confirmed to be positive, and the H gene was successfully amplified from 15 samples. The epidemic strain was identified as type Asia-1 and the novel mutations, A51T, V58I, R179K and D262N, were detected in this strain. Isolated strains, BJ16B53, BJ16B14, and BJ17B8, were used for an animal infection experiment in raccoon dogs. BJ16B53 and BJ16B14 were found to cause clinical symptoms, death, and exten- sive lesions in various organs. These results are expected to facilitate the development of effective strategies to monitor and control CDV infection in China.


Assuntos
Vírus da Cinomose Canina/genética , Vírus da Cinomose Canina/patogenicidade , Cinomose/virologia , Hemaglutininas/genética , Sequência de Aminoácidos , Animais , China , Cinomose/epidemiologia , Cães , Regulação Viral da Expressão Gênica , Genótipo , Mutação
12.
Nat Commun ; 9(1): 4879, 2018 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-30451858

RESUMO

Kainate-type glutamate receptors play critical roles in excitatory synaptic transmission and synaptic plasticity in the brain. GluK1 and GluK2 possess fundamentally different capabilities in surface trafficking as well as synaptic targeting in hippocampal CA1 neurons. Here we find that the excitatory postsynaptic currents (EPSCs) are significantly increased by the chimeric GluK1(SPGluK2) receptor, in which the signal peptide of GluK1 is replaced with that of GluK2. Coexpression of GluK1 signal peptide completely suppresses the gain in trafficking ability of GluK1(SPGluK2), indicating that the signal peptide represses receptor trafficking in a trans manner. Furthermore, we demonstrate that the signal peptide directly interacts with the amino-terminal domain (ATD) to inhibit the synaptic and surface expression of GluK1. Thus, we have uncovered a trafficking mechanism for kainate receptors and propose that the cleaved signal peptide behaves as a ligand of GluK1, through binding with the ATD, to repress forward trafficking of the receptor.


Assuntos
Região CA1 Hipocampal/metabolismo , Potenciais Pós-Sinápticos Excitadores/fisiologia , Sinais Direcionadores de Proteínas/genética , Receptores de Ácido Caínico/metabolismo , Transmissão Sináptica/fisiologia , Animais , Animais Recém-Nascidos , Sítios de Ligação , Região CA1 Hipocampal/citologia , Expressão Gênica , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Hemaglutininas/genética , Hemaglutininas/metabolismo , Humanos , Microtomia , Proteínas Mutantes Quiméricas/genética , Proteínas Mutantes Quiméricas/metabolismo , Plasticidade Neuronal , Técnicas de Cultura de Órgãos , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Ratos , Receptores de Ácido Caínico/química , Receptores de Ácido Caínico/genética , Sinapses/metabolismo , Sinapses/ultraestrutura
13.
Zhonghua Liu Xing Bing Xue Za Zhi ; 39(11): 1465-1471, 2018 Nov 10.
Artigo em Chinês | MEDLINE | ID: mdl-30462955

RESUMO

Objective: To understand the molecular characteristics of hemagglutinin (HA) and neuraminidase (NA) as well as the disease risk of influenza virus A H7N9 in Guizhou province. Methods: RNAs were extracted and sequenced from HA and NA genes of H7N9 virus strains obtained from 18 cases of human infection with H7N9 virus and 6 environmental swabs in Guizhou province during 2014-2017. Then the variation and the genetic evolution of the virus were analyzed by using a series of bioinformatics software package. Results: Homology analysis of HA and NA genes revealed that 2 strains detected during 2014-2015 shared 98.8%-99.2% and 99.2% similarities with vaccine strains A/Shanghai/2/2013 and A/Anhui/1/2013 recommended by WHO, respectively. Two strains detected in 2016 and 14 strains detected in 2017 shared 98.2%-99.3% and 97.6%-98.8% similarities with vaccine strain A/Hunan/02650/2016, respectively. Other 6 stains detected in 2017 shared 99.1%-99.4% and 98.9%-99.3% similarities with strain A/Guangdong/17SF003/2016, respectively. Phylogenetic analysis showed that all the strains were directly evolved in the Yangtze River Delta evolution branch, but they were derived from different small branch. PEVPKRKRTAR↓GLF was found in 6 of 24 strains cleavage site sequences of HA protein, indicating the characteristic of highly pathogenic avian influenza virus. Mutations A134V, G186V and Q226L at the receptor binding sites were found in the HA. All the strains had a stalk deletion of 5 amino acid residue "QISNT" in NA protein, and drug resistance mutation R294K occurred in strain A/Guizhou-Danzhai/18980/2017. In addition, potential glycosylation motifs mutations NCS42NCT were found in the NA of 9 of 24 strains. Conclusions: HA and NA genes of avian influenza A (H7N9) virus showed genetic divergence in Guizhou province during 2014-2017. The mutations of key sites might enhance the virulence of the virus, human beings are more susceptible to it. Hence, the risk of infection is increasing.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Hemaglutininas/genética , Subtipo H7N9 do Vírus da Influenza A/genética , Influenza Humana/virologia , Neuraminidase/genética , Animais , Sequência de Bases , Aves , China/epidemiologia , Genoma Viral , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Humanos , Subtipo H7N9 do Vírus da Influenza A/enzimologia , Subtipo H7N9 do Vírus da Influenza A/imunologia , Subtipo H7N9 do Vírus da Influenza A/isolamento & purificação , Influenza Aviária , Influenza Humana/epidemiologia , Filogenia , RNA Viral/genética , Análise de Sequência de DNA
14.
PLoS One ; 13(11): e0206987, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30439983

RESUMO

Influenza B virus-caused illness has recently been considered as an urgent public health problem due to substantial morbidity, mortality and life-threatening medical complications. In this study, we have reported the main characteristics of influenza B virus in Mongolia, including prevalence, lineages, suitability with vaccine strains and drug susceptibility against the virus. 15768 specimens were tested by qPCR for detecting influenza viruses. From positive specimens for influenza B virus, the clinical isolates were isolated using MDCK cells. Sequencing analysis, hemagglutination inhibition assay and Neuraminidase inhibitor (NAI) drug susceptibility testing were performed for the clinical isolates. Influenza B virus was around in 3.46% of the samples in Mongolia, and B/Victoria clade-1A and B/Yamagata clade-3 lineages were predominant. Importantly, it was confirmed that the lineages corresponded to the vaccine strains. Moreover, drug susceptibility tests revealed that some Mongolian clinical isolates showed reduced susceptibility to antiviral agents. Interestingly, G104R was identified as a novel mutation, which might have a significant role in drug resistance of the virus. These results describe the characteristics of influenza B viruses that have caused respiratory illness in the population of Mongolia between 2013 and 2017.


Assuntos
Vírus da Influenza B/isolamento & purificação , Influenza Humana/diagnóstico , Animais , Antivirais/farmacologia , Cães , Farmacorresistência Viral/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Testes de Inibição da Hemaglutinação , Hemaglutininas/classificação , Hemaglutininas/genética , Humanos , Vírus da Influenza B/efeitos dos fármacos , Vírus da Influenza B/genética , Influenza Humana/epidemiologia , Influenza Humana/virologia , Células Madin Darby de Rim Canino , Mongólia/epidemiologia , Mutação , Neuraminidase/genética , Filogenia , Prevalência , RNA Viral/química , RNA Viral/genética , RNA Viral/metabolismo , Análise de Sequência de RNA
15.
Vet Microbiol ; 224: 107-115, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30269784

RESUMO

A 12-month pilot project for notifiable avian disease (NAD) exclusion testing in chicken and turkey flocks in Great Britain (GB) offered, in partnership with industry, opportunities to carry out differential diagnosis in flocks where NAD was not suspected, and to identify undetected or undiagnosed infections. In May 2014, clinical samples received from a broiler breeder chicken premises that had been experiencing health and production problems for approximately one week tested positive by avian influenza (AI) real-time reverse transcription polymerase chain reaction (RRT-PCR). Following immediate escalation to an official, statutory investigation to rule out the presence of notifiable AI virus (AIV; H5 or H7 subtypes), a non-notifiable H4N6 low pathogenicity (LP) AIV was detected through virus isolation in embryonated specific pathogen free (SPF) fowls' eggs, neuraminidase inhibition test, cleavage site sequencing and AIV subtype H4-specific serology. Premises movement restrictions were lifted, and no further disease control measures were implemented as per the United Kingdom (UK) legislation. Phylogenetic analysis of the haemagglutinin and neuraminidase genes of the virus revealed closest relationships to viruses from Mallard ducks in Sweden during 2007 and 2009. In June 2014, clinical suspicion of NAD was reported in a flock of free-range laying chickens elsewhere in GB, due to increasing daily mortality and reduced egg production over a five-day period. An H4N6 LPAIV with an intravenous pathogenicity index of 0.50 was isolated. This virus was genetically highly similar, but not identical, to the virus detected during May 2014. Full viral genome analyses showed characteristics of a strain that had not recently transferred from wild birds, implying spread within the poultry sector had occurred. A stalk deletion in the neuraminidase gene sequence indicated an adaptation of the virus to poultry. Furthermore, there was unexpected evidence of systemic spread of the virus on post-mortem. No other cases were reported. Infection with LPAIVs often result in variable clinical presentation in poultry, making detection of disease more difficult.


Assuntos
Vírus da Influenza A/genética , Vírus da Influenza A/isolamento & purificação , Influenza Aviária/epidemiologia , Doenças das Aves Domésticas/diagnóstico , Doenças das Aves Domésticas/epidemiologia , Animais , Animais Selvagens/virologia , Galinhas/virologia , Diagnóstico Diferencial , Surtos de Doenças/prevenção & controle , Surtos de Doenças/veterinária , Monitoramento Epidemiológico , Hemaglutininas/genética , Sequenciamento de Nucleotídeos em Larga Escala , Vírus da Influenza A/crescimento & desenvolvimento , Vírus da Influenza A/patogenicidade , Influenza Aviária/virologia , Neuraminidase/genética , Filogenia , Projetos Piloto , Aves Domésticas/virologia , Doenças das Aves Domésticas/virologia , Reação em Cadeia da Polimerase em Tempo Real , Perus/virologia
16.
Vet Microbiol ; 224: 8-16, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30269795

RESUMO

Two reassortant H5N2 viruses in which hemagglutinin (HA) was clustered into clade 2.3.4.4, were isolated from apparently healthy waterfowl in live poultry markets in Eastern China in 2016. We used specific pathogen-free chickens, mallard ducks, and BALB/c mice to evaluate the isolates' biological characteristics in different animal models. The newly isolated reassortant H5N2 viruses were able to cause severe disease in chickens and effective contact transmission, only at high doses. Our pathogenicity studies in ducks yielded an interesting result: the intravenous pathogenicity index (IVPI) indicated that isolate A/goose/Eastern China/1106/2016(1106) was low pathogenic and the other isolate A/duck/Eastern China/YD1516/2016(YD1516) was of highly pathogenicity in ducks. However, our 50% duck lethal dose (DLD50) experiment demonstrated that these viruses were all of low pathogenicity (DLD50 > 107.0 EID50) in ducks. Additionally, despite the fact that reassortant H5N2 were of low pathogenicity in mice, they could bind to both avian-type (SAα-2,3 Gal) and human-type (SAα-2,6 Gal) receptors, suggesting that these isolates still present a high risk for human infection. Therefore, it is of great importance to implement continual surveillance of avian influenza virus (AIV) to protect both veterinary and public health.


Assuntos
Galinhas/virologia , Patos/virologia , Vírus da Influenza A Subtipo H5N2/genética , Vírus da Influenza A Subtipo H5N2/patogenicidade , Influenza Aviária/virologia , Vírus Reordenados/genética , Animais , China/epidemiologia , Feminino , Genoma Viral , Hemaglutininas/genética , Humanos , Vírus da Influenza A Subtipo H5N2/isolamento & purificação , Influenza Aviária/epidemiologia , Influenza Aviária/transmissão , Camundongos , Camundongos Endogâmicos BALB C , Filogenia , Aves Domésticas/virologia , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Ligação Viral
17.
Life Sci ; 213: 158-165, 2018 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-30352241

RESUMO

Aim To investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). MAIN METHODS: H1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized- metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. KEY FINDINGS: Purified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. SIGNIFICANCE: H1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.


Assuntos
Alérgenos/uso terapêutico , Antígenos de Dermatophagoides/metabolismo , Proteínas de Artrópodes/metabolismo , Hemaglutininas/metabolismo , Alérgenos/metabolismo , Animais , Antígenos de Dermatophagoides/genética , Proteínas de Artrópodes/genética , Quimera , Epitopos/metabolismo , Escherichia coli/imunologia , Engenharia Genética/métodos , Hemaglutininas/genética , Humanos , Hipersensibilidade/imunologia , Imunoglobulina E/imunologia , Imunoterapia/métodos , Camundongos , Camundongos Endogâmicos BALB C , Orthomyxoviridae/metabolismo , Pyroglyphidae/imunologia , Proteínas Recombinantes/genética
18.
J Virol ; 92(24)2018 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-30258006

RESUMO

The H1N1 influenza virus responsible for the most recent pandemic in 2009 (H1N1pdm) has spread to swine populations worldwide while it replaced the previous seasonal H1N1 virus in humans. In France, surveillance of swine influenza A viruses in pig herds with respiratory outbreaks led to the detection of 44 H1N1pdm strains between 2009 and 2017, regardless of the season, and findings were not correlated with pig density. From these isolates, 17 whole-genome sequences were obtained, as were 6 additional hemagglutinin (HA)/neuraminidase (NA) sequences, in order to perform spatial and temporal analyses of genetic diversity and to compare evolutionary patterns of H1N1pdm in pigs to patterns for human strains. Following mutation accumulation and fixation over time, phylogenetic analyses revealed for the first time the divergence of a swine-specific genogroup within the H1N1pdm lineage. The divergence is thought to have occurred around 2011, although this was demonstrated only through strains isolated in 2015 to 2016 in the southern half of France. To date, these H1N1pdm swine strains have not been related to any increased virulence in swine herds and have not exhibited any antigenic drift compared to seasonal human strains. However, further monitoring is encouraged, as diverging evolutionary patterns in these two species, i.e., swine and humans, may lead to the emergence of viruses with a potentially higher risk to both animal and human health.IMPORTANCE Pigs are a "mixing vessel" for influenza A viruses (IAVs) because of their ability to be infected by avian and human IAVs and their propensity to facilitate viral genomic reassortment events. Also, as IAVs may evolve differently in swine and humans, pigs can become a reservoir for old human strains against which the human population has become immunologically naive. Thus, viruses from the novel swine-specific H1N1pdm genogroup may continue to diverge from seasonal H1N1pdm strains and/or from other H1N1pdm viruses infecting pigs and lead to the emergence of viruses that would not be covered by human vaccines and/or swine vaccines based on antigens closely related to the original H1N1pdm virus. This discovery confirms the importance of encouraging swine IAV monitoring because H1N1pdm swine viruses could carry an increased risk to both human and swine health in the future as a whole H1N1pdm virus or gene provider in subsequent reassortant viruses.


Assuntos
Vírus da Influenza A Subtipo H1N1/classificação , Infecções por Orthomyxoviridae/epidemiologia , Doenças dos Suínos/virologia , Sequenciamento Completo do Genoma/métodos , Animais , Evolução Molecular , França/epidemiologia , Hemaglutininas/genética , Vírus da Influenza A Subtipo H1N1/genética , Neuraminidase/genética , Infecções por Orthomyxoviridae/virologia , Pandemias , Filogenia , Vigilância da População , Análise Espaço-Temporal , Suínos , Doenças dos Suínos/epidemiologia , Proteínas Virais/genética , Sequenciamento Completo do Genoma/veterinária
19.
Infect Immun ; 86(11)2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30181349

RESUMO

Mycoplasma gallisepticum, the primary etiologic agent of chronic respiratory disease, is a significant poultry pathogen, causing severe inflammation and leading to economic losses worldwide. Immunodominant proteins encoded by the variable lipoprotein and hemagglutinin (vlhA) gene family are thought to be important for M. gallisepticum-host interaction, pathogenesis, and immune evasion, but their exact role remains unknown. Previous work has demonstrated that vlhA phase variation is dynamic throughout the earliest stages of infection, with vlhA 3.03 being the predominant vlhA expressed during the initial infection, and that the pattern of dominant vlhA expression may be nonrandom and regulated by previously unrecognized mechanisms. To further investigate this gene family, we assessed the vlhA profile of two well-characterized vaccine strains, GT5 and Mg7, a vlhA 3.03 mutant strain, and an M. gallisepticum population expressing an alternative immunodominant vlhA Here, we report that two M. gallisepticum vaccine strains show different vlhA profiles over the first 2 days of infection compared to that of wild-type Rlow, while the population expressing an alternative immunodominant vlhA gene reverted to a profile indistinguishable from that of wild-type Rlow Additionally, we observed a slight shift in the vlhA gene expression profile but no reduction in virulence in a vlhA 3.03 mutant. Taken together, these data further support the hypothesis that M. gallisepticum vlhA genes change in a nonstochastic temporal progression of expression and that vlhA 3.03, while preferred, is not required for virulence. Collectively, these data may be important in elucidating mechanisms of colonization and overall pathogenesis of M. gallisepticum.


Assuntos
Proteínas de Bactérias/biossíntese , Regulação Bacteriana da Expressão Gênica , Hemaglutininas/biossíntese , Lipoproteínas/biossíntese , Infecções por Mycoplasma/veterinária , Mycoplasma gallisepticum/genética , Doenças das Aves Domésticas/microbiologia , Animais , Variação Antigênica , Proteínas de Bactérias/genética , Galinhas , Perfilação da Expressão Gênica , Hemaglutininas/genética , Lipoproteínas/genética , Família Multigênica , Infecções por Mycoplasma/microbiologia , Mycoplasma gallisepticum/metabolismo , Doenças das Aves Domésticas/patologia
20.
Mol Biol (Mosk) ; 52(4): 644-658, 2018.
Artigo em Russo | MEDLINE | ID: mdl-30113030

RESUMO

To study the pathogenicity factors of the pandemic A(H1N1) influenza virus, a number of mutant variants of the A/Hamburg/5/2009 (H1N1)pdm09 strain were obtained through passage in chicken embryos, mouse lungs, and MDCK cell culture. After 17 lung-to-lung passages of the A/Hamburg/5/2009 in mice, the minimum lethal dose of the derived variant decreased by five orders of magnitude compared to that of the parental virus. This variant differed from the original virus by nine amino acid residues in the following viral proteins: hemagglutinin (HA), neuraminidase (NA), and components of the polymerase complex. Additional passaging of the intermediate variants and cloning made it possible to obtain pairs of strains that differed by a single amino acid substitution. Comparative analysis of replicative activity, receptor specificity, and virulence of these variants revealed two mechanisms responsible for increased pathogenicity of the virus for mice. Thus, (1) substitutions in HA (Asp225Gly or Gln226Arg) and compensatory mutation decreasing the charge of HA (Lys123Asn, Lys157Asn, Gly158Glu, Asn159Asp, or Lys212Met) altered viral receptor-binding specificity and restored the functional balance between HA and NA; (2) Phe35Leu substitution in the PA protein increased viral polymerase activity.


Assuntos
Hemaglutininas/genética , Vírus da Influenza A Subtipo H1N1/genética , Influenza Humana/genética , Neuraminidase/genética , RNA Replicase/genética , Proteínas Virais/genética , Substituição de Aminoácidos/genética , Animais , Galinhas , Cães , Humanos , Vírus da Influenza A Subtipo H1N1/patogenicidade , Influenza Humana/virologia , Células Madin Darby de Rim Canino , Camundongos , Mutação , Replicação Viral/genética
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