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1.
Artigo em Inglês | MEDLINE | ID: mdl-31694758

RESUMO

Ebola virus (EBOV) is a zoonotic pathogen, the infection often results in severe, potentially fatal, systematic disease in human and nonhuman primates. VP35, an essential viral RNA-dependent RNA polymerase cofactor, is indispensable for Ebola viral replication and host innate immune escape. In this study, VP35 was demonstrated to be phosphorylated at Serine/Threonine by immunoblotting, and the major phosphorylation sites was S187, S205, T206, S208 and S317 as revealed by LC-MS/MS. By an EBOV minigenomic system, EBOV minigenome replication was shown to be significantly inhibited by the phosphorylation-defective mutant, VP35 S187A, but was potentiated by the phosphorylation mimic mutant VP35 S187D. Together, our findings demonstrate that EBOV VP35 is phosphorylated on multiple residues in host cells, especially on S187, which may contribute to efficient viral genomic replication and viral proliferation.

2.
Viruses ; 11(10)2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31597372

RESUMO

Rift Valley fever virus (RVFV), which causes Rift Valley fever (RVF), is a mosquito-borne zoonotic pathogen that causes serious morbidity and mortality in livestock and humans. RVF is a World Health Organization (WHO) priority disease and, together with rabies, is a major health burden in Africa. Here, we present the development and characterization of an inactivated recombinant RVFV and rabies virus (RABV) vaccine candidate (rSRV9-eGn). Immunization with rSRV9-eGn stimulated the production of RVFV-specific IgG antibodies and induced humoral and cellular immunity in mice but did not induce the production of neutralizing antibodies. IgG1 and IgG2a were the main isotypes observed by IgG subtype detection, and IgG3 antibodies were not detected. The ratios of IgG1/IgG2a > 1 indicated a Type 2 humoral immune response. An effective vaccine is intended to establish a long-lived population of memory T cells, and mice generated memory cells among the proliferating T cell population after immunization with rSRV9-eGn, with effector memory T cells (TEM) as the major population. Due to the lack of prophylactic treatment experiments, it is impossible to predict whether this vaccine can protect animals from RVFV infection with only high titres of anti-RVFV IgG antibodies and no neutralizing antibodies induced, and thus, protection confirmation needs further verification. However, this RVFV vaccine designed with RABV as the vector provides ideas for the development of vaccines that prevent RVFV and RABV infections.

3.
Viruses ; 11(10)2019 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-31590353

RESUMO

Peste des petits ruminants is a highly contagious acute or subacute disease of small ruminants caused by the peste des petits ruminants virus (PPRV), and it is responsible for significant economic losses in animal husbandry. Vaccination represents the most effective means of controlling this disease, with virus-like particle (VLP) vaccines offering promising vaccine candidates. In this study, a PPRV VLP-based vaccine was developed using a baculovirus expression system, allowing for the simultaneous expression of the PPRV matrix (M), hemagglutinin (H), fusion (F) and nucleocapsid (N) proteins in insect cells. Immunization of mice and goats with PPRV VLPs elicited a robust neutralization response and a potent cellular immune response. Mouse studies demonstrated that VLPs induced a more robust IFN-γ response in CD4+ and CD8+ T cells than PPRV Nigeria 75/1 and recruited and/or activated more B cells and dendritic cells in inguinal lymph nodes. In addition, PPRV VLPs induced a strong Th1 class response in mice, as indicated by a high IgG2a to IgG1 ratio. Goat studies demonstrated that PPRV VLPs can induce the production of antibodies specific for F and H proteins and can also stimulate the production of virus neutralizing antibodies to the same magnitude as the PPRV Nigeria 75/1 vaccine. Higher amounts of IFN-γ in VLP-immunized animal serum suggested that VLPs also elicited a cellular immune response in goats. These results demonstrated that VLPs elicit a potent immune response against PPRV infection in small ruminants, making PPRV VLPs a potential candidate for PPRV vaccine development.

4.
BMC Vet Res ; 15(1): 316, 2019 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477101

RESUMO

BACKGROUND: Canine distemper (CD) is an acute infectious disease with high morbidity rates caused by a highly contagious pathogen (Canine Morbillivirus, also known as canine distemper virus, CDV). CDV can infect a broad range of carnivores resulting in complex clinical signs. Currently, there is no effective method to treat for CDV infections. Favipiravir (T-705), a pyrazine derivative, was shown to be an effective antiviral drug against RNA viruses, acting on RNA-dependent RNA polymerase (RdRp). However, whether the T-705 has antiviral effects following CDV infection is unclear. Here, we investigated the antiviral effect of T-705 against CDV-3 and CDV-11 strains in Vero and DH82 cell lines. RESULTS: Our data demonstrated that T-705 significantly inhibited the replication of CDV-3 and CDV-11 in both Vero and DH82 cells at different concentrations, ranging from 2.441 µg/ml to 1250 µg/ml. Additionally, T-705 exhibited efficacious antiviral effects when administered at different time points after virus infection. Cytotoxicity tests showed a slight decline in viability in Vero cells after T-705 treatment, and no apparent cytotoxicity was detected in T-705 treated DH82 cells. Comparison of anti-CDV polyclonal serum only inhibition of CDV in supernatant, T-705 directly inhibited viral replication in cells, and indirectly reduced the amount of virions in supernatant. The combination application of T-705 and anti-CDV polyclonal serum exhibited a rapid and robust inhibition against virions in supernatant and virus replication in cells. CONCLUSIONS: Our data strongly indicated that T-705 effectively inhibited viral replication following CDV infection in vitro, and could be a potential candidate for treatment for CD.

5.
Viruses ; 11(9)2019 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-31470645

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV), a new coronavirus that has been causing severe and fatal acute respiratory illnesses in humans since its outbreak in 2012, has raised public fear worldwide. The development of prophylactics and therapeutics is urgently needed to prevent and control MERS-CoV infections. In this study, a bacterium (Lactococcus lactis)-like particle (BLP) vaccine displaying the MERS-CoV receptor-binding domain (RBD) was developed, and gram-positive enhancer matrix (GEM) particles were used as substrates to externally bind to the MERS-CoV RBD through a protein anchor (PA). The designs included different numbers of lysin motif (LysM) repeats in the PAs linked by linkers (RBD-linker-PA2 (RLP2), RBD-linker-PA3 (RLP3) and RBD-PA3 (RP3)), and three LysM repeats and a linker in the fusion proteins increased the binding activity to the RBD. The specific immune responses were tested by intranasally immunizing mice with RLP3-GEM with or without the adjuvant GEL01. The results showed that GEL01-adjuvanted RLP3-GEM increased the systemic humoral, cellular and local mucosal immune responses in the mouse model, especially in the intestinal tract. The above results indicate that the MERS-CoV BLP product has the potential to be developed into a promising mucosal candidate vaccine to protect against MERS-CoV infections.

6.
Virus Res ; 270: 197638, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31173772

RESUMO

Adjuvants are important components of vaccination strategies because they boost and accelerate the immune response. The aim of this study was to investigate the adjuvant activity of PCP-II, a polysaccharide isolated from Poria cocos, together with an inactivated rabies vaccine. The polysaccharide PCP-II was compared with the common veterinary rabies vaccine adjuvant Alhydrogel by co-administration of either adjuvant with the inactivated rabies virus rCVS-11-G to mice via the intramuscular route. Blood samples were collected to determine the virus-neutralizing antibody (VNA) titer and assess activation of B and T lymphocytes. Inguinal lymph node samples were collected, and proliferation of B lymphocytes was measured. Splenocytes were isolated, and antigen-specific cellular immune responses were evaluated by enzyme-linked immunospot and immunosorbent assays (ELISpot assay and ELISA, respectively). The results showed that PCP-II enhanced and promoted an increase in the VNA titer in the mice compared to Alhydrogel. Flow cytometry assays revealed that the polysaccharide activated more B lymphocytes in the lymph nodes and more B and T lymphocytes in the blood. Assessment of antigen-specific cellular immune responses showed that PCP-II strongly induced T lymphocyte proliferation in the spleen and high levels of cytokine secretion from splenocytes. All of these data suggest that PCP-II possesses excellent adjuvant activity and enhances both cellular and humoral immunity in mice. After examining the adjuvant activities of PCP-II in mice, dogs were immunized with rCVS-11-G together with Alhydrogel or PCP-II as an adjuvant; the control group was injected with a commercial rabies vaccine. Serum samples were collected, and the VNA titers were measured. PCP-II caused increases in the VNA titers in both the booster and single-dose immunization tests when co-administered with rCVS-11-G compared with Alhydrogel. The VNA titer of the commercial vaccine group was also significantly lower than that of the PCP-II group. These data indicate that PCP-II is an excellent candidate adjuvant for inactive rabies vaccines in the veterinary setting.

7.
Virus Genes ; 55(4): 550-556, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31161411

RESUMO

Japanese encephalitis virus SA14-14-2 (JEV SA14-14-2) is a widely used vaccine in China and other southeastern countries to prevent Japanese encephalitis in children. In this study, a stable infectious cDNA clone of JEV SA14-14-2 with a low copy number pACYC177 vector dependent on the T7 promoter and T7 terminator was developed. Two introns were inserted into the capsid gene and envelope gene of JEV cDNA for gene stability. Hepatitis delta virus ribozyme (HDVr) was engineered into the 3' UTR cDNA of JEV for authentic 3' UTR transcription. The rescued virus showed biological properties indistinguishable from those of the parent strain (JEV SA14-14-2). The establishment of a JEV SA14-14-2 reverse genetics system lays the foundation for the further development of other flavivirus vaccines and viral pathogenesis studies.

8.
Arch Virol ; 164(8): 2023-2029, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31111259

RESUMO

We previously obtained mouse-adapted variants of H1N2 avian influenza virus that contained PB2-L134H, PB2-I647L, PB2-D701N, HA-G228S, and M1-D231N mutations. Here, we analyzed the effects of these mutations on viral pathogenicity in a mammalian model. By evaluating the virulence of mouse-adapted H1N2 variants at different generations, we found that the PB2-D701N and HA-G228S mutations both contribute to the virulence of this virus in mammals. Furthermore, we found that the PB2-D701N and HA-G228S mutations both enhance the ability of the virus to replicate in vivo and in vitro and that the PB2-D701N substitution results in an expansion of viral tissue tropism. These results suggest that the PB2-D701N mutation and the HA-G228S mutation are the major mammalian determinants of H1N2 virus. These results help us to understand more about the mechanisms by which influenza viruses adapt to mammals, and monitoring of these mutations can be used in continuous influenza surveillance to assess the pandemic potential of avian influenza virus variants.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H1N2/genética , Influenza Aviária/virologia , Mutação/genética , Proteínas Virais/genética , Virulência/genética , Adaptação Biológica/genética , Substituição de Aminoácidos/genética , Animais , Aves , Linhagem Celular , Cães , Feminino , Células Madin Darby de Rim Canino , Mamíferos/virologia , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/virologia , Fenótipo , Replicação Viral/genética
9.
J Virol ; 93(5)2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30541860

RESUMO

Ebola virus (EBOV) infections result in aggressive hemorrhagic fever in humans, with fatality rates reaching 90% and with no licensed specific therapeutics to treat ill patients. Advances over the past 5 years have firmly established monoclonal antibody (MAb)-based products as the most promising therapeutics for treating EBOV infections, but production is costly and quantities are limited; therefore, MAbs are not the best candidates for mass use in the case of an epidemic. To address this need, we generated EBOV-specific polyclonal F(ab')2 fragments from horses hyperimmunized with an EBOV vaccine. The F(ab')2 was found to potently neutralize West African and Central African EBOV in vitro Treatment of nonhuman primates (NHPs) with seven doses of 100 mg/kg F(ab')2 beginning 3 or 5 days postinfection (dpi) resulted in a 100% survival rate. Notably, NHPs for which treatment was initiated at 5 dpi were already highly viremic, with observable signs of EBOV disease, which demonstrated that F(ab')2 was still effective as a therapeutic agent even in symptomatic subjects. These results show that F(ab')2 should be advanced for clinical testing in preparation for future EBOV outbreaks and epidemics.IMPORTANCE EBOV is one of the deadliest viruses to humans. It has been over 40 years since EBOV was first reported, but no cure is available. Research breakthroughs over the past 5 years have shown that MAbs constitute an effective therapy for EBOV infections. However, MAbs are expensive and difficult to produce in large amounts and therefore may only play a limited role during an epidemic. A cheaper alternative is required, especially since EBOV is endemic in several third world countries with limited medical resources. Here, we used a standard protocol to produce large amounts of antiserum F(ab')2 fragments from horses vaccinated with an EBOV vaccine, and we tested the protectiveness in monkeys. We showed that F(ab')2 was effective in 100% of monkeys even after the animals were visibly ill with EBOV disease. Thus, F(ab')2 could be a very good option for large-scale treatments of patients and should be advanced to clinical testing.

10.
Virus Genes ; 2018 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-30519855

RESUMO

Feline panleukopenia virus (FPV) infects cats and can be fatal to kittens. There is evidence that canine parvovirus originated from FPV, which makes FPV important in studies of the family Parvoviridae. In the present study, the entire genome of FPV strain HH-1/86 was converted into a full-length infectious clone (pFPV). The FPV strain HH-1/86 has a 5123-nt single stranded DNA genome with a Y-shaped inverted 3' terminal repeat (ITR) and a U-shaped inverted 5' ITR. Feline kidney cells (F81) were transfected with the pFPV clone which contained a genetic marker, and a rescued virus was obtained (rFPV). The rFPV was identified by its cytopathic effects, indirect immunofluorescence, growth curve analysis, western blot assay and hemagglutination, and was indistinguishable from the parent virus. The FPV infectious clone will facilitate the study of pathogenicity and viral replication of FPV and the inter-species transmission of parvoviruses.

11.
BMC Microbiol ; 18(1): 224, 2018 12 27.
Artigo em Inglês | MEDLINE | ID: mdl-30587113

RESUMO

BACKGROUND: Bovine ephemeral fever virus (BEFV), the causative agent of bovine ephemeral fever, is an economically important pathogen of cattle and water buffalo. MicroRNAs (miRNAs) are endogenous 21-23 nt small non-coding RNA molecules that binding to a multiple of target mRNAs and functioning in the regulation of viral replication including the miRNA-mediated antiviral defense. However, the reciprocal interaction between bovine ephemeral fever virus replication and host miRNAs still remain poorly understood. The aim of our study herein was to investigate the exact function of miR-3470b and its molecular mechanisms during BEFV infection. RESULTS: In this study, we found a set of microRNAs induced by BEFV infection using small RNA deep sequencing, and further identified BEFV infection could significantly up-regulate the miR-3470b expression in Baby Hamster Syrian Kidney cells (BHK-21) after 24 h and 48 h post-infection (pi) compared to normal BHK-21 cells without BEFV infection. Additionally, the target association between miR-3470b and mitochondrial antiviral signaling protein (MAVS) was predicted by target gene prediction tools and further validated using a dual-luciferase reporter assay, and the expression of MAVS mRNA and protein levels was negatively associated with miR-3470b levels. Furthermore, the miR-3470b mimic transfection significantly contributed to increase the BEFV N mRNA, G protein level and viral titer, respectively, whereas the miR-3470b inhibitor had the opposite effect on BEFV replication. Moreover, the overexpression of MAVS or silencing of miR-3470b by its inhibitors suppressed BEFV replication, and knockdown of MAVS by small interfering RNA also promoted the replication of BEFV. CONCLUSIONS: Our findings is the first to reveal that miR-3470b as a novel host factor regulates BEFV replication via directly targeting the MAVS gene in BHK-21 cells and may provide a potential strategy for developing effective antiviral therapy.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/imunologia , Vírus da Febre Efêmera Bovina/fisiologia , Febre Efêmera/imunologia , Febre Efêmera/virologia , Rim/imunologia , MicroRNAs/genética , Replicação Viral , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Bovinos , Cricetinae , Febre Efêmera/genética , Vírus da Febre Efêmera Bovina/genética , Interações Hospedeiro-Patógeno , Rim/virologia , Mesocricetus , MicroRNAs/imunologia , Coelhos
12.
Vaccine ; 36(40): 5990-5998, 2018 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-30172635

RESUMO

We previously demonstrated that intramuscular immunization with virus-like particles (VLPs) composed of the haemagglutinin (HA), neuraminidase (NA), and matrix (M1) proteins of A/meerkat/Shanghai/SH-1/2012 (clade 2.3.2.1) protected mice from lethal challenge with viruses from other H5 HPAI clades. The inclusion of additional proteins that can serve as immunological adjuvants in VLPs may enhance adaptive immune responses following vaccination, and oral vaccines may represent the safest choice. Here, we report the generation of H5N1 VLPs composed of the viral HA, NA, and M1 proteins and membrane-anchored forms of the Escherichia coli heat-labile enterotoxin B subunit protein (LTB) or the Toll-like receptor 5 ligand flagellin (Flic). Mice intramuscularly or orally immunized with VLPs containing LTB or Flic generated greater humoural and cellular immune responses than those administered H5N1 VLPs without LTB or Flic. Intramuscular immunization with VLPs protected mice from lethal challenge with homologous or heterologous H5N1 viruses irrespective of whether the VLPs additionally included LTB or Flic. In contrast, oral immunization of mice with LTB- or Flic-VLPs conferred substantial protection against lethal challenge with both homologous and heterologous H5N1 influenza viruses, whereas mice immunized orally with VLPs lacking LTB and Flic universally succumbed to infection. Mice immunized orally with LTB- or Flic-VLPs showed 10-fold higher virus-specific IgG titres than mice immunized with H5N1-VLPs lacking LTB or Flic. Collectively, these results indicate that the inclusion of immunostimulatory proteins, such as LTB and Flic, in VLP-based vaccines may represent a promising new approach for the control of current H5N1 HPAI outbreaks by eliciting higher humoural and cellular immune responses and conferring improved cross-clade protection.

13.
Int Immunopharmacol ; 64: 217-222, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30199846

RESUMO

BACKGROUND: Rift Valley fever virus (RVFV) is an emerging arbovirus in Africa and the Arabian Peninsula, in which infection with RVFV poses a serious threat to humans and livestock globally. Approved treatments for RVFV infection, especially for use in humans, have not yet been developed. There is an urgent need for effective drugs to prevent RVFV disease. METHODS: In previous study, we developed RVFV virus like particles (VLPs) expressing the surface glycoproteins Gn and Gc. The morphology was shown to be similar to live RVFV under electron microscopy. In this study, we immunized horses with RVFV VLPs, prepared the immunoglobulin F(ab')2 fragments, and characterized its in vitro neutralization and in vivo efficacy in mice. RESULTS: F(ab')2 was found to potently neutralize RVFV in VeroE6 cells, and passive transfer of immunoglobulin F(ab')2 fragments resulting in reduced mortality in RVFV infected mice. CONCLUSION: Our results show that passive immunotherapy with equine immunoglobulin F(ab')2 fragments is a promising strategy to treat RVFV infections.

14.
Mol Cell Probes ; 41: 22-26, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30138696

RESUMO

Bovine respirovirus 3 also known as Bovine parainfluenza virus type 3 (BPIV3) is one of the most important viral respiratory agents of both young and adult cattle. Rapid diagnosis could contribute greatly in containing epidemics and thus avoid economic losses. However, the lack of robust isothermal visual method poses difficulty. In this study, a novel isothermal assay for detecting BPIV3 was established. The method includes a lateral flow dipstick (LFD) assay combined with reverse transcription recombinase polymerase amplification (RT-RPA). First, the analytical sensitivity and specificity of BPIV3 LFD RT-RPA were tested. The LFD RT-RPA assay has a detection limit of up to 100 copies per reaction in 30 min at 38 °C. Then the performance of LFD RT-RPA was evaluated using 95 clinical samples. Compared to qPCR, the LFD RT-RPA assay showed a clinical sensitivity of 94.74%, a clinical specificity of 96.05% and 0.8734 kappa coefficient. These results have demonstrated the efficiency and effectiveness of the method to be developed into a point of care protocol for the diagnosis of BPIV3.

15.
Sci Rep ; 8(1): 11858, 2018 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-30089776

RESUMO

Rabies, caused by rabies virus (RABV), is still the deadliest infectious disease. Mechanism of host immune response upon RABV infection is not yet fully understood. Accumulating evidences suggest that long noncoding RNAs (lncRNAs) plays key roles in host antiviral responses. However, expression profile and function of lncRNAs in RABV infection remain unclear. In the present study, expression profile of lncRNAs and mRNAs profiles were investigated in RABV-infected brain tissues of mice by RNA sequencing. A total of 140 lncRNAs and 3,807 mRNAs were differentially expressed in RABV-infected animals. The functional annotation and enrichment analysis using Gene Oncology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that differentially expressed transcripts were predominantly involved in signaling pathways related to host immune response. The expression profiles of the selected lncRNAs in brains of mice during RABV infections were verified by quantitative real time polymerase chain reaction (qRT-PCR). To our knowledge, this is the first report to profile the lncRNA expression in RABV infected mice. Our findings provide insights into understanding the role of lncRNAs in host immune response against RABV infection.

16.
Int Immunopharmacol ; 63: 119-128, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30081250

RESUMO

Influenza viruses cause significant morbidity and mortality and pose a substantial threat to public health. Vaccination represents the principle means of preventing influenza virus infection. Current vaccine approaches are hindered by the need to routinely reformulate vaccine compositions in an effort to account for the progressive antigenic changes that occur as influenza viruses circulate in the human population. In this study, we evaluated chimeric virus-like particle (cVLP) vaccines containing conserved elements of influenza proteins (HL5M2e (HA stem gene with 5M2e gene inserted) and NP), with or without glycosylphosphatidylinositol-anchored CCL28 (GPI-CCL28) and/or GM-CSF (GPI-GM-CSF) fusion proteins as molecular adjuvants. cVLPs elicited strong humoral and cellular immune responses against homologous and heterologous viruses, and improved survival following lethal challenge with both homologous and heterologous viruses. Inclusion of GPI-anchored adjuvants in cVLP vaccines augmented the generation of influenza-specific humoral and cellular immune responses in mice in comparison to the non-adjuvanted cVLP vaccines. VLPs containing GPI-anchored adjuvants reduced morbidity and improved survival to lethal challenge with homologous and heterologous influenza viruses. This work suggests that VLP vaccines incorporating conserved influenza virus proteins and GPI-anchored molecular adjuvants may serve as a platform for a broadly protective "universal" influenza vaccine.

17.
Front Microbiol ; 9: 1101, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29896174

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel human coronavirus that can cause human respiratory disease. The development of a detection method for this virus that can lead to rapid and accurate diagnosis would be significant. In this study, we established a nucleic acid visualization technique that combines the reverse transcription loop-mediated isothermal amplification technique and a vertical flow visualization strip (RT-LAMP-VF) to detect the N gene of MERS-CoV. The RT-LAMP-VF assay was performed in a constant temperature water bath for 30 min, and the result was visible by the naked eye within 5 min. The RT-LAMP-VF assay was capable of detecting 2 × 101 copies/µl of synthesized RNA transcript and 1 × 101 copies/µl of MERS-CoV RNA. The method exhibits no cross-reactivities with multiple CoVs including SARS-related (SARSr)-CoV, HKU4, HKU1, OC43 and 229E, and thus exhibits high specificity. Compared to the real-time RT-PCR (rRT-PCR) method recommended by the World Health Organization (WHO), the RT-LAMP-VF assay is easy to handle, does not require expensive equipment and can rapidly complete detection within 35 min.

18.
Emerg Microbes Infect ; 7(1): 101, 2018 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-29872043

RESUMO

Ebolavirus vaccines based on several adenoviral vectors have been investigated in preclinical studies and clinical trials. The use of adenovirus serotype 2 as a vector for ebolavirus vaccine has not been reported. Herein, we generated rAd2-ZGP, a recombinant replication-incompetent adenovirus serotype 2 expressing codon-optimized Zaire ebolavirus glycoprotein, and evaluated its immunogenicity in mice and rhesus macaques. rAd2-ZGP induced significant antibody and cell-mediated immune responses at 2 weeks after a single immunization. The glycoprotein (GP)-specific immune responses could be further enhanced with a booster immunization. Compared to protein antigens, Zaire ebolavirus GP and Zaire ebolavirus-like particles, rAd2-ZGP could induce stronger cross-reactive antibody and cell-mediated immune responses to heterologous Sudan ebolavirus in mice and rhesus macaques. In rAd2-ZGP-immunized macaques, GP-specific CD8+ T cells could secret IFN-γ and IL-2, indicating a Th1-biased response. In adenovirus serotype 5 seropositive macaques, rAd2-ZGP could induce robust antibody and cell-mediated immune responses, suggesting that the efficacy of rAd2-ZGP is not affected by pre-existing immunity to adenovirus serotype 5. These results demonstrated that rAd2-ZGP can be considered an alternative ebolavirus vaccine for use in adenovirus serotype 5 seropositive subjects or as a sequential booster vaccine after the subjects have been immunized with a recombinant adenovirus serotype 5-based vaccine.


Assuntos
Adenoviridae/genética , Anticorpos Antivirais/imunologia , Ebolavirus/imunologia , Doença pelo Vírus Ebola/imunologia , Imunidade Celular , Vacinas Virais/imunologia , Adenoviridae/metabolismo , Animais , Modelos Animais de Doenças , Ebolavirus/genética , Feminino , Glicoproteínas/administração & dosagem , Glicoproteínas/genética , Glicoproteínas/imunologia , Doença pelo Vírus Ebola/prevenção & controle , Doença pelo Vírus Ebola/virologia , Humanos , Interferon gama/imunologia , Interleucina-2/imunologia , Macaca mulatta , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Linfócitos T/imunologia , Vacinação , Proteínas Virais/administração & dosagem , Proteínas Virais/genética , Proteínas Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
19.
Int Immunopharmacol ; 58: 109-116, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29571081

RESUMO

The H7N9 influenza virus epidemic has been associated with a high mortality rate in China. Therefore, to prevent the H7N9 virus from causing further damage, developing a safe and effective vaccine is necessary. In this study, a vaccine candidate consisting of virus-like particles (VLPs) based on H7N9 A/Shanghai/2/2013 and containing hemagglutinin (HA), neuraminidase (NA), and matrix protein (M1) was successfully produced using a baculovirus (BV) expression system. Immunization experiments showed that strong humoral and cellular immune responses could be induced by the developed VLPs when administered via either the intramuscular (IM) or intranasal (IN) immunization routes. Notably, VLPs administered via both immunization routes provided 100% protection against lethal infection caused by the H7N9 virus. The IN immunization with 40µg of H7N9 VLPs induced strong lung IgA and lung tissue resident memory (TRM) cell-mediated local immune responses. These results provide evidence for the development of an effective preventive vaccine against the H7N9 virus based on VLPs administered through both the IM and IN immunization routes.


Assuntos
Subtipo H7N9 do Vírus da Influenza A/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Infecções por Orthomyxoviridae/imunologia , Vírion/imunologia , Administração Intranasal , Animais , Anticorpos Antivirais/sangue , Feminino , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Humanos , Imunidade Celular , Imunização , Injeções Intramusculares , Camundongos , Camundongos Endogâmicos BALB C , Neuraminidase/imunologia , Proteínas da Matriz Viral/imunologia
20.
BMC Infect Dis ; 18(1): 43, 2018 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-29338693

RESUMO

BACKGROUND: Brucellosis is an endemic disease in the Inner Mongolia Autonomous Region of China and Ulanqab exhibits the highest prevalence of brucellosis in this region. Due to the complex nature of Brucellosis, a cure for this disease has proven to be elusive. Furthermore, the reduced susceptibility of Brucella spp. to antimicrobial agents has been reported as a potential cause of therapeutic failure. However, detailed in vitro antimicrobial susceptibility patterns pertaining to Brucella isolates from this region have not yet been published. The aim of this study was to evaluate the antibiotic susceptibility profile of Brucella melitensis clinical isolates from Ulanqab, Inner Mongolia, China. METHODS: A total of 85 B. melitesis isolates were obtained from humans in Ulanqab of Inner Mongolia, China; the antimicrobial susceptibility of 85 clinical isolates to nine antibiotics was assessed using the E-test method according to the CLSI (Clinical and Laboratory Standards Institute) guidelines. RESULTS: All of the tested isolates were susceptible to minocycline, sparfloxacin, doxycycline, tetracycline, ciprofloxacin, gentamicin and levofloxacin. Resistance to rifampin and cotrimoxazole was observed in 1.0% (1/85) and 7.0% (6/85) of the isolates, respectively. However, rpoB gene mutations were not observed in single isolates exhibiting resistance to rifampin. CONCLUSIONS: We observed that B. melitensis isolates are susceptible to the majority of the tested antibiotics. Furthermore, minocycline and sparfloxacin exhibited extremely high bactericidal effects in relation to the B. melitensis isolates. The sensitivity of commonly used drugs for the treatment of brucellosis should be regularly monitored. To the best of our knowledge, this is the first report of rifampin and cotrimoxazole resistant isolates of B. melitensis in China. In summary, based on the findings from this study, we suggest that antibiotic administration and use should be rationalized to prevent future drug resistance.


Assuntos
Antibacterianos/farmacologia , Brucella melitensis/efeitos dos fármacos , Brucella melitensis/isolamento & purificação , Brucelose/microbiologia , China , Farmacorresistência Bacteriana/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Rifampina/farmacologia , Combinação Trimetoprima e Sulfametoxazol/farmacologia
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