Evidence of bat origin for Menangle virus, a zoonotic paramyxovirus first isolated from diseased pigs

Menangle virus (MenPV) is a zoonotic paramyxovirus capable of causing disease in pigs and humans. It was first isolated in 1997 from stillborn piglets at a commercial piggery in New South Wales, Australia, where an outbreak of reproductive disease occurred. Neutralizing antibodies to MenPV were detected in various pteropid bat species in Australia and fruit bats were suspected to be the source of the virus responsible for the outbreak in pigs. However, previous attempts to isolate MenPV from various fruit bat species proved fruitless. Here, we report the isolation of MenPV from urine samples of the black flying fox, Pteropus alecto, using a combination of improved procedures and newly established bat cell lines. The nucleotide sequence of the bat isolate is 94% identical to the pig isolate. This finding provides strong evidence supporting the hypothesis that the MenPV outbreak in pigs originated from viruses in bats roosting near the piggery. © 2012 Printed in Great Britain.

Incremento del efecto citotóxico de un fármaco antineoplásico mediante la expresión de la glicoproteína fugogénica del paramyxovirus SV5

Tesis (Doctor en Ciencias con Orientación en Farmacia) UANL, 2010.

Estudo de Paramyxovirus, Mycoplasma e de bacilos Gram-negativos no trato respiratório de serpentes Crotalus durissus terrificus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Envelope protein dynamics in paramyxovirus entry.

Paramyxoviruses include major pathogens with significant global health and economic impact. This large family of enveloped RNA viruses infects cells by employing two surface glycoproteins that tightly cooperate to fuse their lipid envelopes with the target cell plasma membrane, an attachment and a fusion (F) protein. Membrane fusion is believed to depend on receptor-induced conformational changes within the attachment protein that lead to the activation and subsequent refolding of F. While structural and mechanistic studies have considerably advanced our insight into paramyxovirus cell adhesion and the structural basis of F refolding, how precisely the attachment protein links receptor engagement to F triggering remained poorly understood. Recent reports based on work with several paramyxovirus family members have transformed our understanding of the triggering mechanism of the membrane fusion machinery. Here, we review these recent findings, which (i) offer a broader mechanistic understanding of the paramyxovirus cell entry system, (ii) illuminate key similarities and differences between entry strategies of different paramyxovirus family members, and (iii) suggest new strategies for the development of novel therapeutics.

Peptides from conserved regions of paramyxovirus fusion (F) proteins are potent inhibitors of viral fusion.

The synthetic peptides DP-107 and DP-178 (T-20), derived from separate domains within the human immunodeficiency virus type 1 (HIV-1) transmembrane (TM) protein, gp4l, are stable and potent inhibitors of HIV-1 infection and fusion. Using a computer searching strategy (computerized antiviral searching technology, C.A.S.T.) based on the predicted secondary structure of DP-107 and DP-178 (T-20), we have identified conserved heptad repeat domains analogous to the DP-107 and DP-178 regions of HIV-1 gp41 within the glycoproteins of other fusogenic viruses. Here we report on antiviral peptides derived from three representative paramyxoviruses, respiratory syncytial virus (RSV), human parainfluenza virus type 3 (HPIV-3), and measles virus (MV). We screened crude preparations of synthetic 35-residue peptides, scanning the DP-178-like domains, in antiviral assays. Peptide preparations demonstrating antiviral activity were purified and tested for their ability to block syncytium formation. Representative DP-178-like peptides from each paramyxovirus blocked homologous virus-mediated syncytium formation and exhibited EC50 values in the range 0.015-0.250 microM. Moreover, these peptides were highly selective for the virus of origin. Identification of biologically active peptides derived from domains within paramyxovirus F1 proteins analogous to the DP-178 domain of HIV-1 gp4l is compelling evidence for equivalent structural and functional features between retroviral and paramyxoviral fusion proteins. These antiviral peptides provide a novel approach to the development of targeted therapies for paramyxovirus infections.

Hendra Virus and Horse Owners – Risk Perception and Management

Hendra virus is a highly pathogenic novel paramyxovirus causing sporadic fatal infection in horses and humans in Australia. Species of fruit-bats (genus Pteropus), commonly known as flying-foxes, are the natural host of the virus. We undertook a survey of horse owners in the states of Queensland and New South Wales, Australia to assess the level of adoption of recommended risk management strategies and to identify impediments to adoption. Survey questionnaires were completed by 1431 respondents from the target states, and from a spectrum of industry sectors. Hendra virus knowledge varied with sector, but was generally limited, with only 13% of respondents rating their level of knowledge as high or very high. The majority of respondents (63%) had seen their state’s Hendra virus information for horse owners, and a similar proportion found the information useful. Fifty-six percent of respondents thought it moderately, very or extremely likely that a Hendra virus case could occur in their area, yet only 37% said they would consider Hendra virus if their horse was sick. Only 13% of respondents stabled their horses overnight, although another 24% said it would be easy or very easy to do so, but hadn’t done so. Only 13% and 15% of respondents respectively had horse feed bins and water points under solid cover. Responses varied significantly with state, likely reflecting different Hendra virus history. The survey identified inconsistent awareness and/or adoption of available knowledge, confusion in relation to Hendra virus risk perception, with both over-and under-estimation of true risk, and lag in the uptake of recommended risk minimisation strategies, even when these were readily implementable. However, we also identified frustration and potential alienation by horse owners who found the recommended strategies impractical, onerous and prohibitively expensive. The insights gained from this survey have broader application to other complex risk-management scenarios.

Survival of Hendra Virus in the Environment: Modelling the Effect of Temperature

Hendra virus (HeV), a highly pathogenic zoonotic paramyxovirus recently emerged from bats, is a major concern to the horse industry in Australia. Previous research has shown that higher temperatures led to lower virus survival rates in the laboratory. We develop a model of survival of HeV in the environment as influenced by temperature. We used 20 years of daily temperature at six locations spanning the geographic range of reported HeV incidents to simulate the temporal and spatial impacts of temperature on HeV survival. At any location, simulated virus survival was greater in winter than in summer, and in any month of the year, survival was higher in higher latitudes. At any location, year-to-year variation in virus survival 24 h post-excretion was substantial and was as large as the difference between locations. Survival was higher in microhabitats with lower than ambient temperature, and when environmental exposure was shorter. The within-year pattern of virus survival mirrored the cumulative within-year occurrence of reported HeV cases, although there were no overall differences in survival in HeV case years and non-case years. The model examines the effect of temperature in isolation; actual virus survivability will reflect the effect of additional environmental factors

Multicenter Blinded Analysis of RT-PCR Detection Methods for Paramyxoviruses in Relation to Paget's Disease of Bone

Conflicting results have been reported on the detection of paramyxovirus transcripts in Paget's disease, and a possible explanation is differences in the sensitivity of RT-PCR methods for detecting virus. In a blinded study, we found no evidence to suggest that laboratories that failed to detect viral transcripts had less sensitive RT-PCR assays, and we did not detect measles or distemper transcripts in Paget's samples using the most sensitive assays evaluated.

Introduction: There is conflicting evidence on the possible role of persistent paramyxovirus infection in Paget's disease of bone (PDB). Some workers have detected measles virus (MV) or canine distemper virus (CDV) transcripts in cells and tissues from patients with PDB, but others have failed to confirm this finding. A possible explanation might be differences in the sensitivity of RT-PCR methods for detecting virus. Here we performed a blinded comparison of the sensitivity of different RT-PCR-based techniques for MV and CDV detection in different laboratories and used the most sensitive assays to screen for evidence of viral transcripts in bone and blood samples derived from patients with PDB.

Materials and Methods: Participating laboratories analyzed samples spiked with known amounts of MV and CDV transcripts and control samples that did not contain viral nucleic acids. All analyses were performed on a blinded basis.

Results: The limit of detection for CDV was 1000 viral transcripts in three laboratories (Aberdeen, Belfast, and Liverpool) and 10,000 transcripts in another laboratory (Manchester). The limit of detection for MV was 16 transcripts in one laboratory (NIBSC), 1000 transcripts in two laboratories (Aberdeen and Belfast), and 10,000 transcripts in two laboratories (Liverpool and Manchester). An assay previously used by a U.S.-based group to detect MV transcripts in PDB had a sensitivity of 1000 transcripts. One laboratory (Manchester) detected CDV transcripts in a negative control and in two samples that had been spiked with MV. None of the other laboratories had false-positive results for MV or CDV, and no evidence of viral transcripts was found on analysis of 12 PDB samples using the most sensitive RT-PCR assays for MV and CDV.

Conclusions: We found that RT-PCR assays used by different laboratories differed in their sensitivity to detect CDV and MV transcripts but found no evidence to suggest that laboratories that previously failed to detect viral transcripts had less sensitive RT-PCR assays than those that detected viral transcripts. False-positive results were observed with one laboratory, and we failed to detect paramyxovirus transcripts in PDB samples using the most sensitive assays evaluated. Our results show that failure of some laboratories to detect viral transcripts is unlikely to be caused by problems with assay sensitivity and highlight the fact that contamination can be an issue when searching for pathogens by sensitive RT-PCR-based techniques.

Infecciones respiratorias agudas en menores de 5 años vacunados o no contra la Influenza en el periodo del 2004 y 2005

De las enfermedades posibles que afectan el ser humano, las infecciones respiratorias agudas (IRA) son las más prevalentes de todas, con una prevalencia para Colombia según informes de la OPS del año 2005 de 12.6% en niños de 0 a 4 años. A pesar de la naturaleza en su mayoría son de corta duración y su desarrollo es benigno, con lleva repercusiones importantes en la salud y en lo económico: Es una de las principales causas de demanda de servicios médicos, de prescripción injustificada de antibióticos , de ausentismo escolar y laboral secundario. El objetivo de este estudio es comparar el número de eventos de infecciones respiratorias agudas, en niños entre 2 y 5 años de edad afiliados a la EPS Sanitas que consultaron al Clinisanitas de Ciudad Salitre entre el periodo de septiembre del 2004 y febrero del 2006, para determinar si existe o no asociación al hecho de estar vacunado contra la influenza.

Promotion of Hendra virus replication by microRNA 146a

Hendra virus is a highly pathogenic zoonotic paramyxovirus in the genus Henipavirus. Thirty-nine outbreaks of Hendra virus have been reported since its initial identification in Queensland, Australia, resulting in seven human infections and four fatalities. Little is known about cellular host factors impacting Hendra virus replication. In this work, we demonstrate that Hendra virus makes use of a microRNA (miRNA) designated miR-146a, an NF-κB-responsive miRNA upregulated by several innate immune ligands, to favor its replication. miR-146a is elevated in the blood of ferrets and horses infected with Hendra virus and is upregulated by Hendra virus in human cells in vitro. Blocking miR-146a reduces Hendra virus replication in vitro, suggesting a role for this miRNA in Hendra virus replication. In silico analysis of miR-146a targets identified ring finger protein (RNF)11, a member of the A20 ubiquitin editing complex that negatively regulates NF-κB activity, as a novel component of Hendra virus replication. RNA interference-mediated silencing of RNF11 promotes Hendra virus replication in vitro, suggesting that increased NF-κB activity aids Hendra virus replication. Furthermore, overexpression of the IκB superrepressor inhibits Hendra virus replication. These studies are the first to demonstrate a host miRNA response to Hendra virus infection and suggest an important role for host miRNAs in Hendra virus disease.

Use of reverse transcriptase polymerase chain reaction (RT-PCR) in molecular screening of Newcastle disease virus in poultry and free-living bird populations

The aim of this study was to evaluate a simple molecular method of reverse transcriptase polymerase chain reaction (RT-PCR) to differentiate Newcastle disease virus strains according to their pathogenicity, in order to use it in molecular screening of Newcastle disease virus in poultry and free-living bird populations. Specific primers were developed to differentiate LaSota-LS-(vaccine strain) and Sao Joao do Meriti-SJM-strain (highly pathogenic strain). Chickens and pigeons were experimentally vaccinated/infected for an in vivo study to determine virus shedding in feces. Validation of sensitivity and specificity of the primers (SJM and LS) by experimental models used in the present study and results obtained in the molecular analysis of the primers by BLAST made it possible to generalize results. The development of primers that differentiate the level of pathogenicity of NDV stains is very important, mainly in countries where real-time RT-PCR is still not used as a routine test. These primers were able to determine the presence of the agent and to differentiate it according to its pathogenicity. © 2012 Springer Science+Business Media B.V.

Molecular and phylogenetic characterization based on the complete genome of a virulent pathotype of Newcastle disease virus isolated in the 1970s in Brazil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estudo das interações entre proteína NS1 do hRSV e flavonóides utilizando métodos biofísicos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)