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PubMed | National Research Center Disciplinaria en Microbiologia Animal, National Autonomous University of Mexico, Free University of Berlin, SUNY Downstate Medical Center and Leibniz Institute for Zoo and Wildlife Research
Type: Journal Article | Journal: mBio | Year: 2016

Gammaherpesviruses (HVs) are generally considered host specific and to have codiverged with their hosts over millions of years. This tenet is challenged here by broad-scale phylogenetic analysis of two viral genes using the largest sample of mammalian HVs to date, integrating for the first time bat HV sequences available from public repositories and newly generated viral sequences from two vampire bat species (Desmodus rotundus and Diphylla ecaudata). Bat and primate viruses frequently represented deep branches within the supported phylogenies and clustered among viruses from distantly related mammalian taxa. Following evolutionary scenario testing, we determined the number of host-switching and cospeciation events. Cross-species transmissions have occurred much more frequently than previously estimated, and most of the transmissions were attributable to bats and primates. We conclude that the evolution of the Gammaherpesvirinae subfamily has been driven by both cross-species transmissions and subsequent cospeciation within specific viral lineages and that the bat and primate orders may have potentially acted as superspreaders to other mammalian taxa throughout evolutionary history.It has long been believed that herpesviruses have coevolved with their hosts and are species specific. Nevertheless, a global evolutionary analysis of bat viruses in the context of other mammalian viruses, which could put this widely accepted view to the test, had not been undertaken until now. We present two main findings that may challenge the current view of HV evolution: multiple host-switching events were observed at a higher rate than previously appreciated, and bats and primates harbor a large diversity of HVs which may have led to increased cross-species transmissions from these taxa to other mammals.


PubMed | Swedish University of Agricultural Sciences, Private Researcher, National Polytechnic Institute of Mexico and National Research Center Disciplinaria en Microbiologia Animal
Type: | Journal: Protein expression and purification | Year: 2016

Blue eye disease caused by Porcine rubulavirus (PorPV) is an endemic viral infection of swine causing neurological and respiratory disease in piglets, and reproductive failure in sows and boars. The hemagglutinin-neuraminidase (HN) glycoprotein of PorPV is the most abundant component in the viral envelope and the main target of the immune response in infected animals. In this study, we expressed the HN-PorPV-recombinant (rHN-PorPV) protein in an Escherichia coli system and analyzed the immune responses in mice. The HN gene was cloned from the reference strain PorPV-La Piedad Michoacan Virus (GenBank accession number BK005918), into the pDual expression vector. The expressed protein was identified at a molecular weight of 61.7kDa. Three-dimensional modeling showed that the main conformational and functional domains of the rHN-PorPV protein were preserved. The antigenicity of the expressed protein was confirmed by Western blot with a monoclonal antibody recognizing the HN, and by testing against serum samples from pigs experimentally infected with PorPV. The immunogenicity of the rHN-PorPV protein was tested by inoculation of BALB/c mice with AbISCO-100() as adjuvant. Analysis of the humoral immune responses in mice showed an increased level of specific antibodies 14 days after the first immunization, compared to the control group (P<0.0005). The results show the ability of the rHN-PorPV protein to induce an antibody response in mice. Due to its immunogenic potential, the rHN-PorPV protein will be further evaluated in pig trials for its suitability for prevention and control of blue eye disease.


Cuevas-Romero S.,National Research Center Disciplinaria en Microbiologia Animal | Cuevas-Romero S.,Swedish University of Agricultural Sciences | Rivera-Benitez J.F.,National Research Center Disciplinaria en Microbiologia Animal | Blomstrom A.-L.,Swedish University of Agricultural Sciences | And 5 more authors.
Virus Genes | Year: 2016

Since the report of the initial outbreak of Porcine rubulavirus (PorPV) infection in pigs, only one full-length genome from 1984 (PorPV-LPMV/1984) has been characterised. To investigate the overall genetic variation, full-length gene nucleotide sequences of current PorPV isolates were obtained from different clinical cases of infected swine. Genome organisation and sequence analysis of the encoded proteins (NP, P, F, M, HN and L) revealed high sequence conservation of the NP protein and the expression of the P and V proteins in all PorPV isolates. The V protein of one isolate displayed a mutation that has been implicated to antagonise the antiviral immune responses of the host. The M protein indicated a variation in a short region that could affect the electrostatic charge and the interaction with the membrane. One PorPV isolate recovered from the lungs showed a mutation at the cleavage site (HRKKR) of the F protein that could represent an important factor to determine the tissue tropism and pathogenicity of this virus. The HN protein showed high sequence identity through the years (up to 2013). Additionally, a number of sequence motifs of very high amino acid conservation among the PorPV isolates important for polymerase activity of the L protein have been identified. In summary, genetic comparisons and phylogenetic analyses indicated that three different genetic variants of PorPV are currently spreading within the swine population, and a new generation of circulating virus with different characteristics has begun to emerge. © 2015, Springer Science+Business Media New York.


Cuevas-Romero S.,Swedish University of Agricultural Sciences | Cuevas-Romero S.,National Research Center Disciplinaria en Microbiologia Animal | Hernandez-Baumgarten E.,National Autonomous University of Mexico | Kennedy S.,Agri Food and Biosciences Institute of Northern Ireland | And 3 more authors.
Virus Research | Year: 2014

The persistence of porcine rubulavirus (PorPV-LPMV) in five pigs that had survived an outbreak of a natural infection was determined. After the resolution of the outbreak, each animal was housed in an isolation pen together with one sentinel pig. Approximately every 2 months thereafter one group of animals was euthanized and tissue samples taken for virological and serological analysis. Infectious virus was not isolated from any samples; antibodies to PorPV-LPMV were detected in convalescent pigs by virus neutralisation test and blocking ELISA but not in sentinel pigs. PorPV-LPMV mRNA of the nucleoprotein (NP) and phosphoprotein (P) genes was detected by a nested polymerase chain reaction (nPCR) in samples of trigeminal and optic nerves, cervical spinal cord, tonsils, salivary gland, lung and pancreas from convalescent pigs. mRNA was also detected in the midbrain, corpus callosum, or olfactory bulb in four out of five pigs by nRT-PCR, this result was confirmed by the sequencing of a 260. bp PCR product of P gene region. The highest average viral copies/μg of total RNA occurred in the olfactory bulb and pancreas tissues of convalescent pigs and midbrain, tonsil and pancreas of sentinel pigs housed with the convalescent pigs. Satellitosis and gliosis of the midbrain, olfactory bulb, corpus callosum, medulla oblongata or choroid plexus were microscopically observed in four convalescent pigs. The control pig remained negative in all tests. The results indicate that PorPV-LPMV mRNA persists and induces a durable humoral immune response in pigs that have recovered from a natural infection. After a possible reactivation of the virus, it was transmitted to sentinel pigs in contact with the convalescent pigs. © 2014 Elsevier B.V.


PubMed | National Research Center Disciplinaria en Microbiologia Animal, National Autonomous University of Mexico, Research Center Biomedica Of Oriente and Swedish University of Agricultural Sciences
Type: Journal Article | Journal: Virus genes | Year: 2016

Since the report of the initial outbreak of Porcine rubulavirus (PorPV) infection in pigs, only one full-length genome from 1984 (PorPV-LPMV/1984) has been characterised. To investigate the overall genetic variation, full-length gene nucleotide sequences of current PorPV isolates were obtained from different clinical cases of infected swine. Genome organisation and sequence analysis of the encoded proteins (NP, P, F, M, HN and L) revealed high sequence conservation of the NP protein and the expression of the P and V proteins in all PorPV isolates. The V protein of one isolate displayed a mutation that has been implicated to antagonise the antiviral immune responses of the host. The M protein indicated a variation in a short region that could affect the electrostatic charge and the interaction with the membrane. One PorPV isolate recovered from the lungs showed a mutation at the cleavage site (HRKKR) of the F protein that could represent an important factor to determine the tissue tropism and pathogenicity of this virus. The HN protein showed high sequence identity through the years (up to 2013). Additionally, a number of sequence motifs of very high amino acid conservation among the PorPV isolates important for polymerase activity of the L protein have been identified. In summary, genetic comparisons and phylogenetic analyses indicated that three different genetic variants of PorPV are currently spreading within the swine population, and a new generation of circulating virus with different characteristics has begun to emerge.


Escalera-Zamudio M.,Leibniz Institute for Zoo and Wildlife Research | Lisandra Zepeda Mendoza M.,Copenhagen University | Heeger F.,Berlin Center for Genomics in Biodiversity Research | Loza-Rubio E.,National Research Center Disciplinaria en Microbiologia Animal | And 7 more authors.
Journal of Virology | Year: 2015

The Desmodus rotundus endogenous betaretrovirus (DrERV) is fixed in the vampire bat D. rotundus population and in other phyllostomid bats but is not present in all species from this family. DrERV is not phylogenetically related to Old World bat betaretroviruses but to betaretroviruses from rodents and New World primates, suggesting recent cross-species transmission. A recent integration age estimation of the provirus in some taxa indicates that an exogenous counterpart might have been in recent circulation. © 2015, American Society for Microbiology.


Escalera-Zamudio M.,Leibniz Institute for Zoo and Wildlife Research | Zepeda-Mendoza M.L.,Copenhagen University | Loza-Rubio E.,National Research Center Disciplinaria en Microbiologia Animal | Rojas-Anaya E.,National Research Center Disciplinaria en Microbiologia Animal | And 4 more authors.
Molecular Ecology | Year: 2015

We characterized the nucleic acid-sensing Toll-like receptors (TLR) of a New World bat species, the common vampire bat (Desmodus rotundus), and through a comparative molecular evolutionary approach searched for general adaptation patterns among the nucleic acid-sensing TLRs of eight different bats species belonging to three families (Pteropodidae, Vespertilionidae and Phyllostomidae). We found that the bat TLRs are evolving slowly and mostly under purifying selection and that the divergence pattern of such receptors is overall congruent with the species tree, consistent with the evolution of many other mammalian nuclear genes. However, the chiropteran TLRs exhibited unique mutations fixed in ligand-binding sites, some of which involved nonconservative amino acid changes and/or targets of positive selection. Such changes could potentially modify protein function and ligand-binding properties, as some changes were predicted to alter nucleic acid binding motifs in TLR 9. Moreover, evidence for episodic diversifying selection acting specifically upon the bat lineage and sublineages was detected. Thus, the long-term adaptation of chiropterans to a wide variety of environments and ecological niches with different pathogen profiles is likely to have shaped the evolution of the bat TLRs in an order-specific manner. The observed evolutionary patterns provide evidence for potential functional differences between bat and other mammalian TLRs in terms of resistance to specific pathogens or recognition of nucleic acids in general. © 2015 John Wiley & Sons Ltd.


Bernal G.R.,National Autonomous University of Mexico | Sagahon A.V.L.,National Autonomous University of Mexico | Tovar C.G.G.,National Autonomous University of Mexico | Aparicio E.D.,National Research Center Disciplinaria en Microbiologia Animal | Gutierrez V.R.T.,National Research Center Disciplinaria en Microbiologia Animal
Revista Mexicana De Ciencias Pecuarias | Year: 2014

The virulence of 12 Listeria monocytogenes strains from goat and goat by-products was evaluated. Variable virulence levels were found when compared the relative virulence (mortality, %) and the ability to grow infect the spleen of BALB/c mice inoculated intravenously or intragastrically. Two strains from feed produced 100 % mortality, by either inoculation route and seven strains, including those from feed, infected the spleen with both inoculation routes. A strain inoculated intravenously without causing death in the mice produced spleen counts similar to those obtained in dead mice, this could indicate that a high count of the microorganism in the spleen not necessarily ensures its virulence. We also tested the capacity to infect J774 macrophages and Caco-2 epithelial cells, finding higher invasion and intracellular growth in Caco-2 with respect the macrophages and showing quadratic growth kinetics in both cell lines. Only the two feed strains did not show a significant difference from the control strain (P>0.05) regarding kinetic parameters and invasion percentage in both cell lines. Although all isolates were able to invade Caco-2 cells, not all isolates were able to invade the spleen after the intragastrically inoculation. Two goats feed strains were the most virulent, representing a risk for animal and human health.


PubMed | National Research Center Disciplinaria en Microbiologia Animal, National Autonomous University of Mexico, Copenhagen University, Leibniz Institute for Zoo and Wildlife Research and University of Veracruz
Type: Journal Article | Journal: Molecular ecology | Year: 2015

We characterized the nucleic acid-sensing Toll-like receptors (TLR) of a New World bat species, the common vampire bat (Desmodus rotundus), and through a comparative molecular evolutionary approach searched for general adaptation patterns among the nucleic acid-sensing TLRs of eight different bats species belonging to three families (Pteropodidae, Vespertilionidae and Phyllostomidae). We found that the bat TLRs are evolving slowly and mostly under purifying selection and that the divergence pattern of such receptors is overall congruent with the species tree, consistent with the evolution of many other mammalian nuclear genes. However, the chiropteran TLRs exhibited unique mutations fixed in ligand-binding sites, some of which involved nonconservative amino acid changes and/or targets of positive selection. Such changes could potentially modify protein function and ligand-binding properties, as some changes were predicted to alter nucleic acid binding motifs in TLR 9. Moreover, evidence for episodic diversifying selection acting specifically upon the bat lineage and sublineages was detected. Thus, the long-term adaptation of chiropterans to a wide variety of environments and ecological niches with different pathogen profiles is likely to have shaped the evolution of the bat TLRs in an order-specific manner. The observed evolutionary patterns provide evidence for potential functional differences between bat and other mammalian TLRs in terms of resistance to specific pathogens or recognition of nucleic acids in general.


PubMed | Copenhagen University, National Research Center Disciplinaria en Microbiologia Animal, National Autonomous University of Mexico, Leibniz Institute for Zoo and Wildlife Research and 2 more.
Type: Journal Article | Journal: Journal of virology | Year: 2015

The Desmodus rotundus endogenous betaretrovirus (DrERV) is fixed in the vampire bat D. rotundus population and in other phyllostomid bats but is not present in all species from this family. DrERV is not phylogenetically related to Old World bat betaretroviruses but to betaretroviruses from rodents and New World primates, suggesting recent cross-species transmission. A recent integration age estimation of the provirus in some taxa indicates that an exogenous counterpart might have been in recent circulation.

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