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Isolation and characterization of adenoviruses persistently shed from the gastrointestinal tract of non-human primates.

Roy S, Vandenberghe LH, Kryazhimskiy S, Grant R, Calcedo R, Yuan X, Keough M, Sandhu A, Wang Q, Medina-Jaszek CA, Plotkin JB, Wilson JM - PLoS Pathog. (2009)

Bottom Line: Adenoviruses are important human pathogens that have been developed as vectors for gene therapies and genetic vaccines.Previous studies indicated that human infections with adenoviruses are self-limiting in immunocompetent hosts with evidence of some persistence in adenoid tissue.Furthermore, the presence of persistent and/or latent adenovirus infections in the gut should be considered in the design and interpretation of human and non-human primate studies with adenovirus vectors.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, USA.

ABSTRACT
Adenoviruses are important human pathogens that have been developed as vectors for gene therapies and genetic vaccines. Previous studies indicated that human infections with adenoviruses are self-limiting in immunocompetent hosts with evidence of some persistence in adenoid tissue. We sought to better understand the natural history of adenovirus infections in various non-human primates and discovered that healthy populations of great apes (chimpanzees, bonobos, gorillas, and orangutans) and macaques shed substantial quantities of infectious adenoviruses in stool. Shedding in stools from asymptomatic humans was found to be much less frequent, comparable to frequencies reported before. We purified and fully sequenced 30 novel adenoviruses from apes and 3 novel adenoviruses from macaques. Analyses of the new ape adenovirus sequences (as well as the 4 chimpanzee adenovirus sequences we have previously reported) together with 22 complete adenovirus genomes available from GenBank revealed that (a) the ape adenoviruses could clearly be classified into species corresponding to human adenovirus species B, C, and E, (b) there was evidence for intraspecies recombination between adenoviruses, and (c) the high degree of phylogenetic relatedness of adenoviruses across their various primate hosts provided evidence for cross species transmission events to have occurred in the natural history of B and E viruses. The high degree of asymptomatic shedding of live adenovirus in non-human primates and evidence for zoonotic transmissions warrants caution for primate handling and housing. Furthermore, the presence of persistent and/or latent adenovirus infections in the gut should be considered in the design and interpretation of human and non-human primate studies with adenovirus vectors.

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Phylogenetic tree of adenoviruses that infect primates.The tree was reconstructed from an alignment of the polymerase gene, using maximum likelihood under the HKY85 model of substitutions, as described in Materials and Methods. The names of simian isolates include the serotype nomenclature, the animal species of isolation (Hu: human, Ch: chimpanzee, Bo: bonobo, Go: gorilla, Cy: cynomolgus macaque, Rh: rhesus macaque), and the source of adenoviral isolation (ATCC: American Tissue Type Collection, JX: Jacksonville zoo, NI: New Iberia Research Center, MD: MD Anderson, SD: San Diego zoo, At: Atlanta zoo, LR: Little Rock zoo, BF: Buffalo zoo). Isolates that have a closely related hexon structure are referred to as variants of the same serotype (“.1” or “.2”). Names of novel sequences obtained in this study are shown in italics. Colors indicate the six species of viruses that infect higher primates (A, B, C, D, E, and F); grey is used to indicate viruses isolated from monkeys. The inset (upper left) shows the same tree with the inclusion of a tree-shrew isolate as an outgroup, after collapsing poorly supported bifurcations. Bootstrap values less than 80% or close to terminal leaves are suppressed.
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ppat-1000503-g001: Phylogenetic tree of adenoviruses that infect primates.The tree was reconstructed from an alignment of the polymerase gene, using maximum likelihood under the HKY85 model of substitutions, as described in Materials and Methods. The names of simian isolates include the serotype nomenclature, the animal species of isolation (Hu: human, Ch: chimpanzee, Bo: bonobo, Go: gorilla, Cy: cynomolgus macaque, Rh: rhesus macaque), and the source of adenoviral isolation (ATCC: American Tissue Type Collection, JX: Jacksonville zoo, NI: New Iberia Research Center, MD: MD Anderson, SD: San Diego zoo, At: Atlanta zoo, LR: Little Rock zoo, BF: Buffalo zoo). Isolates that have a closely related hexon structure are referred to as variants of the same serotype (“.1” or “.2”). Names of novel sequences obtained in this study are shown in italics. Colors indicate the six species of viruses that infect higher primates (A, B, C, D, E, and F); grey is used to indicate viruses isolated from monkeys. The inset (upper left) shows the same tree with the inclusion of a tree-shrew isolate as an outgroup, after collapsing poorly supported bifurcations. Bootstrap values less than 80% or close to terminal leaves are suppressed.

Mentions: Chimpanzee stool samples collected from natural habitats in Cameroon and the Democratic Republic of Congo (DRC) were made available to us for analysis. DNA isolated from these samples has been used for simian foamy virus studies [20]. These samples underwent a sensitive nested PCR that was designed to directly detect adenovirus genomes using oligonucleotides complementary to a conserved region of the DNA polymerase gene (pol). Adenovirus DNA could be detected in 40% of the stool samples from chimpanzees that exist in the wild (Table 1). Cloning and sequencing of these PCR products identified twenty-four that were species E and three that were species B (data not shown). In this conserved region, species E viruses closely clustered with SAdV-25.1, 26 and 39 whereas species B isolates where mostly similar to SAdV-35.1 and SAdV-35.2 (Figure 1). We also obtained stool from mountain gorillas in the Virunga Massif mountain range in Rwanda and detected adenovirus in 3/6 samples by pol PCR. Stool extracts were also filtered and evaluated for their ability to induce cytopathic effects (CPE) on the human epithelial A549 cell line. Every attempt to grow out live adenovirus on A549 cells was unsuccessful. We were unable to determine whether this failure was due to the technical limitations of sample collection or preservation, the sensitivity of the cellular detection assay or the absence of functional viral particles. Analysis of samples acquired under more controlled conditions in the USA permitted us to address these important questions.


Isolation and characterization of adenoviruses persistently shed from the gastrointestinal tract of non-human primates.

Roy S, Vandenberghe LH, Kryazhimskiy S, Grant R, Calcedo R, Yuan X, Keough M, Sandhu A, Wang Q, Medina-Jaszek CA, Plotkin JB, Wilson JM - PLoS Pathog. (2009)

Phylogenetic tree of adenoviruses that infect primates.The tree was reconstructed from an alignment of the polymerase gene, using maximum likelihood under the HKY85 model of substitutions, as described in Materials and Methods. The names of simian isolates include the serotype nomenclature, the animal species of isolation (Hu: human, Ch: chimpanzee, Bo: bonobo, Go: gorilla, Cy: cynomolgus macaque, Rh: rhesus macaque), and the source of adenoviral isolation (ATCC: American Tissue Type Collection, JX: Jacksonville zoo, NI: New Iberia Research Center, MD: MD Anderson, SD: San Diego zoo, At: Atlanta zoo, LR: Little Rock zoo, BF: Buffalo zoo). Isolates that have a closely related hexon structure are referred to as variants of the same serotype (“.1” or “.2”). Names of novel sequences obtained in this study are shown in italics. Colors indicate the six species of viruses that infect higher primates (A, B, C, D, E, and F); grey is used to indicate viruses isolated from monkeys. The inset (upper left) shows the same tree with the inclusion of a tree-shrew isolate as an outgroup, after collapsing poorly supported bifurcations. Bootstrap values less than 80% or close to terminal leaves are suppressed.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2698151&req=5

ppat-1000503-g001: Phylogenetic tree of adenoviruses that infect primates.The tree was reconstructed from an alignment of the polymerase gene, using maximum likelihood under the HKY85 model of substitutions, as described in Materials and Methods. The names of simian isolates include the serotype nomenclature, the animal species of isolation (Hu: human, Ch: chimpanzee, Bo: bonobo, Go: gorilla, Cy: cynomolgus macaque, Rh: rhesus macaque), and the source of adenoviral isolation (ATCC: American Tissue Type Collection, JX: Jacksonville zoo, NI: New Iberia Research Center, MD: MD Anderson, SD: San Diego zoo, At: Atlanta zoo, LR: Little Rock zoo, BF: Buffalo zoo). Isolates that have a closely related hexon structure are referred to as variants of the same serotype (“.1” or “.2”). Names of novel sequences obtained in this study are shown in italics. Colors indicate the six species of viruses that infect higher primates (A, B, C, D, E, and F); grey is used to indicate viruses isolated from monkeys. The inset (upper left) shows the same tree with the inclusion of a tree-shrew isolate as an outgroup, after collapsing poorly supported bifurcations. Bootstrap values less than 80% or close to terminal leaves are suppressed.
Mentions: Chimpanzee stool samples collected from natural habitats in Cameroon and the Democratic Republic of Congo (DRC) were made available to us for analysis. DNA isolated from these samples has been used for simian foamy virus studies [20]. These samples underwent a sensitive nested PCR that was designed to directly detect adenovirus genomes using oligonucleotides complementary to a conserved region of the DNA polymerase gene (pol). Adenovirus DNA could be detected in 40% of the stool samples from chimpanzees that exist in the wild (Table 1). Cloning and sequencing of these PCR products identified twenty-four that were species E and three that were species B (data not shown). In this conserved region, species E viruses closely clustered with SAdV-25.1, 26 and 39 whereas species B isolates where mostly similar to SAdV-35.1 and SAdV-35.2 (Figure 1). We also obtained stool from mountain gorillas in the Virunga Massif mountain range in Rwanda and detected adenovirus in 3/6 samples by pol PCR. Stool extracts were also filtered and evaluated for their ability to induce cytopathic effects (CPE) on the human epithelial A549 cell line. Every attempt to grow out live adenovirus on A549 cells was unsuccessful. We were unable to determine whether this failure was due to the technical limitations of sample collection or preservation, the sensitivity of the cellular detection assay or the absence of functional viral particles. Analysis of samples acquired under more controlled conditions in the USA permitted us to address these important questions.

Bottom Line: Adenoviruses are important human pathogens that have been developed as vectors for gene therapies and genetic vaccines.Previous studies indicated that human infections with adenoviruses are self-limiting in immunocompetent hosts with evidence of some persistence in adenoid tissue.Furthermore, the presence of persistent and/or latent adenovirus infections in the gut should be considered in the design and interpretation of human and non-human primate studies with adenovirus vectors.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, USA.

ABSTRACT
Adenoviruses are important human pathogens that have been developed as vectors for gene therapies and genetic vaccines. Previous studies indicated that human infections with adenoviruses are self-limiting in immunocompetent hosts with evidence of some persistence in adenoid tissue. We sought to better understand the natural history of adenovirus infections in various non-human primates and discovered that healthy populations of great apes (chimpanzees, bonobos, gorillas, and orangutans) and macaques shed substantial quantities of infectious adenoviruses in stool. Shedding in stools from asymptomatic humans was found to be much less frequent, comparable to frequencies reported before. We purified and fully sequenced 30 novel adenoviruses from apes and 3 novel adenoviruses from macaques. Analyses of the new ape adenovirus sequences (as well as the 4 chimpanzee adenovirus sequences we have previously reported) together with 22 complete adenovirus genomes available from GenBank revealed that (a) the ape adenoviruses could clearly be classified into species corresponding to human adenovirus species B, C, and E, (b) there was evidence for intraspecies recombination between adenoviruses, and (c) the high degree of phylogenetic relatedness of adenoviruses across their various primate hosts provided evidence for cross species transmission events to have occurred in the natural history of B and E viruses. The high degree of asymptomatic shedding of live adenovirus in non-human primates and evidence for zoonotic transmissions warrants caution for primate handling and housing. Furthermore, the presence of persistent and/or latent adenovirus infections in the gut should be considered in the design and interpretation of human and non-human primate studies with adenovirus vectors.

Show MeSH
Related in: MedlinePlus