Simian Foamy Viruses in Central and South America: A New World of Discovery.

Foamy viruses (FVs) are the only exogenous retrovirus to date known to infect neotropical primates (NPs). In the last decade, an increasing number of strains have been completely or partially sequenced, and molecular evolution analyses have identified an ancient co-speciation with their hosts. In this review, the improvement of diagnostic techniques that allowed the determination of a more accurate prevalence of simian FVs (SFVs) in captive and free-living NPs is discussed. Determination of DNA viral load in American primates indicates that oral tissues are the viral replicative site and that buccal swab collection can be an alternative to diagnose SFV infection in NPs. Finally, the transmission potential of NP SFVs to primate workers in zoos and primate centers of the Americas is examined.

Simian Foamy Virus Co-Infections.

Foamy viruses (FVs), also known as spumaretroviruses, are complex retroviruses that are seemingly nonpathogenic in natural hosts. In natural hosts, which include felines, bovines, and nonhuman primates (NHPs), a large percentage of adults are infected with FVs. For this reason, the effect of FVs on infections with other viruses (co-infections) cannot be easily studied in natural populations. Most of what is known about interactions between FVs and other viruses is based on studies of NHPs in artificial settings such as research facilities. In these settings, there is some indication that FVs can exacerbate infections with lentiviruses such as simian immunodeficiency virus (SIV). Nonhuman primate (NHP) simian FVs (SFVs) have been shown to infect people without any apparent pathogenicity. Humans zoonotically infected with simian foamy virus (SFV) are often co-infected with other viruses. Thus, it is important to know whether SFV co-infections affect human disease.

Origin, evolution and innate immune control of simian foamy viruses in humans.

Most viral pathogens that have emerged in humans have originated from various animal species. Emergence is a multistep process involving an initial spill-over of the infectious agent into single individuals and its subsequent dissemination into the human population. Similar to simian immunodeficiency viruses and simian T lymphotropic viruses, simian foamy viruses (SFV) are retroviruses that are widespread among non-human primates and can be transmitted to humans, giving rise to a persistent infection, which seems to be controlled in the case of SFV. In this review, we present current data on the discovery, cross-species transmission, and molecular evolution of SFV in human populations initially infected and thus at risk for zoonotic emergence.

In vivo cellular tropism of gorilla simian foamy virus in blood of infected humans.

UNLABELLED: Simian foamy viruses (SFV) are retroviruses that are widespread among nonhuman primates. SFV can be transmitted to humans, giving rise to a persistent infection. Only a few data are available concerning the distribution of SFV in human blood cells. Here we purified blood mononuclear cell subsets from 11 individuals infected with a Gorilla gorilla SFV strain and quantified SFV DNA levels by quantitative PCR. SFV DNA was detected in the majority of the CD8(+), CD4(+), and CD19(+) lymphocyte samples and rarely in CD14(+) monocyte and CD56(+) NK lymphocyte samples. The median (interquartile range [IQR]) SFV DNA counts were 16.0 (11.0 to 49.8), 11.3 (5.9 to 28.3), and 17.2 (2.0 to 25.2) copies/10(5) cells in CD8(+) T lymphocytes, CD4(+) T lymphocytes, and CD19(+) B lymphocytes, respectively. In the CD4 compartment, SFV DNA was detected in both memory and naive CD4(+) T lymphocytes. SFV DNA levels in CD4(+) T cells were positively correlated with the duration of the infection. Our study shows with a quantitative method that CD8(+), CD4(+), and B lymphocytes are major cellular targets of SFV in the blood of infected humans. IMPORTANCE: Investigation of SFV infections in humans is important due to the origin of human immunodeficiency viruses (HIV) and human T cell lymphotropic viruses (HTLV) from cross-species transmission of their simian counterparts to humans. Surprisingly little is known about many aspects of the biology of SFV in infected humans, including quantitative data concerning the cellular targets of SFV in vivo. Here we show that the distribution of SFV DNA among the different leukocyte populations is not homogeneous and that viral load in CD4(+) T lymphocytes is correlated with the duration of infection. These new data will help in understanding the biology of retroviral infections in humans and can be useful in the growing field of SFV-based gene therapy.

Noncanonical microRNA (miRNA) biogenesis gives rise to retroviral mimics of lymphoproliferative and immunosuppressive host miRNAs.

MicroRNAs (miRNAs) play regulatory roles in diverse processes in both eukaryotic hosts and their viruses, yet fundamental questions remain about which viruses code for miRNAs and the functions that they serve. Simian foamy viruses (SFVs) of Old World monkeys and apes can zoonotically infect humans and, by ill-defined mechanisms, take up lifelong infections in their hosts. Here, we report that SFVs encode multiple miRNAs via a noncanonical mode of biogenesis. The primary SFV miRNA transcripts (pri-miRNAs) are transcribed by RNA polymerase III (RNAP III) and take multiple forms, including some that are cleaved by Drosha. However, these miRNAs are generated in a context-dependent fashion, as longer RNAP II transcripts spanning this region are resistant to Drosha cleavage. This suggests that the virus may avoid any fitness penalty that could be associated with viral genome/transcript cleavage. Two SFV miRNAs share sequence similarity and functionality with notable host miRNAs, the lymphoproliferative miRNA miR-155 and the innate immunity suppressor miR-132. These results have important implications regarding foamy virus biology, viral miRNAs, and the development of retroviral-based vectors. IMPORTANCE Fundamental questions remain about which viruses encode miRNAs and their associated functions. Currently, few natural viruses with RNA genomes have been reported to encode miRNAs. Simian foamy viruses are retroviruses that are prevalent in nonhuman host populations, and some can zoonotically infect humans who hunt primates or work as animal caretakers. We identify a cluster of miRNAs encoded by SFV. Characterization of these miRNAs reveals evolutionarily conserved, unconventional mechanisms to generate small RNAs. Several SFV miRNAs share sequence similarity and functionality with host miRNAs, including the oncogenic miRNA miR-155 and innate immunity suppressor miR-132. Strikingly, unrelated herpesviruses also tap into one or both of these same regulatory pathways, implying relevance to a broad range of viruses. These findings provide new insights with respect to foamy virus biology and vectorology.

A novel Bayesian method for detection of APOBEC3-mediated hypermutation and its application to zoonotic transmission of simian foamy viruses.

Simian Foamy Virus (SFV) can be transmitted from non-human primates (NHP) to humans. However, there are no documented cases of human to human transmission, and significant differences exist between infection in NHP and human hosts. The mechanism for these between-host differences is not completely understood. In this paper we develop a new Bayesian approach to the detection of APOBEC3-mediated hypermutation, and use it to compare SFV sequences from human and NHP hosts living in close proximity in Bangladesh. We find that human APOBEC3G can induce genetic changes that may prevent SFV replication in infected humans in vivo.

A novel small animal model to study the replication of simian foamy virus in vivo.

Preclinical evaluation in a small animal model would help the development of gene therapies and vaccines based on foamy virus vectors. The establishment of persistent, non-pathogenic infection with the prototype foamy virus in mice and rabbits has been described previously. To extend this spectrum of available animal models, hamsters were inoculated with infectious cell supernatant or bioballistically with a foamy virus plasmid. In addition, a novel foamy virus from a rhesus macaque was isolated and characterised genetically. Hamsters and mice were infected with this new SFVmac isolate to evaluate whether hamsters are also susceptible to infection. Both hamsters and mice developed humoral responses to either virus subtype. Virus integration and replication in different animal tissues were analysed by PCR and co-cultivation. The results strongly indicate establishment of a persistent infection in hamsters. These studies provide a further small animal model for studying FV-based vectors in addition to the established models.

New World simian foamy virus infections in vivo and in vitro.

Foamy viruses (FV) are complex retroviruses that naturally infect all nonhuman primates (NHP) studied to date. Zoonotic transmission of Old World NHP simian foamy viruses (SFV) has been documented, leading to nonpathogenic persistent infections. To date, there have been no reports concerning zoonotic transmission of New World monkey (NWM) SFV to humans and resulting infection. In this study, we developed a Western blot assay to detect antibodies to NWM SFV, a nested PCR assay to detect NWM SFV DNA, and a ß-galactosidase-containing indicator cell line to assay replication of NWM SFV. Using these tools, we analyzed the plasma and blood of 116 primatologists, of whom 69 had reported exposures to NWM. While 8 of the primatologists tested were seropositive for SFV from a NWM, the spider monkey, none had detectable levels of viral DNA in their blood. We found that SFV isolated from three different species of NWM replicated in some, but not all, human cell lines. From our data, we conclude that while humans exposed to NWM SFV produce antibodies, there is no evidence for long-term viral persistence.

Viral latency in blood and saliva of simian foamy virus-infected humans.

Simian foamy viruses (SFV) are widespread retroviruses among non-human primates (NHP). SFV actively replicate in the oral cavity and can be transmitted to humans through NHP bites, giving rise to a persistent infection. We aimed at studying the natural history of SFV infection in human. We have analyzed viral load and gene expression in 14 hunters from Cameroon previously shown to be infected with a gorilla SFV strain. Viral DNA could be detected by quantitative polymerase chain reaction (q-PCR) targeting the pol-in region, in most samples of peripheral blood mononuclear cells (PBMCs) (7.1 ± 6.0 SFV DNA copies/105 PBMCs) and saliva (2.4 ± 4.3 SFV DNA copies/105 cells) derived from the hunters. However, quantitative real-time reverse-transcription polymerase chain reaction (RT)-qPCR revealed the absence of SFV viral gene expression in both PBMCs and saliva, suggesting that SFV was latent in the human samples. Our study demonstrates that a latent infection can occur in humans and persist for years, both in PBMCs and saliva. Such a scenario may contribute to the putative lack of secondary human-to-human transmissions of SFV.

Simian foamy virus infection of rhesus macaques in Bangladesh: relationship of latent proviruses and transcriptionally active viruses.

Simian foamy viruses (SFV) are complex retroviruses that are ubiquitous in nonhuman primates (NHP) and are zoonotically transmitted to humans, presumably through NHP saliva, by licking, biting, and other behaviors. We have studied SFV in free-ranging rhesus macaques in Bangladesh. It has been previously shown that SFV in immunocompetent animals replicates to detectable levels only in superficial epithelial cells of the oral mucosa, although latent proviruses are found in most, if not all, tissues. In this study, we compare DNA sequences from latent SFV proviruses found in blood cells of 30 Bangladesh rhesus macaques to RNA sequences of transcriptionally active SFV from buccal swabs obtained from the same animals. Viral strains, defined by differences in SFV gag sequences, from buccal mucosal specimens overlapped with those from blood samples in 90% of animals. Thus, latent proviruses in peripheral blood mononuclear cells (PBMC) are, to a great extent, representative of viruses likely to be transmitted to other hosts. The level of SFV RNA in buccal swabs varied greatly between macaques, with increasing amounts of viral RNA in older animals. Evidence of APOBEC3-induced mutations was found in gag sequences derived from the blood and oral mucosa.

Mother-offspring transmission and age-dependent accumulation of simian foamy virus in wild chimpanzees.

Simian foamy viruses (SFVs) are thought to infect virtually any adult nonhuman primate (NHP). While many data have accumulated about patterns of codivergence with their hosts and cross-species transmission events, little is known about the modalities of SFV transmission within NHP species, especially in the wild. Here we provide a detailed investigation of the dynamics of SFV circulation in a wild community of Western chimpanzees (Pan troglodytes verus). We demonstrate that mother-offspring (vertical) SFV transmission is common and hypothesize that it accounts for a number of primary infections. We also show that multiple infections with several chimpanzee-specific SFV strains (i.e., superinfection) commonly happen in adult chimpanzees, which might point to adult-specific aggressive behaviors as a lifelong source of SFV infection. Our data give evidence for complex SFV dynamics in wild chimpanzees, even at a single community scale, and show that linking wild NHP social interactions and their microorganisms' dynamics is feasible.

[Mechanisms of viral emergence and interspecies transmission: the exemple of simian foamy viruses in Central Africa]. / Mécanismes d'émergence virale et transmission interespèces: l'exemple des rétrovirus Foamy simiens chez l'Homme en Afrique Centrale.

A large proportion of viral pathogens that have emerged during the last decades in humans are considered to have originated from various animal species. This is well exemplified by several recent epidemics such as those of Nipah, Severe Acute Respiratory Syndrome, Avian flu, Ebola, Monkeypox, and Hantaviruses. After the initial interspecies transmission per se, the viruses can disseminate into the human population through various and distinct mechanisms. Some of them are well characterized and understood, thus allowing a certain level of risk control and prevention. Surprisingly and in contrast, the initial steps that lead to the emergence of several viruses, and of their associated diseases, remain still poorly understood. Epidemiological field studies conducted in certain specific high-risk populations are thus necessary to obtain new insights into the early events of this emergence process. Human infections by simian viruses represent increasing public health concerns. Indeed, by virtue of their genetic andphysiological similarities, non-human primates (NHPs) are considered to be likely the sources of viruses that can infect humans and thus may pose a significant threat to human population. This is well illustrated by retroviruses, which have the ability to cross species, adapt to a new host and sometimes spread within these new species. Sequence comparison and phylogenetic studies have thus clearly showed that the emergence of human immunodeficiency virus type 1 (HIV-1) and HIV-2 in humans have resulted from several independent interspecies transmissions of different SIV types from Chimpanzees and African monkeys (including sooty mangabeys), respectively, probably during the first part of the last century. The situation for Human T cell Lymphotropic virus type 1 (HTLV-1) is, for certain aspects, quite comparable. Indeed, the origin of most HTLV-1 subtypes appears to be linked to interspecies transmission between STLV-1-infected monkeys and humans, followed by variable periods of evolution in the human host. In this review, after an introduction on emerging viruses, we will briefly present the results of a large epidemiological study performed in groups of Bantus and Pygmies living in villages and settlements located in the rain forest of the South region of Cameroon. These populations are living nearby the habitats of several monkeys and apes, often naturally infected by different retroviruses including SIV, STLV and simianfoamy virus. Most of the persons included in this study were hunters of such NHPs, thus at high risk of contact with infected body fluids (blood, saliva,...) during hunting activities. After reviewing the current available data on the discovery, cross-species transmission from monkeys and apes to humans of the simian foamy retroviruses, we will report the results of our study. Such infection is a unique natural model to study the different mechanisms of restriction of retroviral emergence in Humans.

Cross-species transmission of simian foamy virus to humans in rural Gabon, Central Africa.

In order to characterize simian foamy retroviruses (SFVs) in wild-born nonhuman primates (NHPs) in Gabon and to investigate cross-species transmission to humans, we obtained 497 NHP samples, composed of 286 blood and 211 tissue (bush meat) samples. Anti-SFV antibodies were found in 31 of 286 plasma samples (10.5%). The integrase gene sequence was found in 38/497 samples, including both blood and tissue samples, with novel SFVs in several Cercopithecus species. Of the 78 humans, mostly hunters, who had been bitten or scratched by NHPs, 19 were SFV seropositive, with 15 cases confirmed by PCR. All but one were infected with ape SFV. We thus found novel SFV strains in NHPs in Gabon and high cross-species transmission of SFVs from gorilla bites.

A nuclear export signal within the structural Gag protein is required for prototype foamy virus replication.

BACKGROUND: The Gag polyproteins play distinct roles during the replication cycle of retroviruses, hijacking many cellular machineries to fulfill them. In the case of the prototype foamy virus (PFV), Gag structural proteins undergo transient nuclear trafficking after their synthesis, returning back to the cytoplasm for capsid assembly and virus egress. The functional role of this nuclear stage as well as the molecular mechanism(s) responsible for Gag nuclear export are not understood. RESULTS: We have identified a leptomycin B (LMB)-sensitive nuclear export sequence (NES) within the N-terminus of PFV Gag that is absolutely required for the completion of late stages of virus replication. Point mutations of conserved residues within this motif lead to nuclear redistribution of Gag, preventing subsequent virus egress. We have shown that a NES-defective PFV Gag acts as a dominant negative mutant by sequestrating its wild-type counterpart in the nucleus. Trans-complementation experiments with the heterologous NES of HIV-1 Rev allow the cytoplasmic redistribution of FV Gag, but fail to restore infectivity. CONCLUSIONS: PFV Gag-Gag interactions are finely tuned in the cytoplasm to regulate their functions, capsid assembly, and virus release. In the nucleus, we have shown Gag-Gag interactions which could be involved in the nuclear export of Gag and viral RNA. We propose that nuclear export of unspliced and partially spliced PFV RNAs relies on two complementary mechanisms, which take place successively during the replication cycle.

Two distinct variants of simian foamy virus in naturally infected mandrills (Mandrillus sphinx) and cross-species transmission to humans.

BACKGROUND: Each of the pathogenic human retroviruses (HIV-1/2 and HTLV-1) has a nonhuman primate counterpart, and the presence of these retroviruses in humans results from interspecies transmission. The passage of another simian retrovirus, simian foamy virus (SFV), from apes or monkeys to humans has been reported. Mandrillus sphinx, a monkey species living in central Africa, is naturally infected with SFV. We evaluated the natural history of the virus in a free-ranging colony of mandrills and investigated possible transmission of mandrill SFV to humans. RESULTS: We studied 84 semi-free-ranging captive mandrills at the Primate Centre of the Centre International de Recherches Médicales de Franceville (Gabon) and 15 wild mandrills caught in various areas of the country. The presence of SFV was also evaluated in 20 people who worked closely with mandrills and other nonhuman primates. SFV infection was determined by specific serological (Western blot) and molecular (nested PCR of the integrase region in the polymerase gene) assays. Seropositivity for SFV was found in 70/84 (83%) captive and 9/15 (60%) wild-caught mandrills and in 2/20 (10%) humans. The 425-bp SFV integrase fragment was detected in peripheral blood DNA from 53 captive and 8 wild-caught mandrills and in two personnel. Sequence and phylogenetic studies demonstrated the presence of two distinct strains of mandrill SFV, one clade including SFVs from mandrills living in the northern part of Gabon and the second consisting of SFV from animals living in the south. One man who had been bitten 10 years earlier by a mandrill and another bitten 22 years earlier by a macaque were found to be SFV infected, both at the Primate Centre. The second man had a sequence close to SFVmac sequences. Comparative sequence analysis of the virus from the first man and from the mandrill showed nearly identical sequences, indicating genetic stability of SFV over time. CONCLUSION: Our results show a high prevalence of SFV infection in a semi-free-ranging colony of mandrills, with the presence of two different strains. We also showed transmission of SFV from a mandrill and a macaque to humans.

[Improving infection efficiency of adult cynomolgus monkey mesenchymal stem cells with lentiviral vectors].

Mesenchymal stem cells (MSCs) have received considerable attention for various therapeutic approaches in recent years. MSCs are also easy to genetically modify to express therapeutic genes by using lentiviral vectors. Because of the similarities in genetics, physiology and metabolism between non-human primates (NHPs) and humans, NHPs models are invaluable for researching human disorders and for developing therapeutic strategies. Therefore, MSCs derived from NHPs could be a powerful tool for cell therapy and genetic engineering. Studies from captive and free-ranging adult NHPs show that up to 100% were infected with simian foamy virus (SFV). In this study, we found that all cultured MSCs derived from adult cynomolgus monkey were infected with SFV by RT-PCR. Therefore, antiviral drugs must be added in MSCs culture. However, because of SFV infection and additive antiviral drugs, the infection efficiency of the lentiviral vectors reduced significantly. In this study, we improved the infection efficiency by disabled antiviral drugs before lentiviral infection. It might be provide technical assistance for the culture of adult cynomolgus monkey MSCs as genetically engineered cells applied to clinical and experimental research.

Molecular ecology and natural history of simian foamy virus infection in wild-living chimpanzees.

Identifying microbial pathogens with zoonotic potential in wild-living primates can be important to human health, as evidenced by human immunodeficiency viruses types 1 and 2 (HIV-1 and HIV-2) and Ebola virus. Simian foamy viruses (SFVs) are ancient retroviruses that infect Old and New World monkeys and apes. Although not known to cause disease, these viruses are of public health interest because they have the potential to infect humans and thus provide a more general indication of zoonotic exposure risks. Surprisingly, no information exists concerning the prevalence, geographic distribution, and genetic diversity of SFVs in wild-living monkeys and apes. Here, we report the first comprehensive survey of SFVcpz infection in free-ranging chimpanzees (Pan troglodytes) using newly developed, fecal-based assays. Chimpanzee fecal samples (n = 724) were collected at 25 field sites throughout equatorial Africa and tested for SFVcpz-specific antibodies (n = 706) or viral nucleic acids (n = 392). SFVcpz infection was documented at all field sites, with prevalence rates ranging from 44% to 100%. In two habituated communities, adult chimpanzees had significantly higher SFVcpz infection rates than infants and juveniles, indicating predominantly horizontal rather than vertical transmission routes. Some chimpanzees were co-infected with simian immunodeficiency virus (SIVcpz); however, there was no evidence that SFVcpz and SIVcpz were epidemiologically linked. SFVcpz nucleic acids were recovered from 177 fecal samples, all of which contained SFVcpz RNA and not DNA. Phylogenetic analysis of partial gag (616 bp), pol-RT (717 bp), and pol-IN (425 bp) sequences identified a diverse group of viruses, which could be subdivided into four distinct SFVcpz lineages according to their chimpanzee subspecies of origin. Within these lineages, there was evidence of frequent superinfection and viral recombination. One chimpanzee was infected by a foamy virus from a Cercopithecus monkey species, indicating cross-species transmission of SFVs in the wild. These data indicate that SFVcpz (i) is widely distributed among all chimpanzee subspecies; (ii) is shed in fecal samples as viral RNA; (iii) is transmitted predominantly by horizontal routes; (iv) is prone to superinfection and recombination; (v) has co-evolved with its natural host; and (vi) represents a sensitive marker of population structure that may be useful for chimpanzee taxonomy and conservation strategies.

Interspecies transmission of simian foamy virus in a natural predator-prey system.

Simian foamy viruses (SFV) are ancient retroviruses of primates and have coevolved with their host species for as many as 30 million years. Although humans are not naturally infected with foamy virus, infection is occasionally acquired through interspecies transmission from nonhuman primates. We show that interspecies transmissions occur in a natural hunter-prey system, i.e., between wild chimpanzees and colobus monkeys, both of which harbor their own species-specific strains of SFV. Chimpanzees infected with chimpanzee SFV strains were shown to be coinfected with SFV from colobus monkeys, indicating that apes are susceptible to SFV superinfection, including highly divergent strains from other primate species.

Foamy virus vectors: an awaited alternative to gammaretro- and lentiviral vectors.

The first vectors derived from foamy viruses were established over ten years ago. Until now only used and further developed by a handful of investigators these vectors have been shown to be promising tools for the gene transfer into haematopoietic stem cells. Several inherent features of foamy virus-derived vectors, such as the high efficiency in targeting CD34-positive stem cells, a favourable integration profile, and the apathogenic nature of the parental virus, indicate that they are superior to gammaretroviral and lentiviral vectors. The effectiveness in different preclinical animal models suggests the exploration of foamy virus vectors in human trials.