Article
To read the full-text of this research, you can request a copy directly from the authors.

Abstract

Pathogens often have a limited host range, but some can opportunistically jump to new species. Anthropogenic activities that mix reservoir species with novel, hence susceptible, species [1] can provide opportunities for pathogens to spread beyond their normal host range. Furthermore, rapid evolution can produce new pathogens by mechanisms such as genetic recombination [2]. Zoos unintentionally provide pathogens with a high diversity of species from different continents and habitats assembled within a confined space. Institutions alert to the problem of pathogen spread to unexpected hosts can monitor the emergence of pathogens and take preventative measures [3]. However, asymptomatic infections can result in the causative pathogens remaining undetected in their reservoir host. Furthermore, pathogen spread to unexpected hosts may remain undiagnosed if the outcome of infection is limited, as in the case of compromised fertility, or if more severe outcomes are restricted to less charismatic species that prompt only limited investigation. We illustrate this problem here with a recombinant zebra herpesvirus infecting charismatic species including zoo polar bears over at least four years. The virus may cause fatal encephalitis and infects at least five mammalian orders, apparently without requiring direct contact with infected animals [4-8].

No full-text available

Request Full-text Paper PDF

To read the full-text of this research,
you can request a copy directly from the authors.

... A recent study of EHV-1 and EHV-4 isolates from horses in Australia and New Zealand demonstrated widespread recombination between EHV-1 and EHV-4, in particular [23]. Among EHVs isolated from wild equids, infection with a recombinant zebra-borne EHV-1/EHV-9 strain was reported in a polar bear and in an Indian rhinoceros [24,25]. Whether recombination occurred in zebras in Africa or zoos elsewhere is unclear. ...
... In the current study, we confirmed latent EHV-1 and EHV-9 infections in zebras by detection of active LAT transcripts with the simultaneous absence of gB mRNA in the sensory ganglia of zebras. Recently, EHV-1 and EHV-9 have been shown to spread beyond their natural hosts and to infect other zoo animal species, causing fatal neurological disorders [4][5][6][7]24,25]. Latency of EHV-1 and EHV-9 in captive zebras suggests that the threat is determined by reactivation in latent individuals, which is currently not predictable. ...
... However, little is known about the exact mechanism of how recombination occurs and if recombination events happen directly after primary infection, during latency, or during reactivation [21]. Recently, a recombinant EHV-1 containing EHV-9 sequences in the Pol gene was detected in a polar bear and Indian rhinoceros, both of which died after nervous manifestations in Germany [24,25]. It was postulated that zebra might be the host where this recombination took place [25]. ...
Article
Full-text available
Alphaherpesviruses are highly prevalent in equine populations and co-infections with more than one of these viruses’ strains frequently diagnosed. Lytic replication and latency with subsequent reactivation, along with new episodes of disease, can be influenced by genetic diversity generated by spontaneous mutation and recombination. Latency enhances virus survival by providing an epidemiological strategy for long-term maintenance of divergent strains in animal populations. The alphaherpesviruses equine herpesvirus 1 (EHV-1) and 9 (EHV-9) have recently been shown to cross species barriers, including a recombinant EHV-1 observed in fatal infections of a polar bear and Asian rhinoceros. Little is known about the latency and genetic diversity of EHV-1 and EHV-9, especially among zoo and wild equids. Here, we report evidence of limited genetic diversity in EHV-9 in zebras, whereas there is substantial genetic variability in EHV-1. We demonstrate that zebras can be lytically and latently infected with both viruses concurrently. Such a co-occurrence of infection in zebras suggests that even relatively slow-evolving viruses such as equine herpesviruses have the potential to diversify rapidly by recombination. This has potential consequences for the diagnosis of these viruses and their management in wild and captive equid populations.
... Fatal disease with neurological disorder in polar bear (Ursus maritimus) and in black bear (Ursus americanus) have been associated with infection with EHV-9 and a recombinant EHV-9/EHV-1 virus and EHV-1, respectively (Schrenzel et al., 2008;Donovan et al., 2009;Wohlsein et al., 2011;Greenwood et al., 2012). In a zoo, EHV-1-positive guinea pigs (Cavia porcellus) died after ataxia and paralytic disorders, showing also abortions and stillbirths (Wohlsein et al., 2011). ...
... Parameter 5 -Wild reservoir species (or family/order) Zebra species and subspecies have been found infected on several occasions by EHV-1 and the closely related EHV-9 with a possible similar epidemiological pattern to that of horses, such as latency infectious status, widespread infection, role of reservoir, prevalent subclinical infection (Greenwood et al., 2012;Abdelgawad et al., 2015Abdelgawad et al., , 2016Guevara et al., 2018). Reported infections and serological positivity in asymptomatic zebras and other animals housed nearby in zoos suggest the possibility that zebras likely carry the virus as the definitive host and could play a role as a reservoir (Kasem et al., 2008;Donovan et al., 2009;Greenwood et al., 2012). ...
... Parameter 5 -Wild reservoir species (or family/order) Zebra species and subspecies have been found infected on several occasions by EHV-1 and the closely related EHV-9 with a possible similar epidemiological pattern to that of horses, such as latency infectious status, widespread infection, role of reservoir, prevalent subclinical infection (Greenwood et al., 2012;Abdelgawad et al., 2015Abdelgawad et al., , 2016Guevara et al., 2018). Reported infections and serological positivity in asymptomatic zebras and other animals housed nearby in zoos suggest the possibility that zebras likely carry the virus as the definitive host and could play a role as a reservoir (Kasem et al., 2008;Donovan et al., 2009;Greenwood et al., 2012). This role has been confirmed for both EHV-1 and EHV-9, and the co-infection by these two viruses in a zebra was demonstrated, suggesting the origin for recombinant strains (Abdelgawad et al., 2016). ...
Article
Full-text available
Equine Herpesvirus-1 infection has been assessed according to the criteria of the Animal Health Law (AHL), in particular criteria of: Article 7 on disease profile and impacts, Article 5 on the eligibility of the disease to be listed, Article 9 for the categorisation of the disease according to disease prevention and control measures as in Annex IV and Article 8 on the list of animal species related to Equine Herpesvirus-1 infection. The assessment has been performed following a methodology composed of information collection and compilation, and expert judgement on each criterion at individual and collective level. The outcome is the median of the probability ranges provided by the experts, which indicates whether the criterion is fulfilled (66-100%) or not (0-33%), or whether there is uncertainty about fulfilment (33-66%). For the questions where no consensus was reached, the different supporting views are reported. According to the assessment performed, Equine Herpesvirus-1 infection can be considered eligible to be listed for Union intervention according to Article 5 of the Animal Health Law with 33-90% certainty. According to the criteria as in Annex IV of the AHL related to Article 9 of the AHL for the categorisation of diseases according to the level of prevention and control, it was assessed with less than 1% certainty that EHV-1 fulfils the criteria as in Section 1 (category A), 1-5% for the criteria as in Section 2 (category B), 10-66% for the criteria as in Section 3 (category C), 66-90% for the criteria as in Section 4 (category D) and 33-90% for the criteria as in Section 5 (category E). The animal species to be listed for EHV-1 infection according to Article 8(3) criteria are the species belonging to the families of Equidae, Bovidae, Camelidae, Caviidae, Cervidae, Cricetidae, Felidae, Giraffidae, Leporidae, Muridae, Rhinocerontidae, Tapiridae and Ursidae.
... Both EHV-1 and EHV-9 have been shown to accomplish species jumps beyond their natural hosts in captivity resulting in infections of non-equid species (Chowdhury et al., 1988;Crandell et al., 1988;Rebhun et al., 1988;Schrenzel et al., 2008). Recently, a zebra-borne recombinant EHV-1 was detected in polar bears in Germany (Greenwood et al., 2012). In the case reported here a pregnant female Indian rhinoceros aborted in mid-pregnancy, suffered neurological disease and ultimately was euthanized. ...
... Viral DNA was detected in different tissues with nested and qPCR. Molecular characterization demonstrated high (99%) similarity with zebra-borne EHV-1 strain T-616 and Onager-derived EHV-1 (T-529) for the gB gene, and the same level of identity to a previously reported EHV-1 isolated from a polar bear for five genes (Greenwood et al., 2012). The results are discussed in terms of the significance of trans-species transmission in zoological collections. ...
... Clonal sequence analyses did not suggest coinfection with more than one strain. The amplified products were phylogenetically analyzed using Maximum Likelihood and Baysian inference methods (Greenwood et al., 2012) (supplementary Fig. 2). At the nucleotide level, all sequences strongly supported that Purana's sequences belong to an EHV-1 clade that includes the polar bear and zebra EHV-1 sequences and is more distant to horse EHV-1 strains or EHV-9 (Fig. 2). ...
... Equine herpesvirus type 1 (EHV-1; genus Varicellovirus, subfamily alphaherpesvirinae) has been isolated from zebras and other zoo animals [1,8,12,16,21]. Especially, the 1 isolated from zebras has been focused as an emerging agent in zoo animals. The EHV-1 associated with zebras has been called as zebra-borne EHV-1 [1]. ...
... EHV-1 has been isolated from non-equine species including camels, antelopes, cattle, fallow deer, alpacas, llamas and Thomson's gazelle (Eudorcas thomsoni) from which EHV-1 94-137 was isolated [12]. A polar bear (Ursus maritimus), named Jerka, kept in a zoo in Berlin died from acute encephalitis [8], and a strain of zebra-borne EHV-1 was subsequently isolated from it. The nucleotide sequence analysis of the Pol gene (ORF30 in EHV-1 gene nomenclature [18], a homologue of herpes simplex virus 1 (HSV-1) UL30) in this virus indicated that the virus was a recombinant virus between EHV-1 and equine herpesvirus type 9 (EHV-9), which was isolated from an epizootic encephalitis of Thomson's gazelles kept in a zoo in Japan [4,5]. ...
... strains have the neuropathogenic marker D752 in ORF30, indicating that the 3 strains might be neuropathogenic. Greenwood et al. [8] reported nucleotide sequences of ORF10 (a homologue of HSV-1 UL49.5), ORF15 (a homologue of HSV-1 UL45), ORF16 (a homologue of HSV-1 UL44), ORF30, ORF33 (a homologue of HSV-1 UL27) and ORF67 (also called IR6) of the zebra borne EHV-1 isolated from Jerka. ...
Article
Full-text available
A strain of equine herpesvirus type 1 (EHV-1) was isolated from zebra. This strain, called “zebra-borne EHV-1”, was also isolated from an onager and a gazelle in zoological gardens in U.S.A. The full genome sequences of the 3 strains were determined. They shared 99% identities with each other, while they shared 98% and 95% identities with the horse derived EHV-1 and equine herpesvirus type 9, respectively. Sequence data indicated that the EHV-1 isolated from a polar bear in Germany is one of the zebra-borne EHV-1 and not a recombinant virus. These results indicated that zebra-borne EHV-1 is a subtype of EHV-1.
... Both EHV-1 and/or EHV-9 have been shown to infect species in captivity other than their known natural hosts, resulting in disease and fatality in non-perissodactyla species such as polar bear (Ursus maritimus), black bear (U. americanus), llamas (Lama glama), alpacas (Vicugna pacos), blackbuck (Antelopa cervicapra), Thomson's gazelle (Gazelle thomsoni) and giraffe (Giraffa camelopardalis) [7,12,[14][15][16][17][18]. A recombinant zebra-EHV-1/EHV-9 infection was reported in a polar bear and in Indian rhinoceros (Rhinoceros unicornis), in both cases resulting in severe and ultimately fatal neurological symptoms [19,20]. The complete host range of EHV-1 and EHV-9 and whether there are differences in captivity that potentially promote cross-species transmission remains unknown. ...
... EHV-1 antibodies have been detected in Somali wild asses [18], however this is the first report of EHV-9 antibodies in this species. Recombination between EHV-1 and EHV-9 has been observed in sequences isolated from polar bear and Asian rhinoceros [19,20]. The co-occurrence of these viruses in equids suggests that closely related equid alphaherpesviruses have natural opportunities to recombine and help explain the origin of recombinant isolates. ...
... The question of the natural reservoir and definitive host is particularly important for EHV-9 as the source of the many fatal infections of ungulates under natural conditions was not identified. Both EHV-1 and EHV-9 have been involved in fatal encephalitis cases in Prevalence of EHV-1 and -9 in Captive and Wild Animals captive polar bears without proximity to equids [12,19]. Rhinoceroses, which were not considered a potential source of infection, may have been involved in these unexplained transmission events. ...
Article
Full-text available
Equine herpesvirus type 1 (EHV-1) causes respiratory disorders and abortion in equids while EHV-1 regularly causes equine herpesvirus myeloencephalopathy (EHM), a stroke-like syndrome following endothelial cell infection in horses. Both EHV-1 and EHV-9 infections of non-definitive hosts often result in neuronal infection and high case fatality rates. Hence, EHV-1 and EHV-9 are somewhat unusual herpesviruses and lack strict host specificity, and the true extent of their host ranges have remained unclear. In order to determine the seroprevalence of EHV-1 and EHV-9, a sensitive and specific peptide-based ELISA was developed and applied to 428 sera from captive and wild animals representing 30 species in 12 families and five orders. Members of the Equidae, Rhinocerotidae and Bovidae were serologically positive for EHV-1 and EHV-9. The prevalence of EHV-1 in the sampled wild zebra populations was significantly higher than in zoos suggesting captivity may reduce exposure to EHV-1. Furthermore, the seroprevalence for EHV-1 was significantly higher than for EHV-9 in zebras. In contrast, EHV-9 antibody prevalence was high in captive and wild African rhinoceros species suggesting that they may serve as a reservoir or natural host for EHV-9. Thus, EHV-1 and EHV-9 have a broad host range favoring African herbivores and may have acquired novel natural hosts in ecosystems where wild equids are common and are in close contact with other perissodactyls.
... Over the past ten years natural recombination has been assessed in alphaherpesviruses from five non-human mammalian hosts; EHV-1, EHV-4, EHV-9, FeHV-1 and PRV (Greenwood et al., 2012; Pagamjav et al., 2005; Vaz et al., 2016a; Vaz et al., 2016b; Ye et al., 2016) (Table 2). This represents only a small proportion of the alphaherpesviruses of importance in veterinary medicine. ...
... Germany by Greenwood et al., (2012) following analysis of the sequence of six virus genes using a distance based method within the Recombination Analysis Tool 1.0 (RAT v1.0). Interestingly, the recombinant was isolated from a polar bear with fatal encephalitis, even though neither EHV-1 nor EHV-9 naturally infect polar bears (Greenwood et al., 2012). ...
... Germany by Greenwood et al., (2012) following analysis of the sequence of six virus genes using a distance based method within the Recombination Analysis Tool 1.0 (RAT v1.0). Interestingly, the recombinant was isolated from a polar bear with fatal encephalitis, even though neither EHV-1 nor EHV-9 naturally infect polar bears (Greenwood et al., 2012). More recent work has suggested that the recombination event involving these two viruses most likely occurred in zebras and was then transmitted to the polar bear (Abdelgawad et al., 2016). ...
Article
Full-text available
Recombination in alphaherpesviruses was first described more than sixty years ago. Since then, different techniques have been used to detect recombination in natural (field) and experimental settings. Over the last ten years, next-generation sequencing (NGS) technologies and bioinformatic analyses have greatly increased the accuracy of recombination detection, particularly in field settings, thus contributing greatly to the study of natural alphaherpesvirus recombination in both human and veterinary medicine. Such studies have highlighted the important role that natural recombination plays in the evolution of many alphaherpesviruses. These studies have also shown that recombination can be a safety concern for attenuated alphaherpesvirus vaccines, particularly in veterinary medicine where such vaccines are used extensively, but also potentially in human medicine where attenuated varicella zoster virus vaccines are in use. This review focuses on the contributions that NGS and sequence analysis have made over the last ten years to our understanding of recombination in mammalian and avian alphaherpesviruses, with particular focus on attenuated live vaccine use download at: https://authors.elsevier.com/a/1URWo5aKq22e5q
... Par ailleurs, ces travaux ont mis en lumière des sites de recombinaison entre des souches d'EHV-1 et d'EHV-4, ou encore des souches d'EHV-1 et d'EHV-8. Une autre étude portant cette fois sur une souche d'EHV-1 ayant franchi la barrière de l'espèce et infecté un ours polaire (Ursus maritimus), a mis en évidence une zone de recombinaison entre l'EHV-1 et l'EHV-9 (Greenwood et al. 2012). Cette dernière suspicion de recombinaison a cependant été contestée par la suite par Guo et al. (Guo et al. 2014). ...
... Il n'existe pas beaucoup d'information, à notre connaissance, quant à d'éventuelles recombinaisons avec l'EHV-3 même si elles avaient été suspectées entre l'EHV-1, l'EHV-3 et l'EHV-4 lors d'une étude menée par Staczek et al. en 1983 (Staczek, Atherton andO'Callaghan 1983). Enfin, concernant l'EHV-9, lors d'une étude menée en 2012, Greenwood et al. ont suspecté une zone de recombinaison entre l'EHV-1 et l'EHV-9 au niveau de l'ORF30 d'une souche d'EHV-1 isolée chez un ours polaire, mais cette suspicion a ensuite été contestée par Guo et al. en 2014(Greenwood et al. 2012Guo et al. 2014). Une autre étude menée à plus large échelle par Kolb et al. (2017) s'est portée sur l'ensemble des herpèsvirus appartenant au genre des Varicellovirus. ...
Thesis
L’herpèsvirus équin 1 (EHV-1), classé parmi les Herpesviridae, est l’un des pathogènes les plus fréquemment rencontrés au sein de la population équine. Ce virus peut être à l’origine de formes de maladie respiratoire, abortive ou néonatale, oculaire ou nerveuse pouvant conduire à l’euthanasie de l’animal. Les mesures de prévention incluant la vaccination constituent la première barrière dans la lutte contre ce virus. En cas d’épizootie, le suivi et l’identification des souches peuvent constituer une aide précieuse pour la compréhension des modes d’infection. Ce travail de thèse a porté sur le suivi moléculaire par différentes approches d’une population de souches d’EHV-1 isolées en France entre 2009 et 2018. Le typage de 137 souches d’EHV-1 pour le polymorphisme en position 2254 du gène de l’ADN polymérase (ORF30) associé à la pathogénicité des souches a montré une association significative des souches de type A2254 avec les cas d’avortements. Quatorze souches d’intérêt ont ensuite été analysées par séquençage de l’ORF30 et Multi Locus Sequence Typing (MLST). Le séquençage complet de l’ORF30 s’est montré peu discriminant mais a permis de mettre en évidence un troisième génotype (C2254 (H752)) en position 2254 de l’ORF30. L’analyse par MLST a permis de classer ces souches parmi 7 des 10 clades UL décrits par l’outil à ce jour. Deux autres souches d’herpèsvirus d’équidés, une d’EHV-8 et une d’EHV-9, ont pu être mises en évidence grâce au séquençage de l’ORF30. Ces différentes souches d’alphaherpèsvirus des équidés ont été caractérisées, comparées et leur sensibilité vis-à-vis de sept molécules antivirales a été évaluée in vitro. Leur proximité génétique a pu être visualisée par la construction d’un network phylogénétique basé sur les séquences de l’ORF30. Une seconde partie du travail a porté sur le suivi du niveau de protection conféré par les anticorps neutralisant l’EHV-1. Une technique innovante de quantification a été mise au point grâce à la technologie xCELLigence® sur cellules E. Derm et a permis de déterminer les titres induisant 50% de la neutralisation d’une concentration de virus (NT50) à partir d’une population d’échantillons qualifiés provenant d’une infection expérimentale avec la souche d’EHV-1 C2254. Les valeurs de NT50 obtenues ont montré une bonne corrélation avec les valeurs de titres obtenues avec la méthode de référence sur cellules RK13. Les réponses de chevaux à la vaccination ont ensuite pu être étudiées selon le protocole de vaccination reçu et selon l’historique des chevaux.
... Although horses are natural hosts for equine herpesviruses, several viruses have recently been discovered that are closely related to EHV-1 and were isolated from both captive and wild animals, including Thompson's gazelles (Eudorcas thomsoni) 4 , Ilamas (Lama glama) 5 , alpacas (Vicugna pacos) 6 , black bears (Ursus americanus), polar bears (Ursus maritimus) 7 and rhinos (Rhinoceros unicornis) 8,9 . Serological and genetic evidence suggests that zebra-conspecific and rhinos in Africa are frequently exposed to EHV-1 and its close relative EHV-9, which may even reservoir in rhinos (Abdelgawad et al. 2015). ...
... As this study has demonstrated EHV-1 to be stable under different environmental water conditions, the water sources would provide EHV-1 with the opportunity to infect potential hosts sharing the water source. This in turn may account for the recent reports of EHV-1-like viruses detected in non-equid species in captivity, as water is a potential transmission conduit for the virus between different species [4][5][6][7][8][9] . ...
Article
Full-text available
For viruses to utilize environmental vectors (hard surfaces, soil, water) for transmission, physical and chemical stability is a prerequisite. There are many factors including pH, salinity, temperature, and turbidity that are known to contribute to the ability of viruses to persist in water. Equine herpesvirus type-1 (EHV-1) is a pathogenic alphaherpesvirus associated with domestic horses and wild equids. EHV-1 and recombinants of EHV-1 and EHV-9 are able to cause infections in non-equid animal species, particularly in captive settings. Many of the captive non-equid mammals are not naturally sympatric with equids and do not share enclosures, however, in many cases water sources may overlap. Similarly, in the wild, equids encounter many species at waterholes in times of seasonal drought. Therefore, we hypothesized that EHV-1 is stable in water and that water may act as a vector for EHV-1. In order to establish the conditions promoting or hindering EHV-1 longevity, infectivity and genomic stability in water; we exposed EHV-1 to varied water environments (pH, salinity, temperature, and turbidity) in controlled experiments over 21 days. The presence and infectivity of the virus was confirmed by both qPCR and cell culture experiments. Our results show that EHV-1 remains stable and infectious under many conditions in water for up to three weeks.
... The zebra-borne EHV-1 strains are known to form a group of EHV-1 viruses, closely similar but distinct from horsederived type EHV-1 strains. 1,[10][11][12]15,17 They are sometimes referred to as "variants" or a "subtype" of horse-derived EHV-1. The full genome sequences of the 3 zebra-borne EHV-1 strains have been determined and compared to the full genome sequences of horse-derived type 1 EHVs and of EHV-9. ...
... Interestingly, other neuropathogenic zebra-borne EHV-1 infections in a Thomson's gazelle, a polar bear, and hamsters were characterized by nonsuppurative meningoencephalitis with no viral inclusions (Table 1). 11,15,16 This variation in neuropathologic findings between several zebra-borne EHV-1 infections might be associated with the virulence characteristics of the specific viral strain, the host's immune status, and the animal species affected. Horse-derived EHV-1 causing EHM is vasculotropic and causes endothelial necrosis, thrombosis, perivascular lymphocytic cuffing, and hemorrhage without intranuclear inclusion bodies and neuron localization. ...
Article
We describe the histopathologic, immunohistochemical, and molecular features of a case of meningoencephalitis in a Thomson's gazelle ( Eudorcas thomsonii) naturally infected with zebra-borne equid herpesvirus 1 (EHV-1) and the implications for the molecular detection of zebra-borne EHV-1. A 4-y-old female Thomson's gazelle was submitted for postmortem examination; no gross abnormalities were noted except for meningeal congestion. Microscopic evaluation demonstrated multifocal nonsuppurative meningoencephalitis with intranuclear eosinophilic and amphophilic inclusion bodies and EHV-9 antigen in neurons. PCR demonstrated the presence of a herpesvirus with a nucleotide sequence 99-100% identical to the corresponding sequences of zebra-borne EHV-1 and of EHV-9 strains. To determine whether EHV-1 or EHV-9 was involved, a PCR with a specific primer set for EHV-9 ORF59/60 was used. The sequence was identical to that of 3 recognized zebra-borne EHV-1 strains and 91% similar to that of EHV-9. This isolate was designated as strain LM2014. The partial glycoprotein G ( gG) gene sequence of LM2014 was also identical to the sequence of 2 zebra-borne EHV-1 strains (T-529 isolated from an onager, 94-137 from a Thomson's gazelle). The histologic lesions of encephalitis and antigen localization in this gazelle indicate prominent viral neurotropism, and lesions were very similar to those seen in EHV-1- and EHV-9-infected non-equid species. Histologic lesions caused by EHV-9 and zebra-borne EHV-1 are therefore indistinguishable.
... With medical intervention Lars survived but Jerka died. The cause of death was identified as a recombinant equine herpesvirus (EHV)-1/EHV-9 derived from zebras (Greenwood et al., 2012). The same study found no evidence that Lars and Knut's illnesses were related, because recombinant EHVs were not detected by genetic or serological methods (Greenwood et al., 2012). ...
... The cause of death was identified as a recombinant equine herpesvirus (EHV)-1/EHV-9 derived from zebras (Greenwood et al., 2012). The same study found no evidence that Lars and Knut's illnesses were related, because recombinant EHVs were not detected by genetic or serological methods (Greenwood et al., 2012). ...
Article
This report describes three possibly related incidences of encephalitis, two of them lethal, in captive polar bears (Ursus maritimus). Standard diagnostic methods failed to identify pathogens in any of these cases. A comprehensive, three stage diagnostic ‘pipeline’, employing both standard serological methods and new DNA microarray and next generation sequencing-based diagnostics was developed,in part as a consequence of this initial failure. This pipeline approach illustrates the strengths, weaknesses and limitations of these tools in determining pathogen caused deaths in non-model organisms such as wildlife species and why the use of a limited number of diagnostic tools may fail to uncover important wildlife pathogens.
... Ruminants have long been known as reservoirs for many alphaherpesvirus species, and the vast majority coevolve with their hosts. However, the presence of the same bovine virus has been documented in many mammal species, with some hosts apparently acting as mixing vessels, whereby animals are coinfected simultaneously with multiple species of alphaherpesviruses, thus increasing the likelihood for evolution through, for example, recombination (39,40). ...
... Of the nine known equine herpesviruses (EHVs), which are alphaherpesviruses (EHV-1, EHV-3, EHV-4, EHV-6, EHV-8, and EHV-9) and gammaherpesviruses (EHV-2, EHV-5, and EHV-7), some show evidence of frequent host transfers in mammals (39). In fact, the main feature that appears to prevent host jumps of equine gammaherpesviruses is complete absence of equids from a given environment. ...
Article
Herpesviruses are ubiquitous and can cause disease in all classes of vertebrates but also in animals of lower taxa, including molluscs. It is generally accepted that herpesviruses are primarily species specific, although a species can be infected by different herpesviruses. Species specificity is thought to result from host-virus coevolutionary processes over the long term. Even with this general concept in mind, investigators have recognized interspecies transmission of several members of the Herpesviridae family, often with fatal outcomes in non-definitive hosts-that is, animals that have no or only a limited role in virus transmission. We here summarize herpesvirus infections in wild mammals that in many cases are endangered, in both natural and captive settings. Some infections result from herpesviruses that are endemic in the species that is primarily affected, and some result from herpesviruses that cause fatal disease after infection of non-definitive hosts. We discuss the challenges of such infections in several endangered species in the absence of efficient immunization or therapeutic options. Expected final online publication date for the Annual Review of Virology Volume 5 is September 29, 2018. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... The virus seems to have originated in zebras, but it is unclear at present whether the recombination occurred in captive zebras in zoos or wild zebras in natural habitats before importation. The recombination resulted in a pathogen fatal for polar bears, while zebras remain asymptomatic and likely carry the virus as the definitive host (Greenwood et al., 2012). It is still unclear whether EHV-1/EHV-9 recombination is a single event or happens more frequently. ...
Article
The equine herpesviruses type 1 (EHV-1) and 4 (EHV-4) are ubiquitous pathogens that affect horse populations on all continents. Despite widespread vaccination, EHV-1 and EHV-4 infections remain a permanent risk. While the two viruses share a high degree of genetic and antigenic similarity, they differ significantly in host range and pathogenicity. Compared to EHV-4, which mainly infects horses and causes respiratory disease, EHV-1 has a broader host range and can result in respiratory disease, abortions, neonatal death, and equine herpesvirusmyeloencephalopathy (EHM). Recent studies have elucidated a number of mechanisms that may, at least partly, explain the differential pathogenic potential of the two viruses. While both EHV-1 and EHV-4 can escape host immune responses and establish latent infection, there are differences with respect to virus entry and their ability to interfere with the innate immune response. Understanding the virus' repertoire of immunomodulatory mechanisms may lead the way to develop more efficient vaccines.
... It is surprising, therefore, that herpesviruses tend to be closely associated with a specific host species. However, this concept is challenged recently as some herpesviruses seem to be endemic in more than one species, observations that go beyond the wellknown species jumps and often fatal diseases in non-definitive hosts (Abdelgawad et al. 2014;Greenwood et al. 2012;Wohlsein et al. 2011;Huff and Barry 2003). ...
Article
The entry process of herpesviruses into host cells is complex and highly variable. It involves a sequence of well-orchestrated events that begin with virus attachment to glycan-containing proteinaceous structures on the cell surface. This initial contact tethers virus particles to the cell surface and results in a cascade of molecular interactions, including the tight interaction of viral envelope glycoproteins to specific cell receptors. These interactions trigger intracellular signaling and finally virus penetration after fusion of the viral envelope with cellular membranes. Based on the engaged cellular receptors and co-receptors, and the subsequent signaling cascades, the entry pathway will be decided on the spot. A number of viral glycoproteins and many cellular receptors and molecules have been identified as players in one or several of these events during virus entry. This chapter will review viral glycoproteins, cellular receptors and signaling cascades associated with the very first interactions of herpesviruses with their target cells.
... These findings are not completely unexpected since such natural inter-species recombination events have already been documented in other mammalian herpesviruses (equine herpesviruses; Pagamjav et al. 2005;Greenwood et al. 2012). What may be more surprising is that recombination within HSV-1 and HSV-2 has often been investigated and detected during the last decade (for a complete review on recombination in alphaherpesviruses, see Loncoman et al. 2016). ...
Article
Full-text available
Herpes simplex viruses 1 and 2 (HSV-1 and HSV-2) are seen as close relatives but also unambiguously considered as evolutionary independent units. Here, we sequenced the genomes of 18 HSV-2 isolates characterized by divergent UL30 gene sequences to further elucidate the evolutionary history of this virus. Surprisingly, genome-wide recombination analyses showed that all HSV-2 genomes sequenced to date contain HSV-1 fragments. Using phylogenomic analyses, we could also show that two main HSV-2 lineages exist. One lineage is mostly restricted to sub-Saharan Africa while the other has reached a global distribution. Interestingly, only the worldwide lineage is characterized by ancient recombination events with HSV-1. Our findings highlight the complexity of HSV-2 evolution, a virus of putative zoonotic origin which later recombined with its human-adapted relative. They also suggest that co-infections with HSV-1 and 2 may have genomic and potentially functional consequences and should therefore be monitored more closely. 3
... Most commonly, herpesvirus infection presents as a benign, self-limiting disease with a mild viraemia. Herpesvirus rarely causes widespread disseminated disease in the natural host, although crossspecies transmission to non-reservoir hosts can also result in lethal infections [2]. In general neonates and immune-compromised hosts are at higher risk of developing a fatal herpesvirus infection [3,4]. ...
Article
Full-text available
Elephants are classified as critically endangered animals by the International Union for Conservation of Species (IUCN). Elephant endotheliotropic herpesvirus (EEHV) poses a large threat to breeding programs of captive Asian elephants by causing fatal haemorrhagic disease. EEHV infection is detected by PCR in samples from both clinically ill and asymptomatic elephants with an active infection, whereas latent carriers can be distinguished exclusively via serological assays. To date, identification of latent carriers has been challenging, since there are no serological assays capable of detecting seropositive elephants. Here we describe a novel ELISA that specifically detects EEHV antibodies circulating in Asian elephant plasma/serum. Approximately 80 % of PCR positive elephants display EEHV-specific antibodies. Monitoring three Asian elephant herds from European zoos revealed that the serostatus of elephants within a herd varied from non-detectable to high titers. The antibody titers showed typical herpes-like rise-and-fall patterns in time which occur in all seropositive animals in the herd more or less simultaneously. This study shows that the developed ELISA is suitable to detect antibodies specific to EEHV. It allows study of EEHV seroprevalence in Asian elephants. Results confirm that EEHV prevalence among Asian elephants (whether captive-born or wild-caught) is high.
... However, there are viruses where this does not apply (e.g. Greenwood et al. 2012;Shi et al. 2014). Adaptation of virus attachment proteins to the cell receptor used by viruses to gain entry to host cells can alter virus spread across host species and has resulted in epidemics in novel hosts (Moya et al. 2004;Allison et al. 2014;Shi et al. 2014). ...
Article
Was the 1993/1994 fatal canine distemper virus (CDV) epidemic in lions and spotted hyaenas in the Serengeti ecosystem caused by the recent spillover of a virulent domestic dog strain or one well adapted to these noncanids? We examine this question using sequence data from 13 ‘Serengeti’ strains including five complete genomes obtained between 1993 and 2011. Phylogenetic and haplotype network analyses reveal that strains from noncanids during the epidemic were more closely related to each other than to those from domestic or wild canids. All noncanid ‘Serengeti’ strains during the epidemic encoded: (1) one novel substitution G134S in the CDV-V protein; and (2) the rare amino acid combination 519I/549H at two sites under positive selection in the region of the CDV-H protein that binds to SLAM (CD 150) host cell receptors. Worldwide, only a few noncanid strains in the America II lineage encode CDV-H 519I/549H. All canid ‘Serengeti’ strains during the epidemic coded CDV-V 134G, and CDV-H 519R/549Y, or 519R/549H. A functional assay of cell entry revealed the highest performance by CDV-H proteins encoding 519I/549H in cells expressing lion SLAM receptors, and the highest performance by proteins encoding 519R/549Y, typical of dog strains worldwide, in cells expressing dog SLAM receptors. Our findings are consistent with an epidemic in lions and hyaenas caused by CDV variants better adapted to noncanids than canids, but not with the recent spillover of a dog strain. Our study reveals a greater complexity of CDV molecular epidemiology in multihost environments than previously thought.
... CDV is not just able to infect multiples species; indeed, the present analysis shows that CDV can infect five different Orders. As far as is known, only a few critically emerging and re-emerging viruses such as the Equid herpesvirus 9 causing fatal encephalitis [73], the West Nile Virus [74] or the Influenza A virus [75] have been reported to infect five or more different Orders. ...
Article
Full-text available
Background Canine distemper virus (CDV) is the etiological agent of one of the most infectious diseases of domestic dogs, also known as a highly prevalent viral infectious disease of carnivores and posing a conservation threat to endangered species around the world. To get a better panorama of CDV infection in different Orders, a retrospective and documental systematic review of the role of CDV in different non-dog hosts was conducted. The bibliographical data were collected from MedLine/PubMed and Scopus databases. Data related to Order, Family, Genus and Species of the infected animals, the presence or absence of clinical signs, mortality, serological, molecular or antigenic confirmation of CDV infection, geographic location, were collected and summarized. Results Two hundred seventeen scientific articles were considered eligible which includes reports of serological evaluation, and antigenic or genomic confirmation of CDV infection in non-dog hosts. CDV infects naturally and experimentally different members of the Orders Carnivora (in 12 Families), Rodentia (four Families), Primates (two Families), Artiodactyla (three Families) and Proboscidea (one Family). The Order Carnivora (excluding domestic dogs) accounts for the vast majority (87.5 %) of the records. Clinical disease associated with CDV infection was reported in 51.8 % of the records and serological evidence of CDV infection in apparently healthy animals was found in 49.5 % of the records. High mortality rate was showed in some of the recorded infections in Orders different to Carnivora. In non-dog hosts, CDV has been reported all continents with the exception of Australasia and in 43 different countries. Conclusions The results of this systematic review demonstrate that CDV is able to infect a very wide range of host species from many different Orders and emphasizes the potential threat of infection for endangered wild species as well as raising concerns about potential zoonotic threats following the cessation of large-scale measles vaccination campaigns in the human population. Electronic supplementary material The online version of this article (doi:10.1186/s12917-016-0702-z) contains supplementary material, which is available to authorized users.
... wileyonlinelibrary.com/journal/zoo fatal (neuropathogenic) infections in non-equine species (Abdelgawad et al., 2014;Greenwood et al., 2012). In equids, EHV typically persists as a latent infection and can be reactivated when the immune system is compromised, for example in response to stress (Pusterla et al., 2009). ...
... Two Grevy's zebras (Equus grevysi) housed near the polar bear were the likely source of EHV-9 in the polar bear (Schrenzel et al. 2008). A recombinant zebra EHV-1 virus was identified as the cause of acute neurologic disease in two captive adult bears with evidence of exposure in another asymptomatic bear; the source of the virus was not identified and the mode of transmission was also unknown (Greenwood et al. 2012). An outbreak of suid herpesvirus 1 has been described among a group of eight circus bears, including four polar bears, in Spain; the source of the infection was suspected to be infected pig heads procured from local slaughterhouses and fed to the circus animals (Banks et al. 1999). ...
Article
Full-text available
Disease was a listing criterion for the polar bear (Ursus maritimus) as threatened under the Endangered Species Act in 2008; it is therefore important to evaluate the current state of knowledge and identify any information gaps pertaining to diseases in polar bears. We conducted a systematic literature review focused on infectious agents and associated health impacts identified in polar bears. Overall, the majority of reports in free-ranging bears concerned serosurveys or fecal examinations with little to no information on associated health effects. In contrast, most reports documenting illness or pathology referenced captive animals and diseases caused by etiologic agents not representative of exposure opportunities in wild bears. As such, most of the available infectious disease literature has limited utility as a basis for development of future health assessment and management plans. Given that ecological change is a considerable risk facing polar bear populations, future work should focus on cumulative effects of multiple stressors that could impact polar bear population dynamics.
... Similarly, they apparently misunderstand our recommendations for managing disease and inbreeding risk when planning translocations, protocols that have successfully fostered population re-establishment (Slotow & Hunter 2009). Finally, they offer nothing to demonstrate that using captive founders, especially those of mongrel, opportunistic provenance promoted by private owners such as ALERT, is a preferable alternative (see Greenwood et al., 2012, for further evidence of the risks of exotic disease for captive carnivores). ...
... It is also unknown why these viruses should so readily coexist, though ecological mechanisms such as simultaneous transmission (codispersal), the availability of requisite resources, and/or shared benefits associated with host immunomodulation by one or more of these vi-ruses may explain the observed cooccurrence. Recombination is also a possible consequence of coinfection and is a common feature in the ecology and evolution of herpesviruses (30)(31)(32)(33)(34)(35)(36). ...
Article
Full-text available
Unlabelled: The majority of emerging zoonoses originate in wildlife, and many are caused by viruses. However, there are no rigorous estimates of total viral diversity (here termed "virodiversity") for any wildlife species, despite the utility of this to future surveillance and control of emerging zoonoses. In this case study, we repeatedly sampled a mammalian wildlife host known to harbor emerging zoonotic pathogens (the Indian Flying Fox, Pteropus giganteus) and used PCR with degenerate viral family-level primers to discover and analyze the occurrence patterns of 55 viruses from nine viral families. We then adapted statistical techniques used to estimate biodiversity in vertebrates and plants and estimated the total viral richness of these nine families in P. giganteus to be 58 viruses. Our analyses demonstrate proof-of-concept of a strategy for estimating viral richness and provide the first statistically supported estimate of the number of undiscovered viruses in a mammalian host. We used a simple extrapolation to estimate that there are a minimum of 320,000 mammalian viruses awaiting discovery within these nine families, assuming all species harbor a similar number of viruses, with minimal turnover between host species. We estimate the cost of discovering these viruses to be ~$6.3 billion (or ~$1.4 billion for 85% of the total diversity), which if annualized over a 10-year study time frame would represent a small fraction of the cost of many pandemic zoonoses. Importance: Recent years have seen a dramatic increase in viral discovery efforts. However, most lack rigorous systematic design, which limits our ability to understand viral diversity and its ecological drivers and reduces their value to public health intervention. Here, we present a new framework for the discovery of novel viruses in wildlife and use it to make the first-ever estimate of the number of viruses that exist in a mammalian host. As pathogens continue to emerge from wildlife, this estimate allows us to put preliminary bounds around the potential size of the total zoonotic pool and facilitates a better understanding of where best to allocate resources for the subsequent discovery of global viral diversity.
... For example, ethical dilemmas surrounding the translocation of wildlife populations from native habitats to new environments, including temporary relocations to ex situ facilities such as zoos and aquariums, raise a set of difficult technical, philosophic, and ethical questions for conservation scientists and wildlife biologists (Minteer and Collins 2010). Beyond the animal welfare or animal rights concerns about handling and moving animals that may experience considerable stress (or even mortality) during this process, such practices will also have implications for (1) the original source ecosystems (i.e., the community-level impacts of removing individuals from populations stressed by climate change), (2) the temporary ex situ facility that houses the animals (including shifts in resources and collection space as well as risks of disease transmission) (e.g., Greenwood et al. 2012), and (3) the native species present in the eventual " recipient " ecosystems once the wildlife are introduced (Ricciardi and Simberloff 2009). Another example is the practice of ecological engineering for species conservation in the wild, which can involve the significant modification (and even invention) of habitat to improve field conservation efforts. ...
Article
Ethical obligations to animals in conservation research and management are manifold and often conflicting. Animal welfare concerns often clash with the ethical imperative to understand and conserve a population or ecosystem through research and management intervention. The accelerating pace and impact of global environmental change, especially climate change, complicates our understanding of these obligations. One example is the blurring of the distinction between ex situ (zoo- and aquarium-based) conservation and in situ (field-based) approaches as zoos and aquariums become more active in field conservation work and as researchers and managers consider more intensive interventions in wild populations and ecosystems to meet key conservation goals. These shifts, in turn, have consequences for our traditional understanding of the ethics of wildlife research and management, including our relative weighting of animal welfare and conservation commitments across rapidly evolving ex situ and in situ contexts. Although this changing landscape in many ways supports the increased use of captive wildlife in conservation-relevant research, it raises significant ethical concerns about human intervention in populations and ecosystems, including the proper role of zoos and aquariums as centers for animal research and conservation in the coming decades. Working through these concerns requires a pragmatic approach to ethical analysis, one that is able to make trade-offs among the many goods at stake (e.g., animal welfare, species viability, and ecological integrity) as we strive to protect species from further decline and extinction in this century.
... A first glimpse at this potential situation where animals are normally segregated in nature begin to cohabitate can already be seen in zoos. For example, there have been reported cases of cross-species herpesvirus transmission resulting in fatalities, such as zebra-borne Equid herpesvirus 1 (EHV-1) infecting an Indian rhinoceros and polar bears (Abdelgawad et al. 2014;Greenwood et al. 2012). Thus, understanding how herpesviruses spread in populations must be better understood. ...
Chapter
Interindividual spread of herpesviruses is essential for the virus life cycle and maintenance in host populations. For most herpesviruses, the virus-host relationship is close, having coevolved over millions of years resulting in comparatively high species specificity. The mechanisms governing interindividual spread or horizontal transmission are very complex, involving conserved herpesviral and cellular proteins during the attachment, entry, replication, and egress processes of infection. Also likely, specific herpesviruses have evolved unique viral and cellular interactions during cospeciation that are dependent on their relationship. Multiple steps are required for interindividual spread including virus assembly in infected cells; release into the environment, followed by virus attachment; and entry into new hosts. Should any of these steps be compromised, transmission is rendered impossible. This review will focus mainly on the natural virus-host model of Marek’s disease virus (MDV) in chickens in order to delineate important steps during interindividual spread.
... When the EHV-1 strains were examined using the PHI recombination test (Table 1), statistically significant recombination was detected (p = <0.001). Wild equine derived EHV-1 strains cause severe infections, often neurological in both equine and non-equine captive animals [66][67][68][69][70], and some domestic horse EHV-1.2 strains can also cause myeloencephalopathy [71,72]. A SNP (D752) within the polymerase gene of domestic horse viruses has been shown to influence the neurological disease phenotype, and is shared among wild equine strains [73,74]. ...
Article
Full-text available
Background: The varicelloviruses comprise a genus within the alphaherpesvirus subfamily, and infect both humans and other mammals. Recently, next-generation sequencing has been used to generate genomic sequences of several members of the Varicellovirus genus. Here, currently available varicellovirus genomic sequences were used for phylogenetic, recombination, and genetic distance analysis. Results: A phylogenetic network including genomic sequences of individual species, was generated and suggested a potential restriction between the ungulate and non-ungulate viruses. Intraspecies genetic distances were higher in the ungulate viruses (pseudorabies virus (SuHV-1) 1.65%, bovine herpes virus type 1 (BHV-1) 0.81%, equine herpes virus type 1 (EHV-1) 0.79%, equine herpes virus type 4 (EHV-4) 0.16%) than non-ungulate viruses (feline herpes virus type 1 (FHV-1) 0.0089%, canine herpes virus type 1 (CHV-1) 0.005%, varicella-zoster virus (VZV) 0.136%). The G + C content of the ungulate viruses was also higher (SuHV-1 73.6%, BHV-1 72.6%, EHV-1 56.6%, EHV-4 50.5%) compared to the non-ungulate viruses (FHV-1 45.8%, CHV-1 31.6%, VZV 45.8%), which suggests a possible link between G + C content and intraspecies genetic diversity. Varicellovirus clade nomenclature is variable across different species, and we propose a standardization based on genomic genetic distance. A recent study reported no recombination between sequenced FHV-1 strains, however in the present study, both splitstree, bootscan, and PHI analysis indicated recombination. We also found that the recently sequenced Brazilian CHV-1 strain BTU-1 may contain a genetic signal in the UL50 gene from an unknown varicellovirus. Conclusion: Together, the data contribute to a greater understanding of varicellovirus genomics, and we also suggest a new clade nomenclature scheme based on genetic distances.
... Once infected leukocytes reach the CNS after establishing a cell-associated viraemia, EHV-1 can infect equid endothelial cells from brain capillaries, resulting in an inflammatory cascade, with thrombosis and necrosis [2,[5][6][7]. Outbreaks of encephalitis caused by EHV-1 have also been reported in non-equid species [8,9]. ...
Article
Equine herpesvirus type 1 (EHV-1) is an emerging pathogen that causes encephalomyelitis in horses and non-equid species. Several aspects of the immune response in the central nervous system (CNS), mainly regarding the role of inflammatory mediators during EHV-1 encephalitis, remain unknown. Moreover, understanding the mechanisms underlying extensive neuropathology induced by viruses would be helpful to establish therapeutic strategies. Therefore, we aimed to evaluate some aspects of the innate immune response during highly neurovirulent EHV-1 infection. C57BL/6 mice infected intranasally with A4/72 and A9/92 EHV-1 strains developed a fulminant neurological disease at 3 days post-inoculation with high viral titres in the brain. These mice developed severe encephalitis with infiltration of monocytes and CD8 ⁺ T cells to the brain. The inflammatory infiltrate followed the detection of the chemokines CCL2, CCL3, CCL4, CCL5, CXCL2, CXCL9 and CXCL-10 in the brain. Notably, the levels of CCL3, CCL4, CCL5 and CXCL9 were higher in A4/72-infected mice, which presented higher numbers of inflammatory cells within the CNS. Pro-inflammatory cytokines, such as interleukins (ILs) IL-1α, IL-1β, IL-6, IL-12β, and tumour necrosis factor (TNF), were also detected in the CNS, and Toll-like receptor (TLR) TLR2, TLR3 and TLR9 genes were also upregulated within the brain of EHV-1-infected mice. However, no expression of interferon-γ (IFN-γ) and IL-12α, which are important for controlling the replication of other herpesviruses, was detected in EHV-1-infected mice. The results show that the activated innate immune mechanisms could not prevent EHV-1 replication within the CNS, but most likely contributed to the extensive neuropathology. The mouse model of viral encephalitis proposed here will also be useful to study the mechanisms underlying extensive neuropathology.
... The co-occurrence of EHV-1 and EHV-8 also needs to be monitored because EHVs have the potential to diversify rapidly by recombination. Recombinant EHV-1/EHV-9 infections that originated in asymptomatic zebras were reported to cause non-fatal and fatal encephalitis in polar bears [55] and abortion and neurological disease in Indian rhinoceroses [56]. ...
Article
Full-text available
Equid herpesvirus 8 (EHV-8), formerly known as asinine herpesvirus 3, is an alphaherpesvirus that is closely related to equid herpesviruses 1 and 9 (EHV-1 and EHV-9). The pathogenesis of EHV-8 is relatively little studied and to date has only been associated with respiratory disease in donkeys in Australia and horses in China. A single EHV-8 genome sequence has been generated for strain Wh in China, but is apparently incomplete and contains frameshifts in two genes. In this study, the complete genome sequences of four EHV-8 strains isolated in Ireland between 2003 and 2015 were determined by Illumina sequencing. Two of these strains were isolated from cases of abortion in horses, and were misdiagnosed initially as EHV-1, and two were isolated from donkeys, one with neurological disease. The four genome sequences are very similar to each other, exhibiting greater than 98.4% nucleotide identity, and their phylogenetic clustering together demonstrated that genomic diversity is not dependent on the host. Comparative genomic analysis revealed 24 of the 76 predicted protein sequences are completely conserved among the Irish EHV-8 strains. Evolutionary comparisons indicate that EHV-8 is phylogenetically closer to EHV-9 than it is to EHV-1. In summary, the first complete genome sequences of EHV-8 isolates from two host species over a twelve year period are reported. The current study suggests that EHV-8 can cause abortion in horses. The potential threat of EHV-8 to the horse industry and the possibility that donkeys may act as reservoirs of infection warrant further investigation.
... The Thomson's gazelles involved in the outbreak in Japan shared pasture and a water source with zebras that were considered a possible source of the virus. Originally, the EHV-9 virus is thought to have evolved in Burchell's zebra (Greenwood et al. 2012), and EHV-9 glycoprotein B (gB)-specific sequences were found in the trigeminal ganglia of a Burchell's zebra from the Serengeti ecosystem (Borchers et al. 2008). The virus has a tropism for neuronal and respiratory tissue during experimental infections (Donovan et al. 2009). ...
Article
Full-text available
Equid herpesviruses types 1 (EHV-1) and 9 (EHV-9) are unusual among herpesviruses in that they lack strong host specificity, and the full extent of their host range remains unclear. The virus establishes latency for long periods and can be reactivated and shed, resulting in clinical disease in susceptible species. A sensitive and specific peptide-based enzyme-linked immunosorbent assay was developed to study the seroprevalence of both viruses in a broad range of species among both wild and captive populations. We used this assay to study the seroprevalences of EHV-1 and EHV-9 in a natural population of the highly endangered Grévy's zebra ( Equus grevyi) in Kenya, sampled during a 4-yr period (2012-2015). The results were compared with those obtained from captive Grévy's zebras from a previous study. The wild population had a significantly higher seroprevalence of EHV-9 compared with the captive population, suggesting that captivity might reduce exposure to this serotype. In contrast, the seroprevalences of EHV-1 between captive and wild groups was not significantly different. The seroprevalence of EHV-9 was not significantly higher than EHV-1 in zebras within the wild Kenyan population.
... Captive-animal environments may inadvertently serve as hot spots for pathogens to concentrate and mix in nonsympatric species. This has led to polar bears being exposed to pathogens they would never encounter in their natural environment (1)(2)(3)(4). Viral pathogens such as equine herpesvirus (EHV) types 1 (1) and 9 (3,5), West Nile virus (WNV) (2), and suid herpesvirus (SuHV) (4) have been responsible for many serious and often lethal infections in captive polar bears in Europe and the United States. Polar bears at the Zoological Gardens, Wuppertal, Germany, were found to be infected with EHV-1, while polar bears at the Zoological Society of San Diego were infected with EHV-9. ...
Article
Full-text available
Cross-species transmission of viral pathogens is becoming an increasing problem for captive-animal facilities. This study highlights how animals in captivity are vulnerable to novel opportunistic pathogens, many of which do not result in straightforward diagnosis from symptoms and histopathology. In this study, a novel pathogen was suspected to have contributed to the death of a juvenile polar bear. HTS techniques were employed, and a novel Mastadenovirus was isolated. The virus was present in both the tissue and blood samples. Phylogenetic analysis of the virus at both the gene and genome levels revealed that it is highly divergent to other known mastadenoviruses. Overall, this study shows that animals in isolated conditions still come into contact with novel pathogens, and for many of these pathogens, the host reservoir and mode of transmission are yet to be determined.
... Virus was thought to have been transmitted via contact with a fomite rather than direct contact with an infected equid. Lesions consist of nonsuppurative encephalitis with neuronal necrosis and gliosis (Greenwood, 2012;Schrenzel, 2008). Intranuclear basophilic inclusions may be present in glial cells and neurons. ...
... Acute EHV infection can cause clinical signs including pharyngitis, pneumonia, pyrexia, lymphadenopathy, abortion, and neuropathies due to acute myeloencephalopathy (3,5,9,10). Furthermore, it has been shown that EHVs can occasionally infect a range of nonequid species in captivity, with potentially fatal outcomes (11)(12)(13)(14)(15)(16). ...
Article
Equid herpesviruses (EHVs) establish latent infections, and many EHVs are shed and transmitted via nasal discharge primarily through droplet and aerosol infection. Obtaining nasal swabs and other invasive samples from wildlife is often not possible without capture and physical restraint of individuals, which are resource intensive and a health risk for the captured animals. Fecal EHV shedding has never been demonstrated for naturally infected equids. We established the conditions for fecal EHV screening, and our results suggest that testing fecal samples is an effective noninvasive approach for monitoring acute EHV shedding in equids.
... EHV-1 and -9 have been reported in captive Indian rhinoceros (Rhinoceros unicornis) [8], black bears (Ursus americanus) [9], Thomson's gazelles (Eudorcas thomsoni) [46], guinea pigs (Cavia porcellus f. dom.) [9], and polar bears (Ursus maritimus) [45]. The infected animals often display typical symptoms associated with EHV including encephalitis and sever neurological disease; the severe nature of the symptoms has often resulted in infected animals being euthanized [47]. Phylogenetic analysis of EHV-1 and -9 sequences isolated from different species of zebra showed they were nearly identical to EHVs isolated from different equine species, highlighting the ability of both EHV-1 and -9 to cross species barriers. ...
Article
Full-text available
Equine herpesviruses (EHV) are a major health concern for domestic and wild equids and represent one of the most economically important disease agents of horses. Most known EHVs are transmitted directly between individuals as a result of direct exposure to exudates and aerosols. However, accumulating evidence suggests that environmental transmission may play a role including air, water, and fomites. Here, we reviewed studies on environmental stability and transmission of EHVs, which may influence viral dynamics and the use of environmental samples for monitoring EHV shedding.
... This has also been observed in herpes simplex virus 1 (HSV-1) strains [36]. Furthermore inter-species recombination has been detected in field samples between EHV-1 and EHV-4 [34,37], EHV-1 and EHV-9 [38], and between EHV-1 and EHV-8 [35]. Thus, although the genotyping method based on the targeted multi-locus approach used in this study and also recently developed for VZV [39] may be more practical than NGS for surveillance purposes, it has the limitation that it does not allow the detection of possible recombination crossovers in the unanalysed parts of the genome [40]. ...
Article
Full-text available
Multiple locus typing based on sequencing heterologous regions in 26 open reading frames (ORFs) of equine herpesvirus 1 (EHV-1) strains Ab4 and V592 was used to characterise 272 EHV-1 isolates from 238 outbreaks of abortion, respiratory or neurological disease over a 28-year period. The analysis grouped the 272 viruses into at least 10 of the 13 unique long region (UL) clades previously recognised. Viruses from the same outbreak had identical multi-locus profiles. Sequencing of the ORF68 region of EHV-1 isolates from 222 outbreaks established a divergence into seven groups and network analysis demonstrated that Irish genotypes were not geographically restricted but clustered with viruses from all over the world. Multi-locus analysis proved a more comprehensive method of strain typing than ORF68 sequencing. It was demonstrated that when interpreted in combination with epidemiological data, this type of analysis has a potential role in tracking virus between premises and therefore in the implementation of targeted control measures. Viruses from 31 of 238 outbreaks analysed had the proposed ORF30 G2254/D752 neuropathogenic marker. There was a statistically significant association between viruses of the G2254/D752 genotype and both neurological disease and hypervirulence as defined by outbreaks involving multiple abortion or neurological cases. The association of neurological disease in those with the G2254/D752 genotype was estimated as 27 times greater than in those with the A2254/N752 genotype.
... However, along with EHV-1, EHV-9 also has been found to be show nervous form in nature as reported in Thomson's gazelles, which died of encephalitis in an outbreak and from calves [104]. EHV-1 and EHV-9 often found to show jumping behavior from equids to other non-equid species including polar bears, Giraffe, and Indian rhinoceros [105][106][107]. Both EHV-1 and EHV-2 are economically important viruses affecting the respiratory tract of horses. ...
Article
Full-text available
The importance of horse (Equus caballus) to equine practitioners and researchers cannot be ignored. An unevenly distributed population of equids harbors numerous diseases, which can affect horses of any age and breed. Among these, the affections of nervous system are potent reason for death and euthanasia in equids. Many episodes associated with the emergence of equine encephalitic conditions have also pose a threat to human population as well, which signifies their pathogenic zoonotic potential. Intensification of most of the arboviruses is associated with sophisticated interaction between vectors and hosts, which supports their transmission. The alphaviruses, bunyaviruses, and flaviviruses are the major implicated groups of viruses involved with equines/humans epizootic/epidemic. In recent years, many outbreaks of deadly zoonotic diseases such as Nipah virus, Hendra virus, and Japanese encephalitis in many parts of the globe addresses their alarming significance. The equine encephalitic viruses differ in their global distribution, transmission and main vector species involved, as discussed in this article. The current review summarizes the status, pathogenesis, pathology, and impact of equine neuro-invasive conditions of viral origin. A greater understanding of these aspects might be able to provide development of advances in neuro-protective strategies in equine population.
... 10,18 A recombinant EHV-1 containing EHV-9 sequences in the Pol gene was isolated from a polar bear (U. maritimus) and Indian rhinoceros (Rhinoceros unicornis) in Germany, both of which had died of acute encephalitis. 1,13 The full genome sequences between the 3 strains of EHV-1 virus isolated from a zebra (E. grevyi), onager (E. ...
Article
Full-text available
Encephalitis in hamsters, which was induced by equine herpesvirus (EHV)–9, EHV-1 strain Ab4p, and zebra-borne EHV-1, was investigated and compared to assess viral kinetics and identify the progression and severity of neuropathological findings. Hamsters were inoculated with EHV-9, EHV-1 strain Ab4p, and zebra-borne EHV-1 via the nasal route and euthanized at 24, 48, 72, 96, 120, 144, and 168 hours postinoculation (HPI). The inoculated hamsters had mild to severe neurological signs at 60 to 72, 96, and 120 HPI, and the mortality rate was 75%, 0%, and 0% for animals inoculated with EHV-9, EHV-1 strain Ab4p, and zebra-borne EHV-1 viruses, respectively. Inoculated hamsters had varying degrees of rhinitis and lymphoplasmacytic meningoencephalitis, as well as differences in the severity and distribution of cerebral lesions. Furthermore, the cellular distribution of viral antigen depended on the inoculated virus. Neuronal necrosis was widely detected in animals inoculated with EHV-9, while marked perivascular cuffs of infiltrating inflammatory cells and gliosis were detected in animals inoculated with EHV-1 strain Ab4p and zebra-borne EHV-1. In the present study, 3 viruses belonging to the herpesvirus family induced encephalitis after initial propagation in the nasal cavity. These viruses might travel to the brain via the olfactory pathway and/or trigeminal nerve, showing different distributions and severities of neuropathological changes.
... Infecția a fost confirmată prin izolarea virusului din creier (38) . Anticorpi au fost identificați şi la urşi negri de Florida (39) . ...
Article
Full-text available
Populația de urși din țara noastră este în continuă creștere, reprezentând în prezent circa 40% din urșii existenți în Europa (1) (2). Studiile făcute asupra populațiilor de urși atât cei sălbatici, cât și cei din grădini zoologice, pe perioade variabile de timp (de 1-2 sau mai mulți ani), evidențiază un complex de agenți microbieni, reprezentat de virusuri, bacterii și paraziți. Prezența virusurilor este evidențiată prin manifestarea clinică a unor boli, cu izolarea sau identificarea lor în umori sau țesuturi (situație mai rar raportată), și prin examene serologice de evidențiere a anticorpilor specifici (cele mai multe studii). În această lucrare ne-am propus să prezentăm, în sinteză, date despre virusurile care circulă în populațiile de urși, bolile în care sunt implicate, conexiunile epidemiologice cu alte specii și riscul pentru sănătatea publică. Au fost identificate virusuri cu următoarea încadrare taxonomică: virusuri cu ARN din familiile Flaviviridae, Togaviridae, Orthomyxoviridae, Paramyxoviridae, Phenuiviridae, Peribunyaviridae, Picornaviridae, Reoviridae, Rhabdoviridae, Caliciviridae) și virusuri cu ADN din familiile Adenoviridae, Herpes-viridae, Parvoviridae.
... L'HVE-8, anciennement appelé herpèsvirus asin 3, est connu pour provoquer des symptômes principalement chez les ânes (Browning et al., 1988;Liu et al., 2012) mais a été récemment détecté chez des chevaux (Garvey et al., 2018). L'HVE-9 est connu pour infecter les zèbres mais a aussi été retrouvé chez de nombreuses espèces de mammifères, telles que la gazelle de Thomson, l'ours polaire et la girafe (Donovan et al., 2009;Fukushi et al., 1997;Greenwood et al., 2012;Kasem et al., 2008;Schrenzel et al., 2008). Les chevaux ne sont pas les hôtes naturels de l'HVE-9 mais ce virus s'est montré capable d'infecter cette espèce lors d'une étude expérimentale . ...
Thesis
Neuf herpèsvirus sont connus pour infecter les équidés. Parmi eux, l’herpèsvirus équin 1 (HVE-1) induit les formes de la maladie les plus graves. En effet, ce virus provoque des troubles respiratoires, des avortements, des morts néonatales et des troubles nerveux qui mènent souvent à l’euthanasie de l’animal. La prophylaxie, reposant sur les bonnes pratiques sanitaires et la vaccination, demeure le meilleur moyen de lutte contre l’ensemble des herpèsvirus équins. Des vaccins qui réduisent efficacement les troubles respiratoires et la dissémination de l’HVE-1 ont été développés. Cependant, ces derniers ne préviennent pas les avortements et n’ont aucun effet démontré contre la forme nerveuse. De plus, la couverture vaccinale demeure insuffisante en France. Les traitements aux antiviraux constituent donc une approche complémentaire dans la lutte contre l’HVE-1. Cependant, trop peu d’études ont évalué l’effet de molécules contre ce virus, limitant les perspectives d’utilisation. Pour répondre à cette problématique, nous avons développé un protocole de criblage à moyen/haut débit à l’aide de la technologie RTCA xCELLigence®, basée sur la mesure d’impédance cellulaire. Suite au criblage de 2891 molécules, 21 candidats ont été identifiés pour leur efficacité contre l’HVE-1. Parmi ceux-ci, l’aphidicoline, la décitabine, le ganciclovir, l’idoxuridine, le pritelivir et le valganciclovir ont présenté les meilleurs effets. L’activité de ces molécules a été confirmée sur différents modèles cellulaires en présence de différentes souches d’HVE-1. Cette étude a conduit à l’identification et à l’étude du mode d’action d’une nouvelle molécule efficace contre l’HVE-1, la décitabine. Cet analogue de la déoxycitidine a également montré un effet synergique in vitro lorsqu’elle est associée au valganciclovir. Lors de la seconde phase de ce travail, nous avons testé l’efficacité d’un traitement au valganciclovir lors d’un challenge expérimental avec infection par nébulisation d’une souche d’HVE-1 (C2254) récemment isolée au cours de l’épizootie de 2018. Cette étude a permis de démontrer qu’une dose de 6,5 mg/kg de valganciclovir, administrée 2 fois par jour, permettait de maintenir un bon niveau de protection avant la mise en place de la réponse immunitaire humorale. En effet, ce traitement permet de réduire les signes cliniques, l’excrétion virale et la virémie cellulaire induits par l’HVE-1. Ces travaux réalisés in vivo démontrent l’efficacité du valganciclovir contre l’HVE-1 et le criblage réalisé in vitro ouvre de nouvelles perspectives de traitement, en particulier avec des associations de molécules.
... Viruses adapted to using water as a vector may compensate for host species heterogeneity by evolving a reduced host specificity and virulence. The absence of host specificity by several EHV species is supported by multiple lines of evidence (Abdelgawad et al., 2014;Kennedy et al., 1996;Lunn et al., 2009;House et al., 1991;Rebhun et al., 1988;Greenwood et al., 2012). Among Namibian wildlife, multiple non-equid species, including rhinoceros and bovids, were shown to be infected with EHV-1 and EHV-9 (Abdelgawad et al., 2014). ...
Article
In climates with seasonally limited precipitation, terrestrial animals congregate at high densities at scarce water sources. We hypothesize that viruses can exploit the recurrence of these diverse animal congregations to spread. In this paper, we test the central prediction of this hypothesis -- that viruses employing this transmission strategy remain stable and infectious in water. Equid herpesviruses (EHVs) were chosen as a model as they have been shown to remain stable and infectious in water for weeks under laboratory conditions. Using fecal data from wild equids from a previous study, we establish that EHVs are shed more frequently by their hosts during the dry season, increasing the probability of water source contamination with EHV. We document the presence of several strains of EHVs present in high genome copy number from the surface water and sediments of waterholes sampled across a variety of mammalian assemblages, locations, temperatures and pH. Phylogenetic analysis reveals that the different EHV strains found exhibit little divergence despite representing ancient lineages. We employed molecular approaches to show that EHVs shed remain stable in waterholes with detection decreasing with increasing temperature in sediments. Infectivity experiments using cell culture reveals that EHVs remain infectious in water derived from waterholes. The results are supportive of water as an abiotic viral vector for EHV.
Article
The Bornean sun bear (Helarctos malayanus euryspilus) is the smallest subspecies of sun bear. Their numbers are declining, and more research is needed to better understand their health and biology. Forty-four bears housed at the Bornean Sun Bear Conservation Centre (BSBCC) in Sabah, Malaysia, were screened for known and novel viruses in November 2018. Ursid γ-herpesvirus type 1 (UrHV-1) is a herpesvirus that has been detected from swab samples of clinically healthy sun bears and biopsy samples of oral squamous cell carcinoma in sun bears. We detected an UrHV-1-related virus from throat and rectal swabs by molecular viral screening in samples from 15.9% of the sun bears at BSBCC. None of the bears with the UrHV-1-related virus in this study had oral lesions. There is no known report of UrHV-1 detection in the wild sun bear population, and its association with oral squamous cell carcinoma is not fully understood. Finding an UrHV-1-related virus in a rehabilitation center is a concern because conditions in captivity may contribute to spreading this virus, and there is the potential of introducing it into wild populations when a bear is released. This study demonstrates an urgent need to carry out similar surveillance for sun bears in captivity as well as those in the wild, to better understand the impact of captivity on the prevalence and spread of UrHV-1-related viruses. Positive bears also should be monitored for oral lesions to better understand whether there is a causal relationship.
Article
Equid alphaherpesvirus 1 (EHV-1) causes myeloencephalopathy in horses and occasionally in non-equid species. Although mouse models have been developed to understand EHV-1 pathogenesis, few EHV-1 strains have been identified as highly neurovirulent to mice. The aim of this study was to evaluate the pathogenesis of 2 neurovirulent EHV-1 strains in mice, and to characterize the inflammatory cells and expression of chemokines and the apoptosis marker caspase-3 in the brain of infected mice. C57BL/6J mice were inoculated intranasally with EHV-1 strains A4/72 or A9/92 and evaluated on 1, 2, and 3 days post inoculation (DPI). EHV-1-infected mice showed severe neurological signs at 3 DPI. Ultrastructural analysis revealed numerous viral nucleocapsids and fewer enveloped virions within degenerated and necrotic neurons and in the surrounding neuropil. Histologically, at 3 DPI, there was severe diffuse neuronal degeneration and liquefactive necrosis, prominent microgliosis, and perivascular cuffing composed of CD3 ⁺ cells (T cells) and Iba-1 ⁺ cells (macrophages), mainly in the olfactory bulb and ventral portions of the brain. In these areas, moderate numbers of neuroglial cells expressed CCL5 and CCL2 chemokines. Numerous neurons, including those in less affected areas, were immunolabeled for cleaved caspase-3. In conclusion, neurovirulent EHV-1 strains induced a fulminant necrotizing lymphohistiocytic meningoencephalitis in mice, with microgliosis and expression of chemokines and caspase-3. This model will be useful for understanding the mechanisms underlying the extensive neuropathology induced by these viral infections.
Article
A herd of seven captive-born Grevy's zebras (Equus grevyi) experienced an outbreak of nasal discharge and sneezing. Clinical signs, including lethargy and anorexia, were severe and acute in three animals, including a 16-mo-old male that died within 48 h. Treatment of two severely affected zebras included valacyclovir (40 mg/kg PO), meloxicam (0.6 mg/kg IM/PO), and cefquinome (2.5 mg/kg IM q48h). An adult female improved rapidly, and clinical signs resolved within 48 h of treatment. Administration of valacyclovir pellets was very complicated in a 2-mo-old female, and death occurred within 48 h. Histologic examination of the two individuals that died revealed severe fibrinonecrotic interstitial pneumonia with prominent hyaline membranes and type II pneumocyte hyperplasia. Additionally, the 16-mo-old male presented systemic endothelial activation with vascular thrombosis and necrosis and mild nonsuppurative meningoencephalitis. Herpesviral DNA was detected in the lungs of both individuals by nested polymerase chain reaction. The nucleic acid sequence of the amplicons showed 100% similarity with previously published equid alphaherpesvirus 9 sequences. Three additional animals developed mild nasal discharge only and recovered spontaneously. The zebras shared housing facilities with other species, including white rhinoceros (Ceratotherium simum), reticulated giraffe (Giraffa camelopardalis reticulata), and several antelope species. None of these animals showed clinical signs. Additionally, nasal swabs and whole blood samples were collected from cohoused white rhinoceroses (n = 3) and springboks (Antidorcas marsupialis, n = 3) as well as nasal swabs from cohoused reticulated giraffes (n = 4). Nucleic acid sequence from equid herpesviruses was not detected in any of these samples. The source of the infection in the zebras remains unclear.
Article
Full-text available
The Alphacoronavirus‐1 species include viruses that infect numerous mammalian species. To better understand the wide host range of these viruses, better knowledge on the molecular determinants of virus‐host cell entry mechanisms in wildlife hosts is essential. We investigated Alphacoronavirus‐1 infection in carnivores using long‐term data on Serengeti spotted hyenas (Crocuta crocuta) and molecular analyses guided by the tertiary structure of the viral spike (S) attachment protein’s interface with the host receptor aminopeptidase N (APN). We sequenced the complete 3’‐end region of the genome of 9 variants from wild African carnivores, plus the APN gene of 15 wild carnivore species. Our results revealed two outbreaks of Alphacoronavirus‐1 infection in spotted hyenas associated with genetically distinct canine coronavirus type II (CCoVII) variants. Within the receptor binding domain (RBD) of the S gene the residues that directly bind to the APN receptor were conserved in all variants studied, even those infecting phylogenetically diverse host taxa. We identified a variable region within RBD located next to a region that directly interacts with the APN receptor. Two residues within this variable region were under positive selection in hyena variants, indicating that both sites were associated with adaptation of CCoVII to spotted hyena APN. Analysis of APN sequences revealed that most residues that interact with the S protein are conserved in wild carnivores, whereas some adjacent residues are highly variable. Of the variable residues, four that are critical for virus‐host binding were under positive selection and may modulate the efficiency of virus attachment to carnivore APN.
Article
Full-text available
Across the world, elephant endotheliotropic herpesvirus is increasingly killing elephant calves and threatening the long-term survival of the Asian elephant, a species that is currently facing extinction. This article presents three open-ended stories of elephant care in times of death and loss: at places of confinement and elephant suffering like the zoos in Seattle and Zürich as well as in the conflict-ridden landscapes of South India, where the country’s last free-ranging elephants live. Our stories of deadly viral-elephant-human becomings remind us that neither human care, love, and attentiveness nor techniques of control and creative management are sufficient to fully secure elephant survival. The article introduces the concept of “viral creep” to explore the ability of a creeping, only partially knowable virus to rearrange relations among people, animals, and objects despite multiple experimental human regimes of elephant care, governance, and organization. The viral creep exceeds the physical and intellectual contexts of human interpretation and control. It reminds us that uncertainty and modes of imaging are always involved when we make sense of the world around us.
Thesis
Full-text available
Cetaceans are sentinels of the marine environment, currently threatened by many factors, mainly anthropogenic. The most easily identified compromising conditions are those affecting the skin and external mucosae - good indicators of the cetacean’s health status. Cutaneous and mucocutaneous lesions have been extensively reported in wild and captive cetaceans, but little is known about the involved etiological factors, evolution of the dermatological lesions and their systemic consequences. Viruses are the most commonly involved agents in cutaneous and mucocutaneous lesions, especially herpesviruses (HV) and, associated with skin and mucosal lesions with varying morphologies, and cetacean poxviruses (CePV), mainly associated with characteristic “tattoo” or “ring” skin lesions. In addition, fungal agents are also recognized as causative agents of dermatological disease in cetaceans, especially in the process known as paracoccidioidomycosis ceti, observed as raised proliferative whitish lesions, caused by non cultivable yeast of Paracoccidioides brasiliensis (order Onygenales). Despite being reported worldwide, the occurrence of these etiological agents in southern Atlantic cetaceans is still poorly understood. The goal of this study was to identify and characterize selected cutaneous and mucocutaneous pathogens (HV, CePV and P. brasiliensis) of free-ranging cetaceans from Brazil, and to design more sensitive diagnostic methods for their detection. All the studied cetaceans stranded along the coast of Brazil, between 2005 and 2015, except three wild riverine dolphins that were physically contained and released after sample collection. In order to achieve our goals, we employed molecular, histopathological, and occasionally immunohistochemical and electron microscopy techniques. The presence of HV and CePV was evaluated in cutaneous, and oral and genital mucosal samples from 115 specimens and skin samples from 113 individuals, respectively; whereas the presence of members of the genus Onygenales sp. was evaluated in four specimens presenting macroscopic compatible lesions. Skin or oral mucosal samples from four animals were HV PCR-positive: a whitish ulcerated skin lesion from a Guiana dolphin (Sotalia guianensis), a lingual sample from an Atlantic spotted dolphin (Stenella frontalis), ulcerative lesions and healthy skin samples from a dwarf sperm whale (Kogia sima), and a proliferative skin lesion from a Bolivian river dolphin (Inia boliviensis). The tree first animals were infected with alphaherpesvirus. A sequence more similar to gammaherpesvirus was obtained from the Bolivian river dolphin’s proliferative skin lesion. The Bolivian river dolphin sequence could possibly be a member of a new gammaherpesvirus genus. Additionally, all other available tissue samples from HV-positive specimens, aside from skin and oral mucosa, were also tested by PCR and histologically evaluated. A different alphaherpesvirus sequence was found in the stomach and in a mesenteric lymph node of the dwarf sperm whale. Microscopic findings in two HV-positive animals (chronic proliferative dermatitis in Bolivian river dolphin and Guiana dolphin) were compatible with HV. CePV was identified in “tattoo” skin lesions of an Atlantic bottlenose dolphin and a Guiana dolphin by established molecular methods, and poxviral particles were observed by electron microscopy. CePV-positive animals presented epidermal ballooning degeneration and occasionally small, pale eosinophilic or amphophilic intracytoplasmic inclusions, compatible with CePV. Specific amino acid motifs for all CePV were also identified, reinforcing the suggestion of the new Cetaceanpoxvirus genus. We also designed novel SYBR® Green real-time and conventional CePV PCR methods significantly more sensitive than those currently available in the literature. An additional Guiana dolphin, previously negative based in established PCR methods was diagnosed positive for CePV through these new techniques. Refractile yeasts (4−9 μm in diameter) were observed under light microscopy in mild granulomatous and necrotic skin lesions of four Atlantic bottlenose dolphin, and for the first time, in a skeletal muscle abscess (the former possibly indicating the invasive potential of the agent).Onygenales sp. yeasts were identified in skin lesions by immunohistochemistry and a sequence of P. brasiliensis, more similar (100% nucleotide identity) to the one described in an Atlantic bottlenose dolphin from Cuba than to human or any other terrestrial mammals cases in Brazil, was obtained from the skin lesion of one of the specimens, confirming the etiological agent of these type of lesions. Herein we report the first molecular identification of HV in South American cetaceans and in riverine dolphins worldwide. This study also describes the first amplification of CePV and P. brasiliensis in odontocetes from South America. Four of the five novel herpesvirus sequences herein identified are possibly novel species, tentatively named Delphinid HV-10, Kogiid HV-2, Kogiid HV-3 and Iniid HV-1.
Article
Full-text available
Equine Herpesviruses (EHV) are prevalent and often latent pathogens of equids which can be fatal when transmitted to non-equids. Stress and elevated glucocorticoids have been associated with EHV reactivation in domestic horses, but little is known about the correlation between stress and viral reactivation in wild equids. We investigated the effect of an environmental stressor (social group restructuring following a translocation event) on EHV reactivation in captive Grévy’s zebras (Equus grevyi). A mare was translocated by road transport from Zoo Mulhouse, France, to join a resident group of three mares in Tierpark Berlin, Germany. We used an indirect sampling method to assess the frequency of EHV shedding for 14 days immediately after the translocation event (termed the ‘experimental period’). The results were compared with those from two control periods, one preceding and one subsequent to the experimental period. In addition, we measured fecal glucocorticoid metabolite (fGCM) concentrations daily in all individuals from 6 days before, to 14 days after translocation. We found significantly higher EHV shedding frequencies during the experimental period, compared to each of the two control periods. All animals showed significantly elevated fGCM concentrations, compared to fGCM levels before translocation. Finally, we found that an increase in fGCM concentration was significantly associated with an increased likelihood of EHV shedding. Although the small number of animals in the study limits the conclusions that can be drawn from the study, taken together, our results support the hypothesis that environmental stressors induce viral reactivation in wild equids. Our results suggest that potentials stressors such as group restructuring and translocation should be considered in the management of zoological collections to reduce the risk of fatal EHV infections in novel hosts. Moreover, environmental stressors may play an important role in EHV reactivation and spreading in wild equid populations.
Article
Age-associated neurodegenerative changes, including amyloid β(Aβ) plaques, neurofibrillary tangles (NFTs), and amyloid angiopathy comparable to those seen in the brains of human patients with Alzheimer’s disease (AD), have been reported in the brains of aged bears. However, the significance of these findings in bears is unclear due to the difficulty in assessing cognitive impairment and the lack of standardized approaches for the semiquantitative evaluation of Aβ plaques and NFTs. In this study, we evaluate the neuropathologic changes in archival brain tissue of 2 aged polar bears ( Ursus maritimus , ages 28 and 37) using the National Institute of Aging-Alzheimer Association (NIA-AA) consensus guidelines for the neuropathologic assessment of Alzheimer’s Disease (AD). Both bears had an Aβ (A) score of 3 of 3, Braak stage (B score) of 2 of 3, and neuritic plaque (C) score of 3 of 3. These findings are consistent with the neurodegenerative changes observed in brains of patients with AD. The application of NIA-AA consensus guidelines, as applied to the neuropathologic assessment of the aged bears in this report, demonstrates the use of standardized semiquantitative assessment systems for comparative, translational studies of aging in a vulnerable wildlife species.
Article
Full-text available
Dengue virus is an emerging infectious agent that infects an estimated 50-100 million people annually worldwide, yet current diagnostic practices cannot detect an etiologic pathogen in ∼40% of dengue-like illnesses. Metagenomic approaches to pathogen detection, such as viral microarrays and deep sequencing, are promising tools to address emerging and non-diagnosable disease challenges. In this study, we used the Virochip microarray and deep sequencing to characterize the spectrum of viruses present in human sera from 123 Nicaraguan patients presenting with dengue-like symptoms but testing negative for dengue virus. We utilized a barcoding strategy to simultaneously deep sequence multiple serum specimens, generating on average over 1 million reads per sample. We then implemented a stepwise bioinformatic filtering pipeline to remove the majority of human and low-quality sequences to improve the speed and accuracy of subsequent unbiased database searches. By deep sequencing, we were able to detect virus sequence in 37% (45/123) of previously negative cases. These included 13 cases with Human Herpesvirus 6 sequences. Other samples contained sequences with similarity to sequences from viruses in the Herpesviridae, Flaviviridae, Circoviridae, Anelloviridae, Asfarviridae, and Parvoviridae families. In some cases, the putative viral sequences were virtually identical to known viruses, and in others they diverged, suggesting that they may derive from novel viruses. These results demonstrate the utility of unbiased metagenomic approaches in the detection of known and divergent viruses in the study of tropical febrile illness.
Article
Full-text available
Pseudorabies is caused by Suid herpesvirus 1, a member of the Alphaherpesvirinae subfamily. Although pigs are the natural host of Pseudorabies virus (PRV), the virus has a broad host range and may cause fatal encephalitis in many species. The United States obtained PRV-free status in 2004 after the virus was eradicated from domestic swineherds, but the virus is still present in feral swine populations. The current report describes PRV infection in 3 dogs that were used to hunt feral swine. The dogs developed clinical signs including facial pruritus with facial abrasions, dyspnea, vomiting, diarrhea, ataxia, muscle stiffness, and death. Two were euthanized, and 1 died within approximately 48 hr after onset of clinical signs. The salient histologic changes consisted of neutrophilic trigeminal ganglioneuritis with neuronophagia and equivocal intranuclear inclusion bodies. Pseudorabies virus was isolated from fresh tissues from 2 of the dogs, and immunohistochemistry detected the virus in the third dog. Virus sequencing and phylogeny, based upon available GenBank sequences, revealed that the virus was likely a field strain that was closely related to a cluster of PRV strains previously identified in Illinois. Though eradicated from domestic swine in the United States, PRV is present in populations of feral swine, and should therefore continue to be considered a possible cause of disease in dogs and other domestic animals with compatible clinical history and signs. Continued surveillance is necessary to prevent reintroduction of PRV into domestic swine.
Article
Full-text available
To identify attributes of zoological institutions and surveillance system factors that were associated with participation in the West Nile Virus Surveillance System for Zoological Institutions in the USA, and to assess the potential effectiveness of zoos as a novel data source for surveillance of emerging infectious zoonoses. Retrospective. The number of specimens submitted between August 2001 and December 2006 for West Nile virus testing was determined for each institution. Descriptive statistics were used to summarize the distribution of number of specimens submitted and features of the institutions. Student's t-test was used to assess potential associations between institutional and animal collection characteristics and the total number of specimens submitted by each institution. Factors associated with institutional participation include: submitting specimens for specific purposes of serosurvey testing, sentinel surveillance, vaccine titre checks, vaccine effectiveness, submitting specimens for multiple reasons, and communication with public health. Understanding how zoo and surveillance system characteristics are associated with participation in this surveillance effort may enhance public health efforts and the design of future zoological surveillance efforts.
Article
Full-text available
The critical assessment of bovine adenoviruses (BAdV) as indicators of environmental fecal contamination requires improved knowledge of their prevalence, shedding dynamics, and genetic diversity. We examined DNA extracted from bovine and other animal waste samples collected in Wisconsin for atadenoviruses and mastadenoviruses using novel, broad-spectrum PCR primer sets. BAdV were detected in 13% of cattle fecal samples, 90% of cattle urine samples, and 100% of cattle manure samples; 44 percent of BAdV-positive samples contained both Atadenovirus and Mastadenovirus DNA. Additionally, BAdV were detected in soil, runoff water from a cattle feedlot, and residential well water. Overall, we detected 8 of 11 prototype BAdV, plus bovine, rabbit, and porcine mastadenoviruses that diverged significantly from previously reported genotypes. The prevalence of BAdV shedding by cattle supports targeting AdV broadly as indicators of the presence of fecal contamination in aqueous environments. Conversely, several factors complicate the use of AdV for fecal source attribution. Animal AdV infecting a given livestock host were not monophyletic, recombination among livestock mastadenoviruses was detected, and the genetic diversity of animal AdV is still underreported. These caveats highlight the need for continuing genetic surveillance for animal AdV and for supporting data when BAdV detection is invoked for fecal source attribution in environmental samples. To our knowledge, this is the first study to report natural BAdV excretion in urine, BAdV detection in groundwater, and recombination in AdV of livestock origin.
Article
Full-text available
Comparative analysis of molecular sequence data is essential for reconstructing the evolutionary histories of species and inferring the nature and extent of selective forces shaping the evolution of genes and species. Here, we announce the release of Molecular Evolutionary Genetics Analysis version 5 (MEGA5), which is a user-friendly software for mining online databases, building sequence alignments and phylogenetic trees, and using methods of evolutionary bioinformatics in basic biology, biomedicine, and evolution. The newest addition in MEGA5 is a collection of maximum likelihood (ML) analyses for inferring evolutionary trees, selecting best-fit substitution models (nucleotide or amino acid), inferring ancestral states and sequences (along with probabilities), and estimating evolutionary rates site-by-site. In computer simulation analyses, ML tree inference algorithms in MEGA5 compared favorably with other software packages in terms of computational efficiency and the accuracy of the estimates of phylogenetic trees, substitution parameters, and rate variation among sites. The MEGA user interface has now been enhanced to be activity driven to make it easier for the use of both beginners and experienced scientists. This version of MEGA is intended for the Windows platform, and it has been configured for effective use on Mac OS X and Linux desktops. It is available free of charge from http://www.megasoftware.net.
Article
Full-text available
A 12-year-old female polar bear (Ursus maritimus) developed a sudden onset of muscle tremors, erratic circling, increased blinking, head shaking, and ptyalism, which progressed to partial and generalized seizures. Ancillary diagnostic tests were inconclusive, and the only significant laboratory finding was nonsuppurative pleocytosis of cerebrospinal fluid. Euthanasia was elected. Microscopic evaluation demonstrated multifocal, random nonsuppurative meningoencephalitis involving most prominently the rostral cerebral cortex, as well as the thalamus, midbrain, and rostral medulla. Lesions consisted of inflammation, neuronal necrosis, gliosis, and both neuronal and glial basophilic intranuclear inclusion bodies. Immunohistochemistry with a polyclonal antibody reactive to several equine herpesviruses was positive within affected areas of the brain, and polymerase chain reaction conclusively demonstrated the presence of only equine herpesvirus 9. The clinical and morphologic features of this case resemble other fatal herpesvirus encephalitides derived from interspecies transmission and underscore the need for extreme caution when managing wild or captive equids.
Article
Full-text available
Epidemiological studies have shown that a single‐nucleotide polymorphism in the equid herpesvirus type 1 DNA polymerase gene is associated with outbreaks of highly lethal neurological disease in horses. Reverse genetics experiments further demonstrated that a G2254 A2254 nucleotide mutation introduced in neurovirulent strain Ab4, which resulted in an asparagine for aspartic acid substitution (D752 N752), rendered the virus nonneurovirulent in the equine. Here, we report that the nonneurovirulent strain equid herpesvirus type 1 strain NY03 caused lethal neurological disease in horses after mutation of A2254 G2254 (N752 D752), thereby providing final proof that the D752 allele in the viral DNA polymerase is necessary and sufficient for expression of the lethal neurovirulent phenotype in the natural host. Although virus shedding was comparable between the N752 and D752 variants, infection with the latter was accompanied by efficient establishment of prolonged cell‐associated viremia in peripheral blood mononuclear cells and neurological disease in 2 of 6 animals.
Article
Full-text available
Viruses related to equine herpesvirus type 1 (EHV-1) were isolated from an aborted fetus of an onager (Equus hemionus) in 1984, an aborted fetus of Grevy's zebra (Equus grevyi) in 1984 and a Thomson's gazelle (Gazella thomsoni) with nonsuppurative encephalitis in 1996, all in the USA. The mother of the onager fetus and the gazelle were kept near plains zebras (Equus burchelli). In phylogenetic trees based on the nucleotide sequences of the genes for glycoproteins B (gB), I (gI), and E (gE), and teguments including ORF8 (UL51), ORF15 (UL45), and ORF68 (US2), the onager, Grevy's zebra and gazelle isolates formed a genetic group that was different from several horse EHV-1 isolates. Within this group, the onager and gazelle isolates were closely related, while the Grevy's zebra isolate was distantly related to these two isolates. The epizootiological origin of the viruses is discussed.
Article
Full-text available
Equine herpesvirus 9 was detected in a polar bear with progressive encephalitis; the source was traced to 2 members of a potential equid reservoir species, Grevy's zebras. The virus was also found in an aborted Persian onager. Thus, the natural host range is extended to 6 species in 3 mammalian orders.
Article
Full-text available
Proventricular dilatation disease (PDD) is a fatal disorder threatening domesticated and wild psittacine birds worldwide. It is characterized by lymphoplasmacytic infiltration of the ganglia of the central and peripheral nervous system, leading to central nervous system disorders as well as disordered enteric motility and associated wasting. For almost 40 years, a viral etiology for PDD has been suspected, but to date no candidate etiologic agent has been reproducibly linked to the disease. Analysis of 2 PDD case-control series collected independently on different continents using a pan-viral microarray revealed a bornavirus hybridization signature in 62.5% of the PDD cases (5/8) and none of the controls (0/8). Ultra high throughput sequencing was utilized to recover the complete viral genome sequence from one of the virus-positive PDD cases. This revealed a bornavirus-like genome organization for this agent with a high degree of sequence divergence from all prior bornavirus isolates. We propose the name avian bornavirus (ABV) for this agent. Further specific ABV PCR analysis of an additional set of independently collected PDD cases and controls yielded a significant difference in ABV detection rate among PDD cases (71%, n = 7) compared to controls (0%, n = 14) (P = 0.01; Fisher's Exact Test). Partial sequence analysis of a total of 16 ABV isolates we have now recovered from these and an additional set of cases reveals at least 5 distinct ABV genetic subgroups. These studies clearly demonstrate the existence of an avian reservoir of remarkably diverse bornaviruses and provide a compelling candidate in the search for an etiologic agent of PDD.
Article
Full-text available
The IR6 gene of equine herpesvirus 1 (EHV-1) is a novel gene that maps within each inverted repeat (IR), encodes a potential protein of 272 amino acids, and is expressed as a 1.2-kb RNA whose synthesis begins at very early times (1.5 h) after infection and continues throughout the infection cycle (C. A. Breeden, R. R. Yalamanchili, C.F. Colle, and D.J. O'Callaghan, Virology 191:649-660,1992). To identify the IR6 protein and ascertain its properties, we generated an IR6-specific polyclonal antiserum to a TrpE/IR6 fusion protein containing 129 amino acids (residues 134 to 262) of the IR6 protein. This antiserum immunoprecipitated a 33-kDa protein generated by in vitro translation of mRNA transcribed from a pGEM construct (IR6/pGEM-3Z) that contains the entire IR6 open reading frame. The anti-IR6 antibody also recognized an infected-cell protein of approximately 33 kDa that was expressed as early as 1 to 2 h postinfection and was synthesized throughout the infection cycle. A variety of biochemical analyses including radiolabeling the IR6 protein with oligosaccharide precursors, translation of IR6 mRNA in the presence of canine pancreatic microsomes, radiolabeling the IR6 protein in the presence of tunicamycin, and pulse-chase labeling experiments indicated that the two potential sites for N-linked glycosylation were not used and that the IR6 protein does not enter the secretory pathway. To address the possibility that the unique IR6 gene encodes a novel regulatory protein, we transiently transfected an IR6 expression construct into L-M fibroblasts alone or with an immediate-early gene expression construct along with a representative EHV-1 immediate-early, early, or late promoter-chloramphenicol acetyltransferase reporter construct. The results indicated that the IR6 protein does not affect the expression of these representative promoter constructs. Interestingly, the IR6 protein was shown to be phosphorylated and to associate with purified EHV-1 virions and nucleocapsids. Lastly, immunofluorescence and laser-scanning confocal microscopic analyses revealed that the IR6 protein is distributed throughout the cytoplasm at early times postinfection and that by 4 to 6 h it appears as "dash-shaped" structures that localize to the perinuclear region. At late times after infection (8 to 12 h), these structures assemble around the nucleus, and three-dimensional image analyses reveal that the IR6 protein forms a crown-like structure that surrounds the nucleus as a perinuclear network.
Article
Full-text available
Emerging infectious diseases (EIDs) of free-living wild animals can be classified into three major groups on the basis of key epizootiological criteria: (i) EIDs associated with “spill-over” from domestic animals to wildlife populations living in proximity; (ii) EIDs related directly to human intervention, via host or parasite translocations; and (iii) EIDs with no overt human or domestic animal involvement. These phenomena have two major biological implications: first, many wildlife species are reservoirs of pathogens that threaten domestic animal and human health; second, wildlife EIDs pose a substantial threat to the conservation of global biodiversity.
Article
Full-text available
Infections with viruses of the feline parvovirus subgroup such as feline panleukopenia virus (FPV), mink enteritis virus (MEV) and canine parvovirus (CPV-2) [together with its new antigenic types (CPV-2a, CPV-2b)] have been reported from several wild carnivore species. To examine the susceptibility of different species to the various parvoviruses and their antigenic types, samples from wild carnivores with acute parvovirus infections were collected. Viral DNA was amplified, and subsequently analysed, from faeces or formalin-fixed small intestines from an orphaned bat-eared fox (Otocyon megalotis), a free-ranging honey badger (Mellivora capensis), six captive cheetahs (Acinonyx jubatus), a captive Siberian tiger (Panthera tigris altaica) and a free-ranging African wild cat (Felis lybica). Parvovirus infection in bat-eared fox and honey badger was demonstrated for the first time. FPV-sequences were detected in tissues of the African wild cat and in faeces of one cheetah and the honey badger, whereas CPV-2b sequences were found in five cheetahs and the bat-eared fox. The Siberian tiger (from a German zoo) was infected with a CPV-type 2a virus. This distribution of feline parvovirus antigenic types in captive large cats suggests an interspecies transmission from domestic dogs. CPV-2 sequences were not detected in any of the specimens and no sequences with features intermediate between FPV and CPV were found in any of the animals examined.
Article
Full-text available
A free-ranging lynx (Lynx lynx) was shot because of its abnormal behavior. Histopathological examination revealed a nonsuppurative meningoencephalitis. In situ hybridization, immunohistochemistry, and reverse transcriptase PCR analysis showed the presence of Borna disease virus infection in the brain. To our knowledge, this is the first confirmed case of Borna disease in a large felid.
Article
Full-text available
Microorganisms have been mutating and evolving on Earth for billions of years. Now, a field of research has developed around the idea of using microorganisms to study evolution in action. Controlled and replicated experiments are using viruses, bacteria and yeast to investigate how their genomes and phenotypic properties evolve over hundreds and even thousands of generations. Here, we examine the dynamics of evolutionary adaptation, the genetic bases of adaptation, tradeoffs and the environmental specificity of adaptation, the origin and evolutionary consequences of mutators, and the process of drift decay in very small populations.
Article
Full-text available
MrBayes 3 performs Bayesian phylogenetic analysis combining information from different data partitions or subsets evolving under different stochastic evolutionary models. This allows the user to analyze heterogeneous data sets consisting of different data types—e.g. morphological, nucleotide, and protein—and to explore a wide variety of structured models mixing partition-unique and shared parameters. The program employs MPI to parallelize Metropolis coupling on Macintosh or UNIX clusters. Availability: http://morphbank.ebc.uu.se/mrbayes Contact: fredrik.ronquist@ebc.uu.se * To whom correspondence should be addressed.
Article
Full-text available
Recombination can be a prevailing drive in shaping genome evolution. RAT (Recombination Analysis Tool) is a Java-based tool for investigating recombination events in any number of aligned sequences (protein or DNA) of any length (short viral sequences to full genomes). It is an uncomplicated and intuitive application and allows the user to view only the regions of sequence alignments they are interested in. RAT was applied to viral sequences. Its utility was demonstrated through the detection of a known recombinant of HIV and a detailed analysis of Noroviruses, the most common cause of viral gastroenteritis in humans. RAT, along with a user's guide, is freely available from http://jic-bioinfo.bbsrc.ac.uk/bioinformatics-research/staff/graham_etherington/RAT.htm.
Article
Full-text available
Viral infections of the central nervous system (CNS) are caused by a variety of viruses, namely, herpesviruses, enteroviruses, and flaviviruses. The similar clinical signs provoked by these viruses make the diagnosis difficult. We report on the simultaneous detection of these major CNS pathogens using amplification by PCR and detection of amplified products using DNA microarray technology. Consensus primers were used for the amplification of all members of each genus. Sequences specific for the identification of each virus species were selected from the sequence alignments of each target gene and were synthesized on a high-density microarray. The amplified products were pooled, labeled, and cleaved, followed by hybridization on a single array. This method was successfully used to identify herpesviruses, namely, herpes simplex virus type 1 (HSV-1), HSV-2, and cytomegalovirus; all serotypes of human enteroviruses; and five flaviviruses (West Nile virus, dengue viruses, and Langat virus). This approach, which used highly conserved consensus primers for amplification and specific sequences for identification, would be extremely useful for the detection of variants and would probably help solve some unexplained cases of encephalitis. The analytical sensitivity of the method was shown to be 500 genome equivalents ml−1 for HSV-1, 0.3 50% tissue culture infectious doses (TCID50s) ml−1 for the enterovirus coxsackievirus A9, and 200 TCID50s ml−1 for West Nile virus. The clinical sensitivity of this method must now be evaluated.
Article
Full-text available
Equid herpes virus 1 (EHV-1) related isolates from a captive blackbuck (strain Ro-1) and Grevy's zebra (strain T965) behaved similarly to EHV-1 and EHV-9 in respect to their host cell range. Restriction enzyme analysis and a phylogenetic tree confirmed that Ro-1 and T965 were identical and more closely related to EHV-1 than to EHV-9. Differences from EHV-1 became obvious firstly, by amino acid alignments revealing two unique substitutions in the gB protein of Ro-1 and T965. Secondly, an EHV-1 type-specific monoclonal antibody did not detect its antigen on Ro-1, T965 or EHV-9 infected cells by immunohistochemistry. The results support the view that Ro-1 and T965 isolates represent a distinct, previously unrecognized species of equid herpesviruses.
Article
Full-text available
Equine herpesvirus 1 was isolated from an onager in 1985, a zebra in 1986 and a Thomson's gazelle in 1996 in USA. The genetic relatedness and pathogenicity of these three viruses were investigated based on the nucleotide sequences of the glycoprotein G (gG) gene, experimental infection in hamsters, and comparison with horse isolates. The gG gene sequences of EHV-1 from onager and zebra were identical. The gG gene sequences of the gazelle isolate showed 99.5% identity to those of onager and zebra isolates. The gG gene sequences of EHV-1 isolated from horses were 99.9-100% identical and 98, 98 and 97.8% similar to gG from onager, zebra and gazelle isolates, respectively. Hamsters inoculated with onager, zebra and gazelle isolates had severe weight loss, compared with hamsters inoculated with horse isolates. The histopathological findings were related to the virulence of each isolate. The results indicated that EHV-1 isolates from onager, zebra and gazelle differ from horse EHV-1 and are much more virulent in hamsters.
Article
Full-text available
Infection with equid herpesvirus type 1 (EHV-1) leads to respiratory disease, abortion, and neurologic disorders in horses. Molecular epidemiology studies have demonstrated that a single nucleotide polymorphism resulting in an amino acid variation of the EHV-1 DNA polymerase (N752/D752) is significantly associated with the neuropathogenic potential of naturally occurring strains. To test the hypothesis that this single amino acid exchange by itself influences neuropathogenicity, we generated recombinant viruses with differing polymerase sequences. Here we show that the N752 mutant virus caused no neurologic signs in the natural host, while the D752 virus was able to cause inflammation of the central nervous system and ataxia. Neurologic disease induced by the D752 virus was concomitant with significantly increased levels of viremia (p = 0.01), but the magnitude of virus shedding from the nasal mucosa was similar between the N752 and D752 viruses. Both viruses replicated with similar kinetics in fibroblasts and epithelial cells, but exhibited differences in leukocyte tropism. Last, we observed a significant increase (p < 0.001) in sensitivity of the N752 mutant to aphidicolin, a drug targeting the viral polymerase. Our results demonstrate that a single amino acid variation in a herpesvirus enzyme can influence neuropathogenic potential without having a major effect on virus shedding from infected animals, which is important for horizontal spread in a population. This observation is very interesting from an evolutionary standpoint and is consistent with data indicating that the N752 DNA pol genotype is predominant in the EHV-1 population, suggesting that decreased viral pathogenicity in the natural host might not be at the expense of less efficient inter-individual transmission.
Article
Zoos are ideally placed to act as epidemiological monitoring stations because for decades, many have been building up detailed collections of serum banks, tissue banks and medical record-keeping systems that could be mined for information that would be beneficial to public health. For example, in 1999 wild Crows Corvus brachyrhynchos in the United States of America started to die of unknown causes but it was not until some died in the grounds of a zoological institution that West Nile virus, which is a threat to both human and animal health, was identified. There is a serious disparity in the type and amount of biosurveillance provided for humans, agricultural livestock and wildlife agencies, often driven by economic factors. There is an argument for public-health entities to contribute funds to the cost of managing serum-banks and testing stations within zoos to enhance biosurveillance in urban settings, in a cost-effective and mutually beneficial manner. The key to sustainable and integrated biosurveillance lies in public-health professionals working with zoo professionals, who care for wild animals on a day-to-day basis, to create electronic surveillance networks. This could be of utmost benefit to everyone.
Article
Using the complete haemagglutinin (HA) gene and partial phosphoprotein (P) gene we investigated the genotype of canine distemper virus (CDV) strains recovered from two wildlife species in Mecklenburg-Vorpommern, Germany. Phylogenetic analyses demonstrated significant differences between the strains from raccoons Procyon lotor (family Procyonidae) obtained in 2007 and strains from red foxes Vulpes vulpes (family Canidae) obtained in 2008. The raccoon strains belonged to the CDV European wildlife lineage whereas the red fox strains belonged to the CDV Europe lineage. We combined our genetic sequence data with published data from 138 CDV stains worldwide to investigate the proposed importance of amino acid substitutions in the SLAM binding region of the CDV HA protein at position 530 (G/E to R/D/N) and 549 (Y to H) to the spread of domestic dog-adapted CDV strains to other carnivores. We found no evidence that amino acid 530 was strongly affected by host species. Rather, site 530 was conserved within CDV lineages, regardless of host species. Contrary to expectation, strains from non-dog hosts did not exhibit a bias towards the predicted substitution Y549H. Wild canid hosts were more frequently infected by strains with 549Y, a pattern similar to domestic dogs. Non-canid strains showed no significant bias towards either H or Y at site 549, although there was a trend towards 549H. Significant differences between the prevalence of 549Y and 549H in wild canid strains and non-canid strains suggests a degree of virus adaptation to these categories of host.
Article
In a zoological collection, four black bears (Ursus americanus) died from neurological disease within six months. Independently in a geographically different zoo, two Thomson's gazelles (Eudorcas thomsoni) and 18 guinea pigs (Cavia porcellus f. dom.) suffered from neurological disorders. In addition, guinea pigs showed abortions and stillbirths. All affected animals displayed a non suppurative meningoencephalitis with intranuclear inclusion bodies. Immunohistology demonstrated equine herpes virus antigen and ultrastructurally herpes viral particles were detected. Virus isolation and molecular analysis identified neurotropic equine herpesvirus (EHV) 1 strains in both epizootics. There is serological evidence of a possible virus transmission from other equids to the affected animals. Cross-species transmission of EHV-1 should be considered in the management of captive wild equids and ungulates, particularly with respect to fatal disease in irreplaceable species.
Article
In 2007, disease related mortality occurred in one African wild dog (Lycaon pictus) pack close to the north-eastern boundary of the Serengeti National Park, Tanzania. Histopathological examination of tissues from six animals revealed that the main pathologic changes comprised interstitial pneumonia and suppurative to necrotizing bronchopneumonia. Respiratory epithelial cells contained numerous eosinophilic intracytoplasmic inclusion bodies and multiple syncytial cells were found throughout the parenchymal tissue, both reacting clearly positive with antibodies against canine distemper virus (CDV) antigen. Phylogenetic analysis based on a 388 nucleotide (nt) fragment of the CDV phosphoprotein (P) gene revealed that the pack was infected with a CDV variant most closely related to Tanzanian variants, including those obtained in 1994 during a CDV epidemic in the Serengeti National Park and from captive African wild dogs in the Mkomazi Game Reserve in 2000. Phylogenetic analysis of a 335-nt fragment of the fusion (F) gene confirmed that the pack in 2007 was infected with a variant most closely related to one variant from 1994 during the epidemic in the Serengeti National Park from which a comparable fragment is available. Screening of tissue samples for concurrent infections revealed evidence of canine parvovirus, Streptococcus equi subsp. ruminatorum and Hepatozoon sp. No evidence of infection with Babesia sp. or rabies virus was found. Possible implications of concurrent infections are discussed. This is the first molecular characterisation of CDV in free-ranging African wild dogs and only the third confirmed case of fatal CDV infection in a free-ranging pack.
Article
Blindness characterized by dilated unresponsive pupils and funduscopic evidence of varying degrees of vitritis, retinal vasculitis, retinitis, chorioretinitis, and optic neuritis developed in 21 alpacas and 1 llama within a 30-day period. The animals were part of a group of approximately 100 animals imported from Chile one year earlier. The animals had spent 6 months in quarantine and then, for the 6 months preceding the epizootic, were housed at an exotic animal import-export farm, where the disease developed. Four of the affected animals also had signs of neurologic dysfunction. A herpesvirus indistinguishable from equine herpesvirus I was isolated from 4 of the affected animals, and antibody titers diagnostic for equine herpesvirus I were demonstrated in the serum of all but one of the affected animals.
Article
Pseudorabies virus (PRV) was not transmitted horizontally from 3 PRV-infected calves to 2 contact control calves during 4 days of comingling in experiment 1. Although these contact control calves developed clinical signs of pseudorabies when infected intranasally with PRV in experiment 2, they did not transmit PRV to a second pair of contact control calves. However, 1 of 2 pigs comingled with these 4 calves seroconverted. During both experiments, moderate amounts (10(2) to 10(5) TCID50) of PRV were present in the nasal secretions of the infected calves during the contact periods. All infected calves traumatized their nares or periorbital tissue. Infected calves developed a nonsuppurative meningoencephalitis mainly involving the brain stem. Four of the 5 infected calves had nonsuppurative ganglioneuritis and acute lymphoid necrosis of germinal centers. Virus could not be recovered from nasal and tonsillar swab samples from contact-control calves and pigs.
Article
A simple, sensitive, and specific polymerase chain reaction (PCR) protocol for detection of rabies virus is described. The process consists of sample preparation, reverse transcription, two-step DNA amplification, and detection of the amplified product. RNA was extracted from animal and human brain byphenol-chloroform using guanidinium thiocyanate. Viral RNA wasthen amplified in a two-step PCR that used two sets of nested primers designed to amplify rabies nucleocapsid (N) sequence. Rabies nucleocapsid sequence was amplified from all brain samples from 95 dogs and 3 humans with rabies confirmed by fluorescent antibody (FAT) and mouse inoculation tests (MIT). Rabies-negative brain samples (110 dogs, 2 humans) were PCR-negative. The process requires <24 h. Detection of viralRNA was still possible in brain material that was left at room temperature for 72 h. As little as 8 pg of rabies virus RNA could be detected. This technique could have practical applications as a confirmatory test to FAT at busy rabies diagnostic centers.
Article
A herpesvirus was isolated from Thomson's gazelle (Gazella thomsoni) kept at a zoological garden in Japan during an outbreak of epizootic acute encephalitis. The virus, gazelle herpesvirus 1 (GHV-1), was serologically related to equine herpesvirus 1 (EHV-1). However, DNA fingerprints of GHV-1 were different from those of EHV-1 and other equine herpesviruses. Southern hybridization with probes of cloned BamHI fragments derived from UL and US segments of EHV-1 revealed differences in the DNA restriction profiles throughout the entire genome. Nucleotide sequences were determined for a conserved region of an essential envelope glycoprotein B (gB) gene and a type-specific glycoprotein G (gG) homologue gene. The predicted amino acid sequence of GHV-1 gB showed 97, 92, 61, and 57% identity to EHV-1, EHV-4, feline herpesvirus, and pseudorabies virus, respectively, indicating that GHV-1 was closer to EHV-1 than any other herpesvirus. The GHV-1 gG gene showed 93.2, 92.3, and 53% identity to EHV-1, EHV-8, and EHV-4 gGs, respectively. GHV-1 was virulent to suckling mice of the ICR strain by intracerebral inoculation and was virulent to 4-week-old BALB/c mice by intranasal inoculation, causing neurological symptoms and death. We conclude that GHV-1 is a new type of equine herpesvirus with strong neurotropism.
Article
We have developed a sensitive nested reverse-transcriptase polymerase chain reaction assay (n RT-PCR) for the detection of the tick-borne encephalitis virus (TBEV) RNA, especially in ticks. The primer pairs were selected from the 5'-terminal noncoding region, a highly conserved part of the virus. The specificity was tested by computer homology searches of sequences as well as by the sequencing of the first and second amplificate, by Southern blot hybridization with a DIG-labelled oligonucleotide probe, and by restriction enzyme analysis. The method has proved to be very sensitive. The detection limit is about 20 fg of TBEV RNA per PCR run (25 microliters), or a single positive tick, i.e. (adult or nymph). The method can be used for comparative studies of the epidemiological situation, as well as for the screening of natural foci for the presence and circulation of TBEV or for the detection of TBEV-genome-sequences in clinical materials.