More than 100 young captive and wild Asian elephants are known to have died from a rapid-onset, acute hemorrhagic disease caused primarily by multiple distinct strains of two closely related chimeric variants of a novel herpesvirus species designated elephant endotheliotropic herpesvirus (EEHV1A and EEHV1B). These and two other species of Probosciviruses (EEHV4 and EEHV5) are evidently ancient and likely nearly ubiquitous asymptomatic infections of adult Asian elephants worldwide that are occasionally shed in trunk wash secretions. Although only a handful of similar cases have been observed in African elephants, they also have proved to harbor their own multiple and distinct species of Probosciviruses-EEHV2, EEHV3, EEHV6, and EEHV7-found in lung and skin nodules or saliva. For reasons that are not yet understood, approximately 20% of Asian elephant calves appear to be susceptible to the disease when primary infections are not controlled by normal innate cellular and humoral immune responses. Sensitive specific polymerase chain reaction (PCR) DNA blood tests have been developed, routine monitoring has been established, the complete large DNA genomes of each of the four Asian EEHV species have now been sequenced, and PCR gene subtyping has provided unambiguous evidence that this is a sporadic rather than epidemic disease that it is not being spread among zoos or other elephant housing facilities. Nevertheless, researchers have not yet been able to propagate EEHV in cell culture, determine whether or not human antiherpesvirus drugs are effective inhibitors, or develop serology assays that can distinguish between antibodies against the multiple different EEHV species.

Systemic infections with Elephant Endotheliotropic Herpesviruses (EEHV) cause a rapid onset acute hemorrhagic disease with an 85% mortality rate. More than 60 cases have been confirmed worldwide occurring predominantly in juvenile Asian elephants. Originally, three virus types EEHV1A, EEHV1B and EEHV2 were identified, all members of the Proboscivirus genus within the Betaherpesvirinae. However, four elephant gammaherpesviruses (EGHV) have also been found by DNA PCR approaches in eye and genital secretions of asymptomatic animals, and two more versions of the Probosciviruses, EEHV3 and EEHV4, were recently detected in acute hemorrhagic disease cases. To ask whether even more species of elephant herpesviruses may exist, we have developed several new diagnostic DNA PCR assays using multiple round primers in the DNA POL region. These have been used routinely for nearly three years to screen samples submitted to the Elephant Herpesvirus Laboratory for diagnosis of possible cases of EEHV disease in blood and necropsy tissue, as well as in biopsies of other suspicious lesions or growths. Several more cases of EEHV1-associated hemorrhagic disease were confirmed, but in addition, we describe here eleven examples of other known and novel herpesviruses detected and evaluated with these reagents. They include the prototypes of four new elephant herpesviruses, two more within the Proboscivirus group EEHV5 and EEHV6, plus two more gammaherpesviruses EGHV3B and EGHV5. We also report initial semi-quantitative PCR assays demonstrating very high viral loads in the blood of the EEHV3 and EEHV4-associated hemorrhagic disease cases.

Objective To investigate the pathogenesis and transmission of elephant endotheliotropic herpesvirus (EEHV1) by analyzing various elephant fluid samples with a novel EEHV1-specific real-time PCR assay. Animals 5 apparently healthy captive Asian elephants (Elephas maximus) from the same herd. Procedures A real-time PCR assay was developed that specifically detects EEHV1. The assay was used to evaluate paired whole blood and trunk-wash samples obtained from the 5 elephants during a 15-week period. Deoxyribonucleic acid sequencing and viral gene subtyping analysis were performed on trunk-wash DNA preparations that had positive results for EEHV1. Viral gene subtypes were compared with those associated with past fatal cases of herpesvirus-associated disease within the herd. Results The PCR assay detected viral DNA to a level of 1,200 copies/mL of whole blood. It was used to detect EEHV1 in trunk secretions of 3 of the 5 elephants surveyed during the 15-week period. Viral gene subtyping analysis identified 2 distinct elephant herpesviruses, 1 of which was identical to the virus associated with a previous fatal case of herpesvirus-associated disease within the herd. Conclusions and Clinical Relevance EEHV1 was shed in the trunk secretions of healthy Asian elephants. Trunk secretions may provide a mode of transmission for this virus. Results of this study may be useful for the diagnosis, treatment, and management of EEHV1-associated disease and the overall management of captive elephant populations.

The first herpesviruses described in association with serious elephant disease were referred to as endotheliotropic herpesviruses (EEHV) because of their ability to infect capillary endothelial cells and cause potentially fatal disease. Two related viruses, EEHV1 and EEHV2, have been described based on genetic composition. This report describes the similarities and differences in clinicopathologic features of 2 cases of fatal endotheliotropic herpesvirus infections in Asian elephants caused by a previously unrecognized virus within the betaherpesvirus subfamily that is markedly divergent from the 2 previously studied fatal probosciviruses, based on polymerase chain reaction sequence analysis of 2 segments of the viral genome. In addition to ascites, widespread visceral edema, petechiae, and capillary damage previously reported, important additional findings with EEHV3 infection were the presence of grossly visible renal medullary hemorrhage, a tropism for larger veins and arteries in various tissues, relatively high density of renal herpetic inclusions, and involvement of the retinal vessels. These findings indicate a less selective organ tropism, and this may confer a higher degree of virulence for EEHV3.

Up to 65% of deaths of young Asian elephants (Elephas maximus) between 3 mo and 15 yr of age in Europe and North America over the past 20 yr have been attributed to hemorrhagic disease associated with a novel DNA virus called elephant endotheliotropic herpesvirus (EEHV). To evaluate the potential role of EEHV in suspected cases of a similar lethal acute hemorrhagic disease occurring in southern India, we studied pathologic tissue samples collected from field necropsies. Nine cases among both orphaned camp and wild Asian elephants were identified by diagnostic PCR. These were subjected to detailed gene subtype DNA sequencing at multiple PCR loci, which revealed seven distinct strains of EEHV1A and one of EEHV1B. Two orphan calves that died within 3 days of one another at the same training camp had identical EEHV1A DNA sequences, indicating a common epidemiologic source. However, the high level of EEHV1 subtype genetic diversity found among the other Indian strains matches that among over 30 EEHV1 strains that have been evaluated from Europe and North America. These results argue against the previous suggestions that this is just a disease of captive elephants and that the EEHV1 virus has crossed recently from African elephant (Loxodonta africana) hosts to Asian elephants. Instead, both the virus and the disease are evidently widespread in Asia and, despite the disease severity, Asian elephants appear to be the ancient endogenous hosts of both EEHV1A and EEHV1B.