key: cord-0041749-2hahwnq4
authors: Evermann, James F.; Laurenson, M. Karen; McKeirnan, Alison J.; Caro, T. M.
title: Infectious disease surveillance in captive and free‐living cheetahs: An integral part of the species survival plan
date: 2005-05-23
journal: Zoo Biol
DOI: 10.1002/zoo.1430120111
sha: 2a97d66be7ba692691e9adbac10df469c2e99979
doc_id: 41749
cord_uid: 2hahwnq4

During the formulative stages of developing the Species Survival Plan (SSP) for the cheetah, the impact of infectious disease upon its survival in captivity was of prime consideration, together with genetics, nutrition, physiology, and behavior. This paper summarizes the results of an infectious disease surveillance program, initially designed to monitor the infectious agents associated with clinically normal and clinically ill cheetahs in captivity, but subsequently supplemented with data from free‐living cheetahs. The focus was on two viral infections, feline infectious peritonitis (FIP) and feline rhinotracheitis virus. Results indicated that between 1989 and 1991, there was an increase in the seroprevalence (number antibody‐positive animals) of cheetahs to feline coronavirus from 41% to 64% in captivity. During this same time period, there were only two documented cases of FIP in cheetahs in the United States. The results suggest that feline coronavirus (feline enteric coronavirus‐‐feline infectious peritonitis group) or a closely related coronavirus of cheetahs is becoming endemic in the captive cheetah population. Further serologic results from 39 free‐living cheetahs demonstrated that there was a high seroprevalence (61%) to feline coronavirus, although serum antibody titers were considerably lower than those encountered in captive cheetahs. The observation of a high percentage of free‐living cheetahs, which were seropositive to feline herpesvirus (44%), was unexpected, since it has been generally regarded that this infection is primarily associated with cheetahs in captivity. © 1993 Wiley‐Liss, Inc.

Cheetahs are endangered in the wild as a result of increasing human agricultural encroachment on their habitat and direct persecution of them and their prey. In order to provide a buffer to dwindling wild populations, a concerted effort has been launched by North American zoos to keep a self-sustaining population in captivity. Unfortunately, low reproductive rates and continued depletion from adult mortality has made this goal difficult to attain at present. However, in November, 1989 , the Cheetah Species Survival Plan (SSP) was started to investigate problems of maintaining a viable population in captivity [Grisham and Lindburg, 1989; Marker and O'Brien, 19891. One protocol was to monitor the health of captive cheetahs and to sample them for the occurrence of infectious microorganisms which may have a negative impact upon their survival in captivity. During the initial phase of the disease surveillance study, serum samples, together with swabs and/or tissues from normal and diseased cheetahs, were solicited for viral antibody determination and viral culture.

The purposes of this paper are to bring together the serological results compiled over the past three years and to present perspectives on the ecology of infectious agents in the cheetah's environment. Although emphasis will be placed upon two viral infections, feline infectious peritonitis (FIP) and feline rhinotracheitis, other infectious agents may also have a detrimental effect upon the survival of the cheetah in captivity [ Barr et al., 1989; Baxby et al., 1982; Letcher and O'Conner, 1991; Sabine and Hyne, 19701 .

Serum samples were collected from captive and wild-caught cheetahs as previously reported [Evermann et al., 1988; Heeney et al., 19901 . The serum was separated from the blood clot, and stored at -20°C until used in serologic assays.

The indirect immunofluorescent antibody (LFA) serologic assay was used to evaluate cheetah serum for antibodies to the feline coronavirus group (feline infectious peritonitis/feline enteric coronavirusicanine coronavirusitransmissible gastroenteritis virus). The IFA had been shown to correlate well with western blot serology comparing coronavirus antibody titers in domestic cats and in cheetahs [Heeney et al., 19901 . IFA antibody titers L 1:25 were regarded as positive. The virus neutralization (VN) assay was used to determine antibody titers to feline herpesvirus (FHV) type 1. VN antibody titers 2 1:2 were regarded as positive. Earlier work had demonstrated that FHV was closely related to the herpesvirus isolated from cheetah cubs with advanced facial cutaneous ulcers [Junge et al., 1991; Scherba et al., 19883 , and from adult cheetahs succumbing to diseases heretofore unrelated to herpesvirus infection [Evermann and McKeirnan, 19911. 

The SSP surveillance of feline coronavirus infection in captive cheetahs commenced in 1989. Table 1 presents a summary of the seroreactivity of captive cheetahs to the feline coronavirus group. In 1989, there were 48 accessions which amounted to 102 samples tested, 42 (41.2%) of which were seropositive. In 1990, there were more accessions for a total of 164 serum samples tested, of which 106 (64.6%) were seropositive. In the third year of the study, 1991, there were 81 accessions and 194 serum samples tested. Of these, 125 (64.4%) were seropositive to the feline coronavirus group.

These results were analyzed further by comparing the distribution of serum antibody titers amongst the three years (Table 2 ). There were three categories established based upon the serum titer levels. These were seronegative (IFA titers < 1:25); seropositive, normal range (IFA titers 1:25 to 1:3,125); and seropositive, "watch" (IFA titers 2 1:15,625). It was observed that there was a change in serum antibody titer distribution from the first year (1989) of the study compared with the succeeding two years. The number of seronegative cheetahs dropped from 58.8% in 1989 to 35.9% in 1990. This percentage remained constant throughout 1991. Concurrent with the decrease in seronegative cheetahs was the increase in seropositive cheetahs in the normal serum titer range (35.2% in 1989 to 59.7% in 1990 and 62.8% in 1991) . The number of cheetahs in the "watch" category decreased from 6 in 1989 to 3 in 1991. During this period of time, there were 2 deaths attributed to FIP; both animals had serum titers in the "watch" category.

In a parallel study of free-living cheetahs, immobilized and sampled in the course of radio collaring [Caro, in press; Laurenson and Caro, in press] , samples were collected from 39 cheetahs. Sixty-one percent were seropositive to feline coronavirus ( Table 3) .

Only free-living cheetahs were included in this portion of the study, since the majority of captive cheetahs in the United States are vaccinated with combination vaccines containing feline herpesvirus, feline calicivirus , and feline panleukopenia virus [Wack, 19911 . The results (Table 3) indicated that there was a high rate of seropositivity to feline herpesvirus (43.6%).

The surveillance of infectious disease microorganisms affecting the cheetah has focused upon just two agents, on the basis of prior disease epizootics with FIP and the isolation of feline herpesviruses from several cases during the course of the current study [Evermann et al., 1988 [Evermann et al., , 1989 Evermann and McKeirnan, 1991; Junge et al., 19911 . The results of the coronavirus serology reflected a trend toward a more endemic infection occurring in the captive cheetah populations of the United States. Infection rates of 64.4% are similar to those observed in some domestic cats housed in catteries (40-85%) [Pedersen, 19911. During the course of this study there were only two cases of FIP reported in the cheetah population, which may indicate one of several things. This low level of FIP mortality may be indicative of the control efforts that have been used in various cheetah collections (based upon serologic-directed quarantine); the changing virulence of the feline-cheetah coronavirus in nature; the lowered number of coronavirus-naive cheetahs in captivity; or a combination of the above.

The earlier findings of feline herpesvirus infection being associated with clinically ill as well as clinically normal cheetahs is compatible with our knowledge of the pathogenesis of this virus in domestic cat populations [Povey, 19861 . The isolation of feline herpesvirus from a clinically normal cheetah confirms that asymptomatic shedding can occur and may account for the infection of susceptible animals, especially cubs [Evermann and McKeirnan, 19911. The serologic profile of the 39 free-living cheetahs points to several interesting observations. First, it exemplifies the use of serologic testing for surveillance purposes to assess the degree of infection within a given population of animals [Hancock, 1988; Heeney et al., 1990; Horzinek and Osterhaus, 1979; Munson, 19911 . Second, the high seroprevalence of feline coronavirus antibodies (61.5%) indicates that a coronavirus of cheetahs is endemic in at least one cheetah population living in East Africa. This observation supports the earlier reports of coronavirus seropositive cheetahs in the wild [Evermann et al., 1988; Horzinek and Osterhaus, 19791 . The fact that certain populations of cheetahs, either in captivity or in the wild, have remained seronegative to feline coronavirus may be reflective of the lack of exposure to a cheetah coronavirus or to the other coronaviruses known to cross-react with the feline coronaviruses, i.e., porcine coronavirus and canine coronavirus [Spencer, 19911. The third observation indicates a high seroprevalence of free-living cheetahs to feline herpesvirus (43.6%), previously considered to be predominantly an infection and disease of captive cheetahs. The occurrence of seroreactive cheetahs in the wild raises many questions in regards to the ecology of these viruses and the practicality of trying to maintain colonies of cheetahs that are free of infection. It is clear that more needs to be known about the ecology of these viruses in nature in order to implement control measures for their control in captive cheetah populations [Morse, 19911 .

Anderson [ 19911 recently posed the question pertaining to populations and infectious diseases of whether one should refer to it as ecology or epidemiology. In reality, it is both, since there is considerable overlap. When studying the ecology of a virus, the questions asked are how does the virus persist in nature at the cellular level, host animal level, and at the population level [Anderson and May, 1986; Mahy, 198.51 . Pathogenesis refers to the potential for the virus to cause disease at the host animal level, which invariably involves target organs and preferential cells for virus replication. Epidemiology is the study of the ecology of an infectious agent in order to understand the pathogenesis, and ultimately control it. Control may occur by culling of infected animals, segregation to reduce infection, and thereby minimize disease, and vaccination if one exists. Table 4 presents the ecology of feline infectious peritonitis virus and feline herpesvirus, based upon current information.

The control of feline coronavirus infections in domestic cats and cheetahs has historically relied upon initial serologic testing, segregation, and periodic serologic testing to monitor the effectiveness of the control program [Addie and Jarret, 1990; Evermann et al., 1988 Evermann et al., , 1991 Hoskins, 1991; Pedersen, 19911 . The placement of domestic cats in small family units of five to six animals has also been an effective control measure for control of FIP in endemic catteries [Evermann and Ott, unpublished data] . The apparent increase in seroprevalence of captive cheetahs from 1989 to 1991 suggests that segregation may not be totally effective in reducing infection. However, there has been a noticeable reduction in the clinical cases of FIP, which may reflect a form of natural immunization with a feline-cheetah coronavirus, as the infection becomes more endemic in captive cheetahs.

The control of FIP in domestic cats has been augmented by the recent development of a modified-live temperature-sensitive FIP virus vaccine [Postorino et al., 19921 . The vaccine appears to be safe, and the effectiveness ranges from 40 to 80% depending on the strain of FIP virus that is used to challenge the cats. The vaccine has not yet been tested for safety in cheetahs. The efficacy of such a vaccine should only be studied in a collection of captive cheetahs that is endemic for feline coronavirus, due to the hazards of experimental challenge in an endangered species [Evermann and McKeirnan, 19911. The control of feline herpesvirus infections has been dependent upon a combination of good hygiene, decreased population density, and an aggressive vaccination program (at least twice per year) [Evermann and McKeirnan, 1991; Povey, 1986; Wack, 19911 . The feline herpesvirus is usually shed when the cats have clinical signs, such as rhinitis and conjunctivitis [Povey, 19861 . The pathogenesis of feline herpes-virus in cheetahs appears to be similar to that described in domestic cats [Evermann and McKeirnan, 19911 . The immune response to the feline herpesvirus is dependent upon maternal antibodies for the first 8 to 12 weeks, and then for continual boosting from either natural exposure or vaccination to maintain protective levels of immunity [Spencer and Burroughs, 1991, 1992; Wack, 19911 .

The experiences observed with infectious diseases of cheetahs, in particular the feline coronavirus and feline herpesvirus infections, raises some questions pertaining to the factors which influence the susceptibility of these animals to infection and the severity of disease in selected individuals. According to Pedersen [ 199 11 , there are many factors which influence the outcome of infections. These include: the host response; the environment in which the host lives; and microbial agents to which the host is exposed. Of particular interest with regard to the cheetah are those factors of the host response, primarily those pertaining to developmental and heritable anomalies of the immune system. O'Brien and Evermann [1988] reviewed reports on the genetic homogeneity of the cheetah and postulated that this homozygosity may reflect lack of immunologic diversity to infectious agents. In a series of experiments, the immune response of the cheetah has been reported as being suboptimal, based on the lack of rejection of skin grafts and the lack of demonstrable neutralizing antibodies to FIP virus [McKeirnan and Evermann, unpublished data; O'Brien et al., 19851. Pedersen [1987] cautions against the use of skin testing for measuring the effectiveness of delayed-type hypersensitivity reactions in domestic cats. Although cats do not respond well to some antigens, they have been shown to develop measurable levels of delayed hypersensitivity to certain chemicals. Nonetheless, there appears to be some differences in the cell-mediated immune response of domestic cats as measured by decreased lymphocyte reactivity to T-cell mitogens, and this observation may apply to cheetahs as well. Recent evidence now supports the altered T-cell function of the cheetah [Miller-Edge and Worley, 19921 . In that study, there was a decrease in T-cell responsiveness of cheetah peripheral blood mononuclear cells, when compared with domestic cats, when the cells were cultured with inactivated feline herpesvirus. These experiments, as well as those of other investigators, suggest that the susceptibility of the cheetah to viral infections and the severity of disease are the result of an immune-compromised host [Miller-Edge and Worley, 19911 . This understanding will allow for us to pursue strategies for augmenting the cheetah's immune response, which should include genetics as well as managing cheetahs to reduce the risk of high exposure to potentially fatal pathogens, and the development of effective and safe vaccines to aid in the control of such diseases [Hohdatsu et al., 1991; Lin, 19921 .

We express our gratitude to Dr. Melody Roelke, who saw a disease situation, and not only pursued a diagnosis, but advocated corrective management strategies as a means of long-term prevention. We would also like to thank those investigators who have shared their research findings with us, in particular Dr. Michael Worley and Dr. Ray Wack. We greatly appreciate those zoo veterinarians, curators, and staff who have cooperated in and assisted with the collection of samples from the cheetahs. Continued thanks to Dr. Tony Gallina for his support of the diagnosis of animal diseases. We extend our appreciation to the Government of Tanzania, Serengeti Wildlife Research Institute, and Tanzania National Parks for permission to conduct research. Biological material was collected and transported in full compliance with federal fish and wildlife permits (CITES: endangered and threatened species) issued to the National Cancer Institute by the U.S. Fish and Wildlife Service of the Department of the Interior.

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