James E. Childs

Bats: Important Reservoir Hosts of Emerging Viruses

SUMMARY Bats (order Chiroptera, suborders Megachiroptera [“flying foxes”] and Microchiroptera) are abundant, diverse, and geographically widespread. These mammals provide us with resources, but their importance is minimized and many of their populations and species are at risk, even threatened or endangered. Some of their characteristics (food choices, colonial or solitary nature, population structure, ability to fly, seasonal migration and daily movement patterns, torpor and hibernation, life span, roosting behaviors, ability to echolocate, virus susceptibility) make them exquisitely suitable hosts of viruses and other disease agents. Bats of certain species are well recognized as being capable of transmitting rabies virus, but recent observations of outbreaks and epidemics of newly recognized human and livestock diseases caused by viruses transmitted by various megachiropteran and microchiropteran bats have drawn attention anew to these remarkable mammals. This paper summarizes information regarding chiropteran characteristics and information regarding 66 viruses that have been isolated from bats. From these summaries, it is clear that we do not know enough about bat biology; we are doing too little in terms of bat conservation; and there remain a multitude of questions regarding the role of bats in disease emergence.

Ehrlichia chaffeensis: a Prototypical Emerging Pathogen

SUMMARY Ehrlichia chaffeensis is an obligately intracellular, tick-transmitted bacterium that is maintained in nature in a cycle involving at least one and perhaps several vertebrate reservoir hosts. The moderate to severe disease caused by E. chaffeensis in humans, first identified in 1986 and reported for more than 1,000 patients through 2000, represents a prototypical “emerging infection.” Knowledge of the biology and natural history of E. chaffeensis, and of the epidemiology, clinical features, and laboratory diagnosis of the zoonotic disease it causes (commonly referred to as human monocytic ehrlichiosis [HME]) has expanded considerably in the period since its discovery. In this review, we summarize briefly the current understanding of the microbiology, pathogenesis, and clinical manifestations associated with this pathogen but focus primarily on discussing various ecological factors responsible for the recent recognition of this important and potentially life-threatening tick-borne disease. Perhaps the most pivotal element in the emergence of HME has been the staggering increases in white-tailed deer populations in the eastern United States during the 20th century. This animal serves as a keystone host for all life stages of the principal tick vector (Amblyomma americanum) and is perhaps the most important vertebrate reservoir host for E. chaffeensis. The contributions of other components, including expansion of susceptible human populations, growth and broadening geographical distributions of other potential reservoir species and A. americanum, and improvements in confirmatory diagnostic methods, are also explored.

Predicting the spatial dynamics of rabies epidemics on heterogeneous landscapes

Often as an epidemic spreads, the leading front is irregular, reflecting spatial variation in local transmission rates. We developed a methodology for quantifying spatial variation in rates of disease spread across heterogeneous landscapes. Based on data for epidemic raccoon rabies in Connecticut, we developed a stochastic spatial model of rabies spread through the states 169 townships. We quantified spatial variation in transmission rates associated with human demography and key habitat features. We found that large rivers act as semipermeable barriers, leading to a 7-fold reduction in the local rates of propagation. By combining the spatial distribution of major rivers with long-distance dispersal we were able to account for the observed irregular pattern of disease spread across the state without recourse to direct assessment of host-pathogen populations.

Long-term studies of hantavirus reservoir populations in the southwestern United States: a synthesis.

A series of intensive, longitudinal, mark-recapture studies of hantavirus infection dynamics in reservoir populations in the southwestern United States indicates consistent patterns as well as important differences among sites and host-virus associations. All studies found a higher prevalence of infection in older (particularly male) mice; one study associated wounds with seropositivity. These findings are consistent with horizontal transmission and transmission through fighting between adult male rodents. Despite very low rodent densities at some sites, low-level hantavirus infection continued, perhaps because of persistent infection in a few long-lived rodents or periodic reintroduction of virus from neighboring populations. Prevalence of hantavirus antibody showed seasonal and multiyear patterns that suggested a delayed density-dependent relationship between prevalence and population density. Clear differences in population dynamics and patterns of infection among sites, sampling periods, and host species underscore the importance of replication and continuity of long-term reservoir studies. Nevertheless, the measurable associations between environmental variables, reservoir population density, rates of virus transmission, and prevalence of infection in host populations may improve our capacity to model processes influencing infection and predict increased risk for hantavirus transmission to humans.

Epidemiology of Human Rabies in the United States, 1980 to 1996

One of the oldest recognized zoonotic diseases, rabies continues to plague humankind and causes more than 35 000 deaths annually [1]. These potentially preventable deaths occur primarily in Asia, Africa, and Latin America, where animal control, vaccination programs, and effective human postexposure prophylaxis are not widely available. In contrast, in the United States, deaths in humans caused by rabies totaled 99 in the 1950s, 15 in the 1960s, 23 in the 1970s, 10 in the 1980s, and 22 from 1990 through 1996 [2, 3]. The epidemiology of human rabies is ultimately linked to cycles of rabies virus transmission in animals. With the interruption of dog-to-dog transmission in most regions, the incidence of human rabies in the United States has reached a level that cannot be further reduced without targeting wildlife. An understanding of epidemiologic patterns of rabies virus maintenance in natural populations has emerged in the past 20 years, largely because of advances in immunology and molecular biology. Monoclonal antibody and genetic sequence analyses of rabies virus variants permit detailed descriptions of enzootic maintenance cycles of specific virus variants in the United States [4, 5]. These analyses have led to an understanding of how variants of rabies virus are maintained in natural reservoirs within geographic regions and have provided information on variability of the virus itself. Current epidemiologic patterns of rabies in the United States can be summarized as follows: The annual reports of rabies in wildlife exceed those of rabies in domestic animals [6]; rabies variants in bats are associated with a disproportionate number of infections in humans, although bats constitute only about 10% of all reported rabies cases in animals annually; most other cases of human rabies diagnosed in the United States can be attributed to infections acquired in areas of enzootic canine rabies outside of the United States; most persons with a case of rabies that originated in the United States have no history of an animal bite; and rabies is diagnosed after death in more than one third of the latter group. The last published summary of cases of human rabies in the United States covered the period from 1960 to 1979 [3]. This review discusses the clinical and epidemiologic features of cases of human rabies in the United States from 1980 to 1996. Methods Case Definition This report includes all laboratory-confirmed cases of human rabies in the United States or its territories from 1980 to 1996 [7-31]. All of the cases were reported to the Centers for Disease Control and Prevention (CDC) by health authorities as part of ongoing national surveillance. Variable Definitions Onset of illness was defined as either the first day of reported symptoms attributable to rabies or the date of initial presentation for medical care before confirmation of rabies. Clinical signs attributable to rabies included paresthesia, anxiety, agitation, confusion, disorientation, hydrophobia, aerophobia, hypersalivation, dysphagia, paresis, paralysis, and fluctuating levels of consciousness [32, 33]. The type of transmitting animal and the geographic location of exposure were listed if the case history included a definite animal bite. The reliability of information that linked rabies exposure to a human was assessed by subsequent laboratory typing of the rabies virus variant. All other exposures were defined as unknown. The diagnosis of rabies was considered antemortem if it was tentatively made and samples were obtained specifically for rabies testing before the patients death. Laboratory Tests The diagnosis of rabies was confirmed by using standard tests [34] conducted at the CDC or at a state laboratory. Serology Two tests were used to detect rabies antibody: the rapid fluorescent focus inhibition test and the indirect immunofluorescence assay. The rapid fluorescent focus inhibition test measures neutralizing antibody. An antibody titer of 1:5 or more, as defined by the reciprocal of the serum or cerebrospinal fluid dilution that reduces the challenge virus by 50%, was considered positive. An indirect immunofluorescence assay, using patient serum or cerebrospinal fluid diluted 1:4 or more, detects serum reactive with rabies antigen in infected cell cultures. The presence of antibody in serum was considered diagnostic if no vaccine or antirabies serum was given to the patient. Antibody in the cerebrospinal fluid, regardless of the rabies immunization history, was considered indicative of rabies virus infection. Virus Isolation Suspensions of brain or saliva specimens were added to mouse neuroblastoma cells and cultured for 24 and 48 hours. Culture slides were fixed and examined by direct immunofluorescence assay for antigen. Samples that were initially negative were maintained for an additional 3 to 4 days and retested. The negative result was considered definitive if it occurred both times. Antigen Detection Antigen detection was performed by direct immunofluorescence of assay serial frozen sections of nuchal skin biopsy specimens, touch impressions of corneal epithelial cells, or fresh brain matter. Paraffin-embedded fixed brain matter was sectioned and enzyme-digested before direct immunofluorescence. RNA Detection Standard extraction procedures and reagents were used to obtain nucleic acids from samples of undiluted saliva; from fresh or paraffin-embedded fixed samples of the brain; or, occasionally, from other tissues. Reverse transcription of RNA and complementary DNA amplification were performed by polymerase chain reaction (PCR) with primers derived from the sequence of the N protein gene. The nucleotide sequence of all PCR products was obtained by standard dideoxynucleotide sequencing methods. Rabies virus variants were identified by comparing samples of rabies virus obtained from all known reservoirs for rabies in the United States [5] with samples of rabies virus obtained from dogs in Asia, Africa, and Latin America [35]. Statistical Analysis Data analyses were performed by using EPI INFO 6 (Centers for Disease Control and Prevention, Atlanta, Georgia) or SPSS 6.0 for Windows (SPSS Inc., Chicago, Illinois) [36, 37]. Specific tests are identified in the text. Some variables were dichotomized before statistical comparisons for determination of odds ratios and 95% CIs. All reported P values are for two-tailed tests of significance. Results Demographic Information Thirty-two persons died of rabies in the United States from 1980 through 1996. Patients ranged in age from 4 to 82 years (median, 27 years) and 20 (63%) were male (Table 1). Cases were reported from 20 states; 7 cases (22%) were reported in California and 6 in Texas. Eleven patients were exposed to rabies in eight foreign countries on the basis of variant typing. The onset of illness occurred in all months and had no apparent seasonal pattern. Dates of exposure, based on the history of an animal bite, were obtained for 7 patients (22%) (Table 1). Table 1. Human Rabies in the United States, 1980-1996 Exposure History A definite history of animal exposure was identified in 7 of the 32 patients (22%), and 25 remained unknown or indefinite (Table 1). Of the 7 cases of definite exposure, 6 resulted from a dog bite received in a foreign country and 1 was from a bat bite received in the United States. Although rabies was not diagnosed in any of the animals that inflicted bites, in each case the rabies virus variant identified in the human sample was consistent with that in the animal species implicated as the source of infection (Table 1). Contact with an animal, thereby suggesting the source for infection, was identified in 12 persons (8 with a bat, 2 with a dog, 1 with a cow, and 1 with a cat). This human-animal contact, however, could not be linked to a bite or mucous membrane contact with the saliva of an animal potentially infected with rabies virus. The remaining 13 patients did not report animal contact; thus, a potential source of exposure was not identified. Histories were obtained before death from friends or relatives in 9 cases and from 4 children aged 11 to 13 years. Prophylaxis None of the 32 patients received a complete series of rabies prophylaxis after exposure; patient 7 reported receiving a single injection of an unknown type after a dog bite in Guatemala, and patients 15, 29, and 30 received human rabies immune globulin during the course of their clinical illness. Clinical Presentation For the 7 patients in which a definite animal bite occurred, the median incubation period was 85 days (range, 53 to 150 days). The first signs and symptoms of rabies were often nonspecific, including fever, sore throat, chills, malaise, anorexia, headache, nausea, vomiting, dyspnea, cough, and weakness. Specific symptoms, such as paresthesias at or near the presumed exposure site, were also reported early in the clinical course, and 19 of the 32 patients (59%) had three or more clinical findings suggestive of rabies during the course of their illness (Table 2). The 32 patients were seen by physicians on an outpatient basis a median of one time (range, 0 to 5 times) before hospitalization, and the median length of time from the onset of illness attributable to rabies to hospitalization was 4 days (range, 1 to 10 days). Table 2. Clinical Findings Suggestive of Rabies in 32 Patients* On admission, 21 of the 32 patients (66%) were febrile (oral temperature > 37.8C), including 12 patients with temperatures greater than 39.5C. Of the 11 patients who were afebrile on admission, 5 reported being febrile before admission, 2 became febrile within 2 days of admission, and 4 had no additional temperatures recorded. The antemortem diagnosis of rabies was first considered at the time of hospitalization in 5 patients, within 1 day of hospitalization in 5 patients, and after a median of 6 days of hospitalization (range, 2 to 12 days) in 10 patients. In 12 patients, rabies was diagnosed after death. Th

Guidelines for Working with Rodents Potentially Infected with Hantavirus

Because of the high morbidity and mortality associated with hantavirus pulmonary syndrome and the possibility of aerosol transmission of hantaviruses, persons handling known reservoir species in the field, laboratory, or classroom should take special precautions to minimize the risk of infection. We provide specific guidelines for personal safety while trapping, handling and releasing, transporting, sampling, and performing necropsy on potentially infected rodents or teaching field classes in areas occupied by reservoir species. Special consideration should be given to respiratory protection, choice and use of disinfectants, decontamination of instruments and traps, proper disposal of infectious wastes, and preservation and shipment of samples intended for hantavirus testing. Precautionary testing of wild rodents used to start laboratory colonies is recommended. Although we specifically address hantaviruses, the procedures described are applicable for any study of populations of small mammals when an infectious zoonotic agent transmissible by aerosol and capable of causing high morbidity and mortality is involved.

Q Fever in Humans and Animals in the United States

Coxiella burnetii, the etiologic agent of Q fever, is a worldwide zoonotic pathogen. Although Q fever is present in the United States, little is known about its current incidence or geographic distribution in either humans or animals. Published reports of national disease surveillance, individual cases, outbreak investigations, and serologic surveys were reviewed to better characterize Q fever epidemiology in the United States. In national disease surveillance reports for 1948-1986, 1,396 human cases were reported from almost every state. Among published individual case reports and outbreak investigations, occupational exposures (research facilities, farm environments, slaughterhouses) were commonly reported, and sheep were most frequently implicated as a possible source of infection. In studies conducted on specific groups, livestock handlers had a significantly higher prevalence of antibodies to C. burnetii than did persons with no known risk. Animal studies showed wide variation in seroprevalence, with goats having a significantly higher average seroprevalence (41.6%) than sheep (16.5%) or cattle (3.4%). Evidence of antibody to C. burnetii was reported among various wild-animal species, including coyotes, foxes, rodents, skunks, raccoons, rabbits, deer, and birds. This literature review suggests that C. burnetii is enzootic among ruminants and wild animals throughout much of the United States and that there is widespread human exposure to this pathogen. Sheep and goats appear to be a more important risk for human infection in the United States than cattle or wild animals, and research studies examining the natural history and transmission risk of Q fever in sheep and goats in this country should be encouraged.

Strategies for containing Ebola in West Africa.

The ongoing Ebola outbreak poses an alarming risk to the countries of West Africa and beyond. To assess the effectiveness of containment strategies, we developed a stochastic model of Ebola transmission between and within the general community, hospitals, and funerals, calibrated to incidence data from Liberia. We find that a combined approach of case isolation, contact-tracing with quarantine, and sanitary funeral practices must be implemented with utmost urgency in order to reverse the growth of the outbreak. As of 19 September, under status quo, our model predicts that the epidemic will continue to spread, generating a predicted 224 (134 to 358) daily cases by 1 December, 280 (184 to 441) by 15 December, and 348 (249 to 545) by 30 December. A combination of hygienic practices could feasibly check Ebola within 6 months. Recharging Ebola mitigation measures Effective drugs and vaccines for Ebola virus are not available, so what can be done? Pandey et al. used a mathematical model to analyze transmission in different scenarios: the community, hospitals, and at funerals. Achieving full compliance with any single control measure, such as case isolation, is impossible under prevailing conditions. However, with a minimum of 60% compliance, a combination of case isolation, hygienic burial, and contact tracing could reduce daily case numbers to single figures in 5 to 6 months. Success will also require persistence and sensitivity to local customs. Science, this issue p. 991

Association of intraspecific wounding with hantaviral infection in wild rats (Rattus norvegicus).

The potential for hantaviral transmission among wild Norway rats by wounding associated with aggressive interactions was evaluated using a prospective sero-epidemiological study coupled with a mark-release-recapture survey. There was a significant association between an animals serological status and the presence of wounds. Longitudinal studies of marked and released animals showed seroconversion between captures was associated with wounding between captures more often (33%) than expected by chance, while unwounded animals seroconverted less often (8%) than expected. Typically, less than a 5% difference was found when comparing the incidence of seroconversion with the predicted rate based on wounding and seroprevalence. Infection was highly associated with the onset of sexual maturity and aggression but decoupled from rat age and the length of environmental exposure. Seroconversions occurred at times most associated with aggressive encounters and least associated with amicable behaviours that could lead to aerosol transmission.

Rats of the genus Rattus are reservoir hosts for pathogenic Bartonella species: an Old World origin for a New World disease?

Bartonella species were isolated from the blood of 63 of 325 Rattus norvegicus and 11 of 92 Rattus rattus from 13 sites in the United States and Portugal. Infection in both Rattus species ranged from 0% (e.g., 0/87) to approximately 60% (e.g., 35/62). A 337-bp fragment of the citrate synthase (gltA) gene amplified by polymerase chain reaction was sequenced from all 74 isolates. Isolates from R. norvegicus were most similar to Bartonella elizabethae, isolated previously from a patient with endocarditis (93%-100% sequence similarity), followed by Bartonella grahamii and other Bartonella species isolated from Old World rodents (Clethrionomys species, Mus musculus, and Rattus species). These data suggest that Rattus species are a reservoir host for pathogenic Bartonella species and are consistent with a hypothesized Old World origin for Bartonella species recovered from Rattus species introduced into the Americas.