Rift Valley fever MP-12 vaccine elicits an early protective immune response in mice.

Rift Valley fever virus (RVFV) is an important mosquito-borne pathogen that causes outbreaks of severe disease in people and livestock throughout Africa and the Arabian Peninsula. The development of an effective veterinary and human vaccine to protect against Rift Valley fever (RVF) disease remains a high priority. The live attenuated RVFV MP-12 is a promising vaccine candidate for the prevention of RVF in both human and domestic ruminants. The aim of this study was to determine the onset of protective immunity elicted in mice by a single dose of this vaccine. Groups of CD-1 mice were vaccinated intraperitoneally with RVFV MP-12 vaccine and challenged on days 2, 5, 6 and 7 post-vaccination (PV) with a lethal dose of virulent RVFV. The mice were observed once daily for terminal morbidity and blood samples were obtained from the retro-orbital sinus complex on days 23 and 28 PV of surviving mice to determine RVFV neutralizing antibody titers. In one test, 2 of 3 mice challenged on day 2 PV survived and all 3 mice challenged at days 5 and 7 PV also survived. A second test of 10 mice per group was performed, and half (5) of those challenged at day 2 PV survived while all (10) survived challenge at day 4 and 6 PV. All surviving animals develop antibody that ranged from 1:80 to 1:1,280 PV. In a separate experiment, RVFV MP-12 vaccinated CD-1 mice, but not challenged developed a low viremia for the first 3 days PV and neutralzing antibody was detected on days 5 through day 28 PV. These findings demonstrated that the RVFV MP-12 vaccine elicited a rapid protective immune response in mice as early as 2 days PV, thus further supporting the effectiveness of this vaccine candidate for preventing RVF among humans and domestic ruminants.

Safety and immunogenicity of the Rift Valley fever arMP-12 ΔNSm21/384 candidate vaccine in pregnant ewes.

Rift Valley fever (RVF) poses a threat to human and animal health as well as economic losses due to abortion, new-born teratogenic effect and mortality. Safe and effective vaccines are critically needed to prevent the disease in humans and livestock. The objective of this study was to assess safety and immunogenicity of the Rift Valley fever virus (RVFV) arMP-12DNSm21/384 attenuated vaccine in 32 pregnant ewes at different stages of pregnancy including 17 ewes vaccinated during the early stage (G1) of pregnancy (<35 days) and 15 ewes vaccinated during the last two stages (G2) of pregnancy (>35 days). Ewes were monitored for clinical observations, rectal temperature and abortions and lambs were monitored for general health and rectal temperature. Vaccinated ewes and lambs were periodically sampled for their neutralizing antibody response to RVFV vaccination. All ewes were positive for antibody two weeks post-vaccination and 79% of ewes were positive at delivery. None of the 32 ewes aborted during pregnancy and all ewes vaccinated during the G2 stages of pregnancy gave birth to healthy lambs. However, among the 17 ewes vaccinated during the G1 stage of pregnancy, 2 ewes gave birth to 2 lambs with fore limb malformations that died at 1-day of age. One ewe gave birth to 2 punny twins that died at 2 days of age. Another ewe, gave birth to one lamb with a deformed tail that died at 20 days of age. At post-mortem, tissues of dead lambs (spleen, lung, brain and long bone) were negative for RVFV by PCR assay. While the findings did not link the malformed lambs directly to infection by the vaccine virus, these results indicated that pregnant sheep should not be vaccinated with the RVFV arMP-12DNSm21/384 vaccine during the first month of gestation.

Transovarial transmission of LaCrosse virus (California encephalitis group) in the mosquito, Aedes triseriatus.

LaCrosse virus (California encephalitis group) was recovered from F(1) eggs, larvae, and adults produced by experimentally infected Aedes triseriatus. The F(1) females transmitted the virus by bite to suckling mice and chipmunks. This, plus isolations of LaCrosse virus from larvae collected from their natural habitats in enzootic areas and from males and females reared from them, suggests that transovarial transmission is the overwintering mechanism for this arbovirus in northern United States.

Isolation of St. Louis encephalitis virus from overwintering Culex pipiens mosquitoes.

Two strains of St. Louis encephalitis virus were isolated from overwintering mosquitoes collected in Maryland and Pennsylvania during January and February 1977. There isolations from Culex pipiens constitute evidence that a mosquito-borne flavivirus can persist in a vector mosquito in temperate climates during the winter season.

Safety and immunogenicity of a live attenuated Rift Valley Fever recombinant arMP-12ΔNSm21/384 vaccine candidate for sheep, goats and calves.

Rift Valley fever (RVF) causes serious health and economic losses to the livestock industry as well as a significant cause of human disease. The prevention of RVF in Africa is a global priority, however, available vaccines have only been partially effective. Therefore, the objective of this study was to evaluate the safety and immunogenicity of a live, attenuated recombinant RVFV arMP-12ΔNSm21/384 nucleotide deletion vaccine candidate in domestic ruminants. Evaluation involved testing to determine the infectivity titer of the vaccine virus in Vero cells for industrial scale up vaccine production. Safety experiments were conducted to determine the potential of the vaccine virus to revert to virulence by serial passages in sheep, the possibility of virus spread from vaccinated sheep and calves to unvaccinated animals, and the potential health effects of administering overdoses of the vaccine to sheep, goats and calves. The immunogenicity of 3 doses of 104, 105 and 106 Tissue Culture Infectious Doses50% (TCID50) of the vaccine was assessed in 3 groups of 10 sheep and 3 groups of 10 goats, and doses of 105, 106 and 107 TCID50 was evaluated in 3 groups of 10 calves subcutaenous vaccintation. The results showed that the infectivity titer of the vaccine virus was 108.4 TCID50/ml, that the vaccine did not spread from vaccinated to un-vaccinated animals, there was no evidence of reversion to virulence in sheep and the vaccine overdoses did not cause any adverse effects. The immunogenicity among sheep, goats and calves indicated that doses of 104-106 TCID50 elicited detectable antibody by day 7 post-vaccination (PV) with antibody titers ranging from 0.6 log to 2.1 log on day 14 PV with sustained titers through day 28 PV. Overall, these findings indicated that the RVFV arMP-12ΔNSm21/384 vaccine is a promising candidate for the prevention of RVF among domestic ruminants.

Evaluation of cotton rats as a model for severe acute respiratory syndrome.

Experimental studies were conducted to evaluate two species of cotton rats, Sigmodon hispidus and Sigmodon fulviventer, as a model for severe acute respiratory syndrome (SARS). Blood and turbinate wash samples, and lung tissue were collected from each animal at different time points after SARS coronavirus (CoV) infection for determining the growth curve of virus, if any, by the standard infectivity assay in Vero E6 cells. In addition, sections of the lung, liver, spleen, and kidney were taken and used for histology analysis. All animals were observed daily for signs of illness, and in some experiments, animals were weighed on the day when they were sacrificed. The results indicated that the cotton rat species, S. hispidus and S. fulviventer, were not a useful model for either SARS-CoV infection or disease. This observation was supported by the absence of any signs of illness, the failure to consistently demonstrate virus in the blood and tissues, and the absent of any notable histopathology. However, infected animals were capable of producing neutralizing antibodies against SARS-CoV, suggesting the seroconversion did occur. Further studies are warranted to consider other animal species in efforts to find better animal models for the evaluation of SARS-CoV vaccines and antiviral drugs.

Anatomical flow phantoms of the nonplanar carotid bifurcation, part I: computer-aided design and fabrication.

Doppler ultrasound is widely used in the diagnosis and monitoring of arterial disease. Current clinical measurement systems make use of continuous and pulsed ultrasound to measure blood flow velocity; however, the uncertainty associated with these measurements is great, which has serious implications for the screening of patients for treatment. Because local blood flow dynamics depend to a great extent on the geometry of the affected vessels, there is a need to develop anatomically accurate arterial flow phantoms with which to assess the accuracy of Doppler blood flow measurements made in diseased vessels. In this paper, we describe the computer-aided design and manufacturing (CAD-CAM) techniques that we used to fabricate anatomical flow phantoms based on images acquired by time-of-flight magnetic resonance imaging (TOF-MRI). Three-dimensional CAD models of the carotid bifurcation were generated from data acquired from sequential MRI slice scans, from which solid master patterns were made by means of stereolithography. Thereafter, an investment casting procedure was used to fabricate identical flow phantoms for use in parallel experiments involving both laser and Doppler ultrasound measurement techniques.

Isolation of viruses from mosquitoes (Diptera: Culicidae) collected in the Amazon Basin region of Peru.

As part of a comprehensive study on the ecology of arthropod-borne viruses in the Amazon Basin region of Peru, we assayed 539,694 mosquitoes captured in Loreto Department, Peru, for arboviruses. Mosquitoes were captured either by dry ice-baited miniature light traps or with aspirators while mosquitoes were landing on human collectors, identified to species, and later tested on Vero cells for virus. In total, 164 virus isolations were made and included members of the Alphavirus (eastern equine encephalomyelitis, Trocara, Una, Venezuelan equine encephalomyelitis, and western equine encephalomyelitis viruses), Flavivirus (Ilheus and St. Louis encephalitis), and Orthobunyavirus (Caraparu, Itaqui, Mirim, Murutucu, and Wyeomyia viruses) genera. In addition, several viruses distinct from the above-mentioned genera were identified to the serogroup level. Eastern equine encephalomyelitis virus was associated primarily with Culex pedroi Sirivanakarn & Belkin, whereas Venezuelan equine encephalomyelitis virus was associated primarily with Culex gnomatos Sallum, Huchings & Ferreira. Most isolations of Ilheus virus were made from Psorophora ferox (Von Humboldt). Although species of the Culex subgenus Melanoconion accounted for only 45% of the mosquitoes collected, 85% of the virus isolations were made from this subgenus. Knowledge of the viruses that are being transmitted in the Amazon Basin region of Peru will enable the development of more effective diagnostic assays, more efficient and rapid diagnoses of clinical illnesses caused by these pathogens, risk analysis for military/civilian operations, and development of potential disease control measures.

The epidemiology of HIV-1 infection in Peru, 1986-1990.

OBJECTIVE: To determine the extent of the HIV-1 epidemic in Peru. DESIGN: Part of a national serosurvey in Peru. METHODS: Between January 1986 and December 1990, 140,976 serum samples were tested for HIV-1 antibody. RESULTS: HIV-1 antibody was found in a high percentage of serum samples provided by 4300 homosexual men (26%), 2204 male sexually transmitted disease patients (10%), 145 drug users (13%), 269 hemophiliacs (10%), and 146 unlicensed female prostitutes (10%). In addition, the prevalence of HIV-1 infection increased substantially among these groups between the beginning and end of the survey period. A low but rising prevalence of HIV-1 antibody was found during this period among serum samples provided by 83,526 blood donors and 11,101 military personnel:total period prevalence, 0.25 and 0.32%, respectively. CONCLUSION: These data indicate that HIV-1 infection is epidemic in Peru among groups at high risk of sexually and parenterally transmitted diseases, and that the risk of infection appears to be low but possibly increasing among the general population.

PIP: The findings of a national seroprevalence survey conducted in Peru during 1986-90 indicate accelerating rates of human immunodeficiency virus (HIV) among population groups at high risk of sexually transmitted diseases. Two databases were maintained: 1) January 1986-December 1988 and 2) January 1989-December 1990. Of the 140,976 survey participants, 3345 (2.4%) were HIV-positive by Western blot. 2591 participants were selected because of clinical signs suggestive of acquired immunodeficiency syndrome (AIDS); 46.7% were HIV-positive, but the prevalence increased from 19% in the 1986-88 period to 60% during 1989-90. Among the 4300 men who identified themselves as homosexual or bisexual, 26% were seropositive (8% during 1986-88 and 41% during 1989-90). HIV prevalence among 2204 men attending a sexually transmitted diseases clinic was 10.3%, with an increase from 2.0% in the first period to 19.0% in the later period. 10.4% of the 269 hemophiliacs were HIV-infected, with an increase from 8% to 36%. Among the 145 intravenous drug users, the prevalence rose from 1% during 1986-88 to 27% during 1989-90. Among 5827 registered female prostitutes, the prevalence rose from 0.3% to 0.7%; however, a 1990 analysis of 146 unregistered prostitutes revealed a rate of 9.6%. The HIV rates among 285 female and 105 male heterosexual partners of known HIV-positive persons were 50.2% and 40.0%, respectively. HIV prevalence increased from 0.8% during 1986-88 to 8.0% during 1989-90 among 1532 men and 1247 women who requested anonymous HIV testing. The prevalence among 542 male and 615 female medical personnel was 2.3%. Among 78,793 volunteer and 4733 paid blood donors, HIV prevalence was 0.2% (0.3% among paid donors). The period prevalence among 11,101 male military recruits and active duty members increased from 0.009% to 0.5%. Finally, only 0.3% of 21,595 applicants for immigration visas were HIV-positive, and there were no HIV cases among 1234 pregnant women attending antenatal clinics. Although the very low HIV prevalence among military personnel and pregnant women suggests that the virus is not yet widely disseminated within the general population, the finding that 28% of HIV-positive men were married and engaged in bisexual behavior suggests potential for heterosexual transmission in the years ahead.

Emerging genetic diversity of HIV-1 in South America.

OBJECTIVE: Genotype determination and risk group analysis of HIV-1 infected individuals in selected regions of South America. DESIGN: Cross-sectional convenience sampling of HIV-1-positive individuals in Peru, Ecuador, Uruguay and Paraguay from March, 1994 through September, 1998. METHODS: HIV-1-positive subjects were identified through the national AIDS surveillance program in each country. A standardized questionnaire was used to obtain demographic, clinical and risk factor data on each study subject. Viral DNA was extracted from participants' peripheral blood mononuclear cells either directly or after co-cultivation. A nested PCR was used to obtain selected fragments of the envelope genes for genotyping by the heteroduplex mobility assay (HMA). A 600 bp sequence encompassing the V3 loop was sequenced from a selection of 23 of these samples for phylogenetic analysis and confirmation of HMA genotype. RESULTS: Among the 257 successfully genotyped HIV-1-positive samples, genotype B was found in 98.3% (228/232) of those obtained from subjects in Peru, Ecuador, and Paraguay. In contrast, 56% (14/25) of the samples from Uruguay were genotype F, and the remainder were genotype B. Genotype F was detected for the first time in Peru (2/224) and Paraguay (1/4), and genotype A for the first time in Peru (1/224). Phylogenetic analysis confirmed the genotype identified by HMA in the 23 samples sequenced. There was no detectable genetic clustering of HIV-1 within the different high-risk groups or geographic locations. CONCLUSIONS: These findings verify and extend the presence of several different HIV-1 genotypes in South America.

Transovarial transmission of LaCrosse virus in Aedes triseriatus.

As part of a continuing investigation on the ecology of LaCrosse virus in Wisconsin, field and laboratory studies were conducted to explore the possibility that the virus is transmitted transovarially in A. triseriatus mosquitoes. In laboratory experiments, A. triseriatus mosquitoes were infected by ingesting LaCrosse virus in defibrinated blood. LaCrosse virus was recovered from F1 eggs, larvae, and adults that originated from the infected parent mosquitoes. In a subsequent field study aimed at determining if transovarial transmission accounted for the survival of LaCrosse virus during the winter season, larvae that originated from overwintering A. triseriatus eggs were collected from a LaCrosse virus enzootic area in southwestern Wisconsin. LaCrosse virus was isolated from these larvae and from adult A. triseriatus that were reared from field-collected larvae. These findings strongly imply that A. triseriatus is the reservoir of LaCrosse virus and that transovarial transmission is the mechanism responsible for the maintenance of the virus during the winter season in the north central region of the United States.

Inhibition of Crimean-Congo hemorrhagic fever viral infectivity yields in vitro by ribavirin.

Ribavirin was evaluated as a potential therapeutic for Crimean-Congo hemorrhagic fever (CCHF). Viral yields for strains of CCHF virus from Europe, Asia, and Africa in African green monkey kidney (Vero) cells were markedly reduced by this drug. Some CCHF viral strains appeared more sensitive than others, but in general, ribavirin doses as low as 5 micrograms/ml caused a transient reduction of viral yields. A further reduction in viral yields was induced by a dose of 25 micrograms/ml, and evidence of viral replication was not demonstrated in cells treated with 50 or 250 micrograms/ml. In contrast, a dose of ribavirin at least 9 times greater was required to induce a comparable inhibitory effect on the yields of Rift Valley fever virus, for which the drug has been shown to inhibit replication in monkeys and rodents.

Vertical transmission of West Nile virus by Culex and Aedes species mosquitoes.

Experiments were conducted to determine whether West Nile (WN) virus was transmitted vertically by colonized strains of Aedes albopictus, Ae. aegypti, and Culex tritaeniorhynchus. Female mosquitoes were infected by intrathoracic inoculation with WN virus, and the F1 progeny were tested for virus by the fluorescence antibody technique and the newborn mouse assay. Each of the three mosquito species transmitted WN virus to F1 adults derived from immature forms reared at 26 degrees C. The minimal filial infection rate (MFIR) ranged from 1:124 to 1:138 for Ae. albopictus, from 1:62 to 1:172 for Ae. aegypti, and from 1:325 to 1:859 for Cx. tritaeniorhynchus. The MFIR for Cx. tritaeniorhynchus reared at 20 degrees C was 1:213 for larvae and 1:390 for pupae, and 1:208 for larvae and 1:554 for pupae reared at 26 degrees C. These data are the first reported evidence of vertical transmission of WN virus by mosquitoes, and therefore warrant further studies to determine whether vertical transmission occurs among WN viral-infected mosquitoes in nature.

Experimental transmission of trivittatus virus (California virus group) by Aedes trivittatus.

The mosquito, Aedes trivittatus, when fed through a membrane a trivittatus virus dosage of 10(3.4) to 10(5.5) suckling mouse LD50/0.03 ml of blood, transmitted the virus to suckling mice. Virus multiplication indicative of a biological vector occurred in this species. When Aedes vexans and A. triseriatus ingested similar doses of trivittatus virus, both the infection and transmission rates were low and virus multiplication was poor. These results, added to evidence based on virus isolations from mosqito populations in nature, indicate that A. trivittatus is the primary vector of trivittatus virus in the north central United States.

Effect of temperature on the vector efficiency of Aedes aegypti for dengue 2 virus.

The effect of temperature on the ability of Aedes aegypti to transmit dengue (DEN) 2 virus to rhesus monkeys was assessed as a possible explanation for the seasonal variation in the incidence of dengue hemorrhagic fever in Bangkok, Thailand. In two laboratory experiments, a Bangkok strain of Ae. aegypti was allowed to feed upon viremic monkeys infected with DEN-2 virus. Blood-engorged mosquitoes were separated into two groups and retained at constant temperatures. Virus infection and transmission rates were determined for Ae. aegypti at intervals ranging from 4 to 7 days during a 25-day incubation period. Results of the first experiment for mosquitoes infected with a low dose of DEN-2 virus and maintained at 20, 24, 26, and 30 degrees C, indicated that the infection rate ranged from 25% to 75% depending on the incubation period. However, DEN-2 virus was transmitted to monkeys only by Ae. aegypti retained at 30 degrees C for 25 days. In the second experiment, the infection rate for Ae. aegypti that ingested a higher viral dose, and incubated at 26, 30, 32, and 35 degrees C ranged from 67% to 95%. DEN-2 virus was transmitted to monkeys only by mosquitoes maintained at greater than or equal to 30 degrees C. The extrinsic incubation period was 12 days for mosquitoes at 30 degrees C, and was reduced to 7 days for mosquitoes incubated at 32 degrees C and 35 degrees C. These results imply that temperature-induced variations in the vector efficiency of Ae. aegypti may be a significant determinant in the annual cyclic pattern of dengue hemorrhagic fever epidemics in Bangkok.

Experimental transmission of Crimean-Congo hemorrhagic fever virus by Hyalomma truncatum Koch.

Larval Hyalomma truncatum ticks were infected with Crimean-Congo hemorrhagic fever (CCHF) virus by allowing them to engorge on viremic newborn mice. The overall tick infection rate was 4.4% (24/542). Virus was detected in specimens for greater than or equal to 160 days postinfection. Transstadial transmission to the adult tick stage was observed and horizontal transmission to a mammalian host was demonstrated. Horizontal transmission of CCHF virus to uninfected adult ticks occurred while feeding with transstadially infected ticks on the same host. No evidence of transovarial virus transmission from infected female ticks to their 1st generation progeny was observed.

Experimental infection of six species of ixodid ticks with Dugbe virus (family Bunyaviridae, genus Nairovirus).

The vector potential of each of 6 species of colonized North American and African ixodid ticks was assessed by intracoelomic inoculation with Dugbe virus (IbAr 1792, 14th passage in suckling mouse brain) and viral titers were monitored after selected incubation periods. Persistence of Dugbe virus for greater than or equal to 53 days in 5 species (Dermacentor andersoni, D. variabilis, Amblyomma americanum, Rhipicephalus appendiculatus, and R. sanguineus) indicates that infection occurred. Viral titers were significantly higher in female vs. male D. variabilis, R. appendiculatus, and A. americanum after blood feeding. Blood feeding had no significant effect on the viral titers of either female or male R. sanguineus. D. andersoni males also exhibited no significant change in viral titers after blood-feeding, but 100% (20/20) of drop-off females and 96% (24/25) of post-oviposition females (36 days postinoculation) contained no detectable virus even though virus was still found in unfed specimens less than or equal to 124 days postinoculation. Virus was not recovered from greater than 30,000 1st generation progeny (eggs, larvae, nymphs, adults) collected as eggs from inoculated female D. andersoni, D. variabilis, R. sanguineus, and R. appendiculatus 27-51 days postinoculation. R. sanguineus and R. appendiculatus transmitted Dugbe virus to guinea pigs when allowed to feed 1-3 weeks postinoculation.

Oropouche virus transmission in the Amazon River basin of Peru.

Seroepidemiologic studies were conducted to determine the prevalence of Oropouche (ORO) viral antibody, risk factors, and the incidence of infection among residents of the Amazon region of Peru. Blood samples, as well as demographic, cultural, and medical history data, were collected from residents in a sector of the city of Iquitos and in an adjacent rural and three neotropical rain forest communities. Blood specimens were obtained approximately one year later from a cohort of the same study subjects who were negative for ORO antibody on the initial cross-sectional survey. Sera were tested for ORO IgG antibody by an enzyme-linked immunosorbent assay. Antibody prevalences were 35% for residents of the urban population, 24-46% for the forest communities, and 18% for the rural community. Antibody prevalence increased with age, and subjects who were seropositive were significantly (P = 0.001) older (mean = 33 years) than the seronegative subjects (mean = 15 years). Multivariate analysis revealed that only age, urban and forest residence, and occupation as a farmer or housekeeper remained significantly associated with seropositivity. Seroconversion data for the same populations one year later demonstrated evidence of ORO viral infection among 28% of the residents in the rural community and 2% or less in the forest and urban communities. Oropouche virus infection was significantly associated with older age (P = 0.04) in the rural community (P < 0.001). These data support prior evidence of ORO viral infection among residents of Iquitos and surrounding villages and suggest that transmission of this virus occurs continuously in the population of this area of the Amazon basin.

Identification and genetic analysis of Panama-genotype Venezuelan equine encephalitis virus subtype ID in Peru.

Venezuelan equine encephalitis (VEE) virus was isolated in 1993, 1994, and 1995 from human cases of acute, undifferentiated, febrile illness in the Peruvian Amazon Basin. Two virus isolates were recovered in 1994 from Peruvian soldiers at a jungle outpost near Pantoja in northern Peru, and 10 isolates were obtained from military personnel and civilians in 1993-1995 in Iquitos, an urban center in northeastern Peru. The genetic relationship of these isolates to other VEE virus strains was determined by sequencing 856-867 nucleotide reverse transcription-polymerase chain reaction fragments derived from the PE2 glycoprotein gene. The sequences were compared with those of other VEE virus strains, including representatives of the IAB, IC, ID, IE, II, and IIIC subtypes. The two Pantoja isolates were most closely related to subtype IC and ID viruses previously isolated in Colombia and Venezuela, and to the ID viruses isolated during the 1970s in Iquitos. All of the recent Iquitos isolates were similar to one another, but they were more closely related to Panamanian ID strains than to isolates previously obtained in Iquitos, Peru, or in Colombia and Venezuela. The recent Iquitos VEE viral isolates were the first Panama-genotype VEE ID virus strains identified outside of the Republic of Panama.

Serologic evidence of Jamestown Canyon and Keystone virus infection in vertebrates of the DelMarVa Peninsula.

Serological data accumulated during the past decade indicated that a variety of feral and domestic animals of the Delaware-Maryland-Virginia (DelMarVa) Peninsula were infected with Jamestown Canyon (JC) and/or Keystone (KEY) viruses (Bunyaviridae, California serogroup). Neutralizing (N) antibody to JC virus was most prevalent in white-tailed deer, sika deer, cottontail rabbits and horses. KEY virus N antibody was detected most frequently in gray squirrels and domestic goats. N antibody indicative of past infection by one or both viruses also was found in raccoons, horses and humans. JC and/or KEY virus N antibodies were not demonstrable in sera of several other species of small mammals and reptiles. Investigations were extended to evaluate the role of domestic goats as an amplifying host of JC and KEY viruses and to assess their potential as sentinels of virus transmission. Goats maintained in the Pocomoke Cypress Swamp during the summer season of 1978, acquired N antibodies to JC and KEY viruses. Following experimental inoculation with either JC or KEY virus, all goats developed N antibody despite the absence of a demonstrable viremia in most animals. Goats proved to be effective as sentinels for monitoring the transmission of JC and KEY viruses; however, the exceptionally low titers or absence of viremia following inoculation with these viruses would seem to preclude a potential virus-amplifying role for this species. Although findings implicated primarily gray squirrels and white-tailed deer as possible amplifying hosts of KEY and JC virus, respectively, further investigations will be required to clarify their role, particularly since both viruses may be maintained entirely by transovarial transmission.