Genetic identification of a hantavirus associated with an outbreak of acute respiratory illness.

A mysterious respiratory illness with high mortality was recently reported in the southwestern United States. Serologic studies implicated the hantaviruses, rodent-borne RNA viruses usually associated elsewhere in the world with hemorrhagic fever with renal syndrome. A genetic detection assay amplified hantavirus-specific DNA fragments from RNA extracted from the tissues of patients and deer mice (Peromyscus maniculatus) caught at or near patient residences. Nucleotide sequence analysis revealed the associated virus to be a new hantavirus and provided a direct genetic link between infection in patients and rodents.

Orthobunyavirus antibodies among humans in selected parts of the Rift Valley and northeastern Kenya.

Ngari, Bunyamwera, Ilesha, and Germiston viruses are among the mosquito-borne human pathogens in the Orthobunyavirus genus, family Bunyaviridae, associated with febrile illness. Although the four orthobunyaviruses have been isolated from mosquito and/or tick vectors sampled from different geographic regions in Kenya, little is known of human exposure in such areas. We conducted a serologic investigation to determine whether orthobunyaviruses commonly infect humans in Kenya. Orthobunyavirus-specific antibodies were detected by plaque reduction neutralization tests in 89 (25.8%) of 345 persons tested. Multivariable analysis revealed age and residence in northeastern Kenya as risk factors. Implementation of acute febrile illness surveillance in northeastern Kenya will help to detect such infections.

Update: outbreak of hantavirus infection--southwestern United States, 1993.

An outbreak of illness associated with hantavirus infection continues to be investigated by state health departments in New Mexico, Arizona, Colorado, and Utah; the Indian Health Service; and CDC, with the assistance of the Navajo Nation Division of Health. This report updates information regarding the outbreak and presents information on two cases that occurred in the 10 months preceding this outbreak.

Update: hantavirus pulmonary syndrome--United States, 1993.

A unique hantavirus has been identified as the cause of the outbreak of respiratory illness (hantavirus pulmonary syndrome [HPS]) first recognized in the southwestern United States in May 1993. The habitat of the principal rodent reservoir for this virus, Peromyscus maniculatus (deer mouse), extends throughout most of the United States except the Southeast. Through October 21, 1993, HPS has been confirmed in 42 persons reported to CDC from 12 states (Figure 1). This report summarizes major clinical, pathologic, and diagnostic findings in patients with this newly recognized syndrome; addresses the use of the investigational antiviral drug ribavirin; and presents revised screening criteria for national surveillance.

Update: outbreak of hantavirus infection--southwestern United States, 1993.

Since May 1993, the New Mexico Department of Health, the Arizona Department of Health, the Colorado Department of Health, the Utah Department of Health, the Indian Health Service, and CDC, with the assistance of the Navajo Nation Division of Health, have been investigating an outbreak of illness associated with hantavirus infection. This report updates information regarding the relation between illness and infection with a previously unrecognized hantavirus.

Update: outbreak of hantavirus infection--southwestern United States, 1993.

Since May 1993, the New Mexico Department of Health, the Arizona Department of Health Services, the Colorado Department of Health, the Utah Department of Health, the Indian Health Service (IHS), and CDC, with the assistance of the Navajo Nation Division of Health, have been investigating an outbreak of acute illness characterized by a prodrome most commonly including fever, myalgias, headache, and cough, followed rapidly by respiratory failure. Preliminary laboratory findings have suggested this outbreak is associated with infection with a hantavirus or a closely related agent. This report updates the ongoing investigation of this outbreak.

Utilization of autopsy RNA for the synthesis of the nucleocapsid antigen of a newly recognized virus associated with hantavirus pulmonary syndrome.

A newly recognized hantavirus was recently found to be associated with an outbreak of acute respiratory illness in the southwestern United States. The disease, which has become known as hantavirus pulmonary syndrome, has an unusually high mortality (64%). Virus isolation attempts have been unsuccessful thus far, resulting in a lack of homologous antigen for use in diagnostic assays. For this reason, a molecular approach was initiated to produce recombinant homologous antigen. The virus nucleocapsid (N) protein was selected, since N has been shown to be a sensitive antigenic target in other hantavirus systems. The N protein open reading frame of the virus S genome segment was synthesized from frozen autopsy tissue by polymerase chain reaction amplification, followed by cloning and expression in Hela cells (vaccinia-T7 RNA polymerase system) and Escherichia coli. N protein-expressing Hela cells served as excellent antigens for an improved indirect immunofluorescence assay. Use of the E. coli-expressed N protein in an enzyme-linked immunosorbent assay improved the sensitivity and specificity when compared with heterologous antigens used previously. Preliminary analysis also indicates that the higher sensitivity could result in earlier detection of infected persons. These data demonstrate that even in the absence of a virus isolate, the necessary homologous antigen can be produced and can serve to improve the detection and diagnostic capabilities needed to combat this newly recognized fatal respiratory illness in the United States.

Characterization of murine Caraparu Bunyavirus liver infection and immunomodulator-mediated antiviral protection.

A rapid, peripheral disease model utilizing the Bunyavirus, Caraparu, was established in mice for the evaluation of antiviral therapy with immunomodulators. 4-6-week-old B6C3F1 female mice, inoculated intraperitoneally with virus, developed coagulative liver necrosis and died between 4-6 days after infection. This Caraparu disease model was relatively resistant to treatment with immunomodulators, such as ABMP, Ampligen, alpha-interferon (IFN-alpha) or beta-interferon (IFN-beta). However, a significant increase in median survival time (MST) was consistently observed upon treatment with gamma-interferon (IFN-gamma). The nucleoside analog--ribavirin--was highly effective against Caraparu virus in repeated treatment schedules begun on either day -1, day 0, or day +1 of infection. Ribavirin gave little protection when initiation of treatment was delayed until day +2. However, combined treatment with IFN-gamma, starting on day 0 and ribavirin starting on day +2, significantly reduced mortality.

Effect of human, recombinant interleukin 2 on Punta Toro virus infections in C57BL/6 mice.

The effect of human recombinant interleukin-2 (rIL-2) on Punta Toro virus (PTV) infection was investigated in C57BL/6 mice. Immunologic and viral parameters were assessed after mice were treated i.p. with rIL-2 for 5 days. Treatment of mice with 25000 and 12500 units/mouse of rIL-2 resulted in significant inhibition of the disease as indicated by increases in survival of mice as well as decreases in liver and serum virus titers. Serum glutamic oxalic acid and pyruvic acid transaminase levels were also lowered indicating reduced liver damage. Murine IL-2 production returned to normal or above-normal levels in rIL-2 treated mice. Natural killer cell activity was also moderately stimulated by rIL-2 treatment. Significant amounts of interferon were not detected in the sera of treated mice. Weight gain and survival rates were similar for both toxicity and normal controls indicating that rIL-2 treatments had no toxic effect.

Outbreak of Hantavirus pulmonary syndrome in the southwestern United States. Response of pathologists and other laboratorians.

During late spring and early summer of 1993, national and international media called worldwide attention to a cluster of deaths in the southwestern United States. These patients succumbed to a rapidly progressive severe respiratory distress syndrome. After notification of state and national health agencies in mid-May, a major effort was launched to determine the cause of this often fatal respiratory distress syndrome, to advise the public on safety measures, and to determine the method of spread of this "mystery illness." Within weeks of recognition of the early cases, the Centers for Disease Control and Prevention announced the probable agent, a Hantavirus. This report details the response of pathologists, medical technologists, and other laboratory scientists to this new viral epidemic, with emphasis on activities that occurred within New Mexico.

Kairi virus identified from a febrile horse in Argentina.

A virus isolated from the blood of a febrile horse in Argentina was identified as a strain of Kairi virus. This is the fifth Bunyamwera serogroup virus isolated from livestock and wild animals in the Americas. Bunyamwera serogroup viruses have been isolated from febrile humans in the Americas and Africa.

Isolation of the causative agent of hantavirus pulmonary syndrome.

Investigation of a recent outbreak of acute respiratory illness in the southwestern United States resulted in the recognition of a new disease, hantavirus pulmonary syndrome (HPS) with high mortality. Different animals and cell lines were used in attempts to isolate the causative agent. A previously unknown hantavirus was passaged in laboratory-bred deer mice, recovered from lung tissues of a deer mouse, Peromyscus maniculatus, and propagated in the E6 clone of Vero cells. Virus antigen was readily detected in the infected cells by an indirect immunofluorescence assay, using convalescent-phase sera from HPS patients. By electron microscopy, the virus was shown to have the typical morphologic features of members of the genus Hantavirus, family Bunyaviridae. Virus sequences corresponded to those previously detected by a nested reverse transcriptase-polymerase chain reaction assay of hantavirus-infected specimens from rodents and humans. This newly recognized virus, the etiologic agent of HPS, has been tentatively named Muerto Canyon virus.

Transovarial maintenance of San Angelo virus in sequential generations of Aedes albopictus.

An Aedes albopictus line originally selected for efficient transovarial transmission (TOT) of San Angelo (SA) virus displayed a progressive decline in filial infection rate (FIR) whenever artificial selection pressure was relaxed. This observation brought into question whether the efficiency of TOT in this vector-virus model was sufficiently high to permit persistent vertical transmission over numerous freely reproducing generations. By tracing descendants of individual transovarially infected females for several generations, it was found in the present study that this decline in FIR resulted not only from the spontaneous appearance of uninfected females, but also from an accumulation of transovarially infected females which were themselves inefficient transovarial transmitters. Some efficient transovarial transmitters continued to be produced even when FIR had declined to low levels, assuring transmission of virus to subsequent generations, irrespective of the overall infection rate. Outcrossing experiments suggested that a "refractory" genetic factor or factors had been selected out of the TOT-efficient line during its long history of inbreeding. New TOT-efficient lines were more readily established from parenterally infected Ae. albopictus of a Taiwan strain than had been the original TOT-efficient line from a Hawaiian mosquito strain. One of the newly selected lines differed from the others in producing progenies with FIRs no higher than 36% in the third and fourth generations of transovarial passage. The familial and strain variation revealed by these studies suggests a genetic influence on both the establishment and maintenance of persistent oogonial infections.

Distribution of Bunyamwera serogroup viruses in North America, 1956-1984.

We attempted to tabulate all Bunyamwera serogroup (family Bunyaviridae, genus Bunyavirus) isolates from North America. By summarizing information from the laboratories of the Centers for Disease Control, data generously shared by other laboratories, and the published literature, we were able to accumulate data regarding 1,372 Bunyamwera serogroup viruses. These were: Tensaw (664, including 8 from vertebrates), Cache Valley (396, including 6 from vertebrates), Main Drain (160, including 14 from vertebrates), Lokern (69, including 8 from vertebrates), Northway (13, including 5 from vertebrates), Tlacotalpan (7), Santa Rosa (2), Santa Cruz (1 from a horse), and 60 of undetermined serotype. Virus isolation rates by month of collection were correlated with collection efforts, but associations of viruses and arthropod vectors varied by location, vertebrate host, and arthropod distribution. Tensaw virus was isolated principally from Anopheles crucians mosquitoes (466/656 isolates from arthropods) in the southeastern United States; Cache Valley virus principally from An. quadrimaculatus (94), Coquillettidia perturbans (59), Culiseta inornata (45), Aedes sollicitans (30), Psorophora columbiae (23), An. punctipennis (18), and Ae. vexans and trivittatus (18 each) mosquitoes (total = 305/382 isolates from arthropods from all of the United States and Canada, except the southeastern United States); Main Drain virus from Culicoides variipennis (31), Culicoides (Selfia) sp. (65), and Psorophora (23) and Aedes (21) species mosquitoes in the western United States; Lokern virus from Culicoides species (55/61 isolates from arthropods) in the western United States. Relationships between vector and vertebrate host distributions are discussed briefly in regard to geographic distribution of the Bunyamwera serogroup viruses.

Isolation of a newly recognized Bunyamwera serogroup virus from a febrile human in Panama.

A virus, strain 86MSP18, was isolated from the acute phase serum of a U.S. soldier with a febrile illness. He was stationed at Fort Sherman in the Republic of Panama when the onset of his illness occurred. A rise in neutralizing antibody to the viral isolate was observed between the patient's acute and convalescent-phase serum samples. Virus strain 86MSP18 has been shown by plaque reduction neutralization to be closely related to but distinct from Cache Valley virus and known subtypes. It appears to be a newly recognized subtype of Cache Valley virus and is believed to be the second isolation of a Cache Valley virus subtype from a human with a febrile illness. The name "Fort Sherman" virus for strain 86MSP18 is proposed.

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.

Viruses isolated from Panamanian sloths.

Seven virus strains were isolated in Vero cells from whole blood samples from 80 wild-caught sloths, Bradypus variegatus and Choloepus hoffmanni, from Central Panamá. Four strains of at least two different serotypes are related to Changuinola virus; two of these were associated with prolonged or recrudescent viremias. One strain is an antigenic subtype of Punta Toro virus, and another, described here as Bradypus-4 virus, is a new, antigenically ungrouped virus. A second new virus from sloths, Utive virus, forms an antigenic complex within the Simbu serogroup with Utinga and Pintupo viruses. Tests on sequential plasma samples from radio-marked free-ranging sloths and from recently captured animals maintained in captivity showed that both species develop neutralizing antibodies following naturally acquired virus infections. Antibodies against the Changuinola and Simbu serogroup viruses are widespread in both sloth species and are especially prevalent in Choloepus, but are virtually absent in all other wild vertebrate species tested.

Prevalence of neutralizing antibodies against California and Bunyamwera serogroup viruses in deer from mountainous areas of California.

Plaque reduction-serum dilution neutralization was used to evaluate the status of bunyavirus activity in deer in mountainous areas of California. Antibodies against 9 bunyaviruses were measured in 337 mule deer (Odocoileus hemionus hemionus, O. hemionus californicus, and O. hemionus inyoensis) and black-tailed deer (O. hemionus columbianus). More deer from high mountainous areas had neutralizing antibodies against Jamestown Canyon virus than did deer from low mountainous areas (23% vs. 9%; P less than 0.01). This finding is consistent with transmission by snow pool Aedes mosquitoes. Results for Jerry Slough virus were nearly identical to those for Jamestown Canyon virus, which is further evidence that these are strains of the same virus. Neutralizing antibodies against Northway virus were present in 26% of deer from high mountainous areas and 23% of deer from low mountainous areas, suggesting the involvement of a widespread vector, such as Culiseta inornata. Northway virus is not known to occur outside of Alaska and northwestern Canada. Low prevalences of antibodies were detected in deer to California encephalitis, La Crosse, and snowshoe hare viruses of the California serogroup; and Cache Valley, Lokern, and Main Drain viruses of the Bunyamwera serogroup.

Central nervous system involvement during infection by Phlebovirus toscana of residents in natural foci in central Italy (1977-1988).

Central nervous system (CNS) involvement was detected during infection caused by the sand fly-transmitted Phlebovirus Toscana. One hundred fifty-five cases of Toscana virus-associated meningitis or meningoencephalitis were identified in a survey that lasted ten years, conducted in two regions of central Italy. Diagnosis was performed by different serologic tests. A combination of hemagglutination-inhibition and plaque-reduction neutralization or indirect immunofluorescence for IgM, and enzyme-linked immunosorbent assays for IgM were considered the most suitable tests for the diagnosis of Toscana virus infection. A few strains of Toscana virus were isolated from the cerebrospinal fluid of seropositive patients. Toscana virus-associated CNS disease occurred during the summer, reaching a peak value in August, when the maximum activity of the sand fly vector occurs and virus isolates are obtained in their natural foci. The results suggest that Toscana virus should be considered as a possible cause of CNS disease in Mediterranean countries where sand flies of the genus Phlebotomus are known to be present.

Isolation of Batai virus (Bunyaviridae:Bunyavirus) from the blood of suspected malaria patients in Sudan.

From August through November 1988, 77,500 patients with fever presented to the municipal hospital and to eight government health centers in Kassala, a town of approximately 400,000 individuals in eastern Sudan. A diagnosis of malaria, based primarily on clinical presentation, was made in 14,395 individuals during this four-month period; fevers of unknown origin were diagnosed in 29 patients. A Bunyavirus that was antigenically similar or identical to Batai virus by complement fixation and plaque-reduction neutralization tests was recovered from two of 196 sera collected from patients with acute fever admitted to the municipal hospital in Kassala in October 1988. IgM antibody against this virus was detected by enzyme-linked immunosorbent assay in 7% of the sera from patients with acute fever tested and IgG antibody was detected in 61%.