Origin of the West Nile virus responsible for an outbreak of encephalitis in the northeastern United States.

In late summer 1999, an outbreak of human encephalitis occurred in the northeastern United States that was concurrent with extensive mortality in crows (Corvus species) as well as the deaths of several exotic birds at a zoological park in the same area. Complete genome sequencing of a flavivirus isolated from the brain of a dead Chilean flamingo (Phoenicopterus chilensis), together with partial sequence analysis of envelope glycoprotein (E-glycoprotein) genes amplified from several other species including mosquitoes and two fatal human cases, revealed that West Nile (WN) virus circulated in natural transmission cycles and was responsible for the human disease. Antigenic mapping with E-glycoprotein-specific monoclonal antibodies and E-glycoprotein phylogenetic analysis confirmed these viruses as WN. This North American WN virus was most closely related to a WN virus isolated from a dead goose in Israel in 1998.

Culex flavivirus isolates from mosquitoes in Guatemala.

A new strain of Culex flavivirus (family Flaviviridae, genus Flavivirus, CxFV), an insect virus first described in Japan, was isolated from adult Culex quinquefasciatus Say (Diptera: Culicidae) collected in 2006 from Izabal Department on the Caribbean coast of Guatemala. Mosquito pools were assayed for flavivirus RNA by using flavivirus group-specific primers that amplified a 720-bp region of the nonstructural (NS) 5 gene by standard reverse transcriptase-polymerase chain reaction. From 210 pools (1,699 mosquitoes), eight tested positive, and six of these mosquito pools produced virus isolates in Aedes albopictus Skuse C6/36 cells. Nucleotide sequence comparison of the eight flavivirus RNA-positive pools showed that there was 100% identity among them, and phylogenetic analysis of the NS5 and envelope gene regions indicated that they represent a strain of the recently described CxFV from Japan. This is the first report of an insect flavivirus from Central America.

West Nile virus in livestock and wildlife.

WN virus is one of the most ubiquitous arboviruses occurring over a broad geographical range and in a wide diversity of vertebrate host and vector species. The virus appears to be maintained in endemic foci on the African continent and is transported annually to temperate climates to the north in Europe and to the south in South Africa. Reports of clinical disease due to natural WN virus infection in wild or domestic animals were much less common than reports of infection (virus isolation or antibody detection). Until recently, records of morbidity and mortality in wild birds were confined to a small number of cases and infections causing encephalitis, sometimes fatal, in horses were reported infrequently. In the period 1996-2001, there was an increase in outbreaks of illness due to WN virus in animals as well as humans. Within the traditional range of WN virus, encephalitis was reported in horses in Italy in 1998 and in France in 2000. The first report of disease and deaths caused by WN virus infection in domestic birds was reported in Israel in 1997-1999, involving hundreds of young geese. In 1999 WN virus reached North America and caused an outbreak of encephalitis in humans in the New York area at the same time as a number of cases of equine encephalitis and deaths in American crows and a variety of other bird species, both North American natives and exotics. Multi-state surveillance for WN virus has been in place since April 2000 and has resulted in the detection of WN virus in thousands of dead birds from an increasing number of species in North America, and also in several species of mammals. The surveillance system that has developed in North America because of the utility of testing dead birds for the rapid detection of WN virus presence has been a unique integration of public health and wildlife health agencies. It has been suggested that the recent upsurge in clinical WN virus infection in wild and domestic animals as well as in humans may be related to the emergence of one or more new strains of WN virus. Virus isolated in New York in 1999 was found to be identical to that from Israel. It was alarming for WN virus to so easily invade the United States and surprising that it became established so quickly in the temperature climate of New York. Its persistence and rapid expansion in the United States leave a number of unanswered questions. New disease characteristics and patterns have occurred and more are evolving as WN virus further invades the western hemisphere. Additional animal research is needed to answer these questions. Some of the research needs include bird migration as a mechanism of virus dispersal, vector and vertebrate host relationships, virus persistence mechanisms, laboratory diagnosis, viral pathogenesis, risk factor studies, vaccine development, and WN virus impact on wildlife (CDC 2001a). Determination of the primary reservoir host species that are involved in the epidemiology of WN virus and the suitable sentinel species for active surveillance are also important research areas.

Arbovirus infection increases with group size.

Buggy Creek (BCR) virus is an arthropod-borne alphavirus that is naturally transmitted to its vertebrate host the cliff swallow (Petrochelidon pyrrhonota) by an invertebrate vector, namely the cimicid swallow bug (Oeciacus vicarius). We examined how the prevalence of the virus varied with the group size of both its vector and host. The study was conducted in southwestern Nebraska where cliff swallows breed in colonies ranging from one to 3700 nests and the bug populations at a site vary directly with the cliff swallow colony size. The percentage of cliff swallow nests containing bugs infected with BCR virus increased significantly with colony size at a site in the current year and at the site in the previous year. This result could not be explained by differences in the bug sampling methods, date of sampling, sample size of the bugs, age structure of the bugs or the presence of an alternate host, the house sparrow (Passer domesticus). Colony sites that were reused by cliff swallows showed a positive autocorrelation in the percentage of nests with infected bugs between year t and year t+1, but the spatial autocorrelation broke down for year t+2. The increased prevalence of BCR virus at larger cliff swallow colonies probably reflects the larger bug populations there, which are less likely to decline in size and lead to virus extinction. To the authors' knowledge this is the first demonstration of arbovirus infection increasing with group size and one of the few known predictive ecological relationships between an arbovirus and its vectors/hosts. The results have implications for both understanding the fitness consequences of coloniality for cliff swallows and understanding the temporal and spatial variation in arboviral epidemics.

West Nile virus in the United States: guidelines for detection, prevention, and control.

The epidemic/epizootic of West Nile (WN) encephalitis in the northeastern United States in the summer and fall of 1999 was an unprecedented event, underscoring the ease with which emerging infectious pathogens can be introduced into new geographic areas in today's era of rapid transportation and increased movement of people, animals, and commodities. This epidemic/epizootic and the increased frequency of other exotic pathogens being imported into the United States raises the issue of whether local, state, and national public health agencies are prepared to deal with epidemics/epizootics of vector-borne infectious diseases. The overwintering of WN virus and the epizootic transmission in the summer of 2000 reinforces the need to rebuild the public health infrastructure to deal with vector-borne diseases in this country. This article summarizes guidelines for surveillance, prevention, and control of WN virus that were drafted in December 1999 to help prepare state and local health departments for monitoring WN virus activity in the spring and summer of 2000 and also summarizes the data collected from those surveillance systems through September 2000.

West Nile virus surveillance using sentinel birds.

Captive and free-ranging birds have been used for decades as living sentinels in arbovirus surveillance programs. This review summarizes information relevant to selecting sentinel bird species for use in surveillance of West Nile (WN) virus. Although experience using avian sentinels for WN virus surveillance is limited, sentinels should be useful for both detecting and monitoring WN virus transmission; however, sentinel bird surveillance systems have yet to be adequately tested for use with the North American strain of WN virus. Captive chickens are typically used for arbovirus surveillance, but other captive species may be used as well. Serosurvey and experimental infection data suggest that both chickens and pigeons show promise as useful captive sentinels; both species were naturally exposed during the epizootics in New York City, 1999-2000, and both species develop antibodies after infection without becoming highly infectious to Culex pipiens vectors. Wild bird species that should be targeted for use as free-ranging sentinels include house sparrows and pigeons. The ideal wild bird should be determined locally on the basis of seroprevalence studies. Interpreting serological data generated from studies using free-ranging sentinel birds is complex, however. Sentinel bird monitoring sites should be selected in enzootic transmission foci. Several years of observation may be required for selection of effective sentinel monitoring sites.

Emergence of eastern encephalitis in Massachusetts.

The 20th century emergence in Massachusetts of zoonotic eastern encephalitis was interpreted in terms of recorded environmental change. The main mosquito vector of the infection, Cs. melanura, appears to have been scarce in eastern North America before the 1930s. Its relative scarcity resulted from destruction of the swamps that had been lumbered or drained for farming in the 18th and 19th centuries. When swamps matured once again early in the 1900s, the formation of subsurface pools of water beneath mature trees would have increased the availability of breeding sites for this mosquito. Transmission would have further been enhanced by the simultaneous proliferation of wetland-roosting robins and the extinction of such vagile birds as the passenger pigeon. Although numerous horses were maintained in Massachusetts at the time, no outbreaks of "equine sleeping sickness" came to public notice between the 1830s and the 1930s, when mature trees were scarce and the fauna was most disturbed. The severity of the first major outbreak in 1938 may have been potentiated by the absence of herd-immunity in a rapidly proliferating population of reservoir birds. These considerations suggest that recent landscape and faunal changes caused zoonotic EE to emerge in Massachusetts after waning for a century.

The economic burden imposed by a residual case of eastern encephalitis.

To estimate the economic burden imposed by eastern encephalitis (EE), we identified a series of residents of eastern Massachusetts who had survived EE infection and enumerated any costs that could be attributed to their experience. The records of three people who suffered only a transient episode of disease were analyzed as well as those of three who suffered severe residual sequelae. Transiently affected subjects mainly required assistance for direct medical services; the average total cost per case was $21,000. Those who suffered persistent sequelae remained at home and seemed likely to live a normal span of years, but without gainful employment. Early in the course of their chronic illness, costs ranged as high as $0.4 million per year, but plateaued at about $0.1 million after three years. Hospital costs, which dominated early in the disease experience, approached $0.3 million per patient. Educational costs tended to replace hospital costs after two years as the dominant economic burden and totaled about $0.3 million per patient during the first six years. Total costs then averaged almost $0.8 million. By the time that these subjects will have reached 22 years of age, disease-related costs will have totaled about $1.5 million. Institutionalization will impose an additional lifetime cost of $1.0 million. Insecticidal interventions designed to avert outbreaks of human EE infection cost between $0.7 million and $1.4 million, depending on the extent of the treated region. The direct costs of an intervention are less than the $3 million imposed on one person suffering residual sequelae of EE.

Eastern equine encephalitis virus in birds: relative competence of European starlings (Sturnus vulgaris).

To determine whether eastern equine encephalitis (EEE) virus infection in starlings may be more fulminant than in various native candidate reservoir birds, we compared their respective intensities and durations of viremia. Viremias are more intense and longer lasting in starlings than in robins and other birds. Starlings frequently die as their viremia begins to wane; other birds generally survive. Various Aedes as well as Culiseta melanura mosquitoes can acquire EEE viral infection from infected starlings under laboratory conditions. The reservoir competence of a bird is described as the product of infectiousness (proportion of feeding mosquitoes that become infected) and the duration of infectious viremia. Although starlings are not originally native where EEE is enzootic, a starling can infect about three times as many mosquitoes as can a robin.

High-dose i.v. contrast in CT scanning of the postoperative lumbar spine.

Evaluation of the postoperative lumbar spine is sometimes difficult because of obliteration of epidural fat by hypertrophic scarring. We examined 70 patients using a high-dose intravenous contrast technique in an attempt to distinguish hypertrophic scarring from herniated disk. The CT interpretation was confirmed in all 17 patients who had follow-up operations. Thirteen had herniated disk material associated with scar and four had hypertrophic scarring only. The latter four patients underwent reoperation because of concomitant foraminal or spinal canal stenosis seen on CT. Twelve of the herniated disks had the expected appearance of a nonenhancing mass surrounded by a rim of enhancing scar tissue. In the 13th patient, homogeneous enhancement of the herniated disk was seen. It is thought that chronically herniated disks, such as this one, may incite enough surrounding scar to give the CT appearance of an enhancing disk. Finally, marginal enhancement in the anulus fibrosus region was seen in over 90% of disk spaces examined. Although an anatomical explanation cannot be given at present, this phenomenon is thought to represent a normal finding.

Radiological techniques in the examination of patients with headaches.

It can be said that in the majority of headache patients who have a long typical history, a negative neurologic examination, and a noncontributory general physical examination, radiologic studies will probably not give further pertinent information. Nevertheless, in a sufficient number of these patients, the noninvasive and essentially atraumatic procedures of plain skull radiography, computed tomography, and (as indicated) echoencephalography and radionuclide brain scanning will provide a considerable amount of information pertinent to the clinical problem. Hospitalization for angiography and, on occasion, pneumoencephalography or other special procedures, will then become indicated for those patients in whom an abnormality is found in these screening procedures.

Bunyavirus infections in North Carolina white-tailed deer (Odocoileus virginianus).

We analyzed serum from white-tailed deer (Odocoileus virginianus) collected in southeastern North Carolina in 1991 for neutralizing antibodies to six mosquito-borne bunyaviruses (Lacrosse, Jamestown Canyon, Keystone,Cache Valley, Potosi, and Tensaw), including several of public health importance. Evidence was found for all six to be locally transmitted, although greatest seroprevalence was found for Potosi, Jamestown Canyon, and Cache Valley viruses.

Serologic evidence for West Nile virus infection in birds in Staten Island, New York, after an outbreak in 2000.

After an outbreak of West Nile virus (WNV) infections in people, horses, and wildlife in Staten Island, NY, during the summer of 2000, we surveyed the bird population of the island for evidence of infection. Neutralizing antibodies were detected in 59 of 257 (23.0%) resident birds and none of 96 transient (migrating) birds sampled in early October. Species with the greatest seroprevalence were northern cardinal (Cardinalis cardinalis) (69.2%) and rock dove (Columba livia) (54.5%). House sparrows (Passer domesticus) and chickens (Gallus gallus) had lower than expected seroprevalences, 8.6% and 5.5%, respectively. The geographic distribution of seropositivity suggested focal transmission at several locations on the island. The concentration of seropositive birds among resident bird populations on Staten Island supports the concept that many birds survive WNV infection and that some of these play an important role in the WNV-bird-mosquito transmission cycle.

Interrupted blood-feeding by Culiseta melanura (Diptera: Culicidae) on European starlings.

To determine whether Culiseta melanura (Coquillett) mosquitoes tend to take multiple blood meals when birds of certain species serve as hosts, we compared the frequencies with which such mosquitoes fed upon caged starlings and robins and determined whether similar volumes of blood were imbibed from each. The blood of robins (Turdus migratorius) and European starlings (Sturnus vulgaris) was marked contrastingly by injecting birds with rubidium or cesium salts. Caged birds were placed together in a natural wetland setting overnight. Mosquitoes captured nearby on the following morning were analyzed for each of the elemental markers. Where marked robins and starlings were equally abundant, 43% of freshly engorged Cs. melanura fed on more than or equal to two hosts. More Cs. melanura fed on robins than on starlings. Individual mosquitoes tended to contain far more robin- than starling-associated marker, indicating that mosquitoes "feasted" on robins but only "nibbled" on starlings. Mosquitoes marked with both elements apparently fed meagerly on the starlings then abundantly on the robins. Our estimates of bloodmeal volume indicate that 85% of mosquitoes that fed on marked starlings obtained < 0.5 microliter of blood from them. We suggest that defensive behavior by starlings interrupts mosquito blood-feeding and that, in a communal roost of starlings, each mosquito will tend to feed on more than one bird, thereby promoting rapid transmission of such ornithonotic arboviruses as eastern equine encephalomyelitis virus and West Nile virus.

Sensitive and specific colorimetric dot assay to detect eastern equine encephalomyelitis viral RNA in mosquitoes (Diptera: Culicidae) after polymerase chain reaction amplification.

A sensitive and specific colorimetric dot assay following polymerase chain reaction (PCR) method has been developed to detect 0.1 pg of eastern equine encephalomyelitis viral (EEEV) RNA. The assay is 250-fold more sensitive than analysis by electrophoresis and is based on converting a 291-nucleotide sequence of the viral coat protein amino terminus into a double-stranded DNA (dsDNA) and amplifying the DNA using a specific primer pair and PCR. The amplified complementary DNA (cDNA) is denatured adsorbed onto a nylon strip, baked, and detected with a digoxigenin-labeled probe. Dots with viral cDNA are stained dark red, whereas controls do not stain or stain lightly. The assay is very specific and sensitive and detects only EEEV. RNA of Venezuelan equine encephalitis, St. Louis encephalitis, Keystone, Flanders, Tensaw, and western equine encephalitis viruses were not detected. EEEV (Ten Broeck) RNA was detected at the 10-ng level, indicating that the prototype we used may have different nucleotides in the region where the primer pair binds. The PCR amplified EEEV cDNA that was 92% homologous to the consensus sequence of EEEV. The detection of EEEV in the liver of an infected Emu bird and in field-collected mosquitoes from Florida and Massachusetts that were analyzed concurrently as blind samples by tissue culture plaque assay and by PCR dot analysis proved that the assay is sensitive and can be used to detect infected mosquitoes. The assay can detect at least 1 infected mosquito in a pool of 1,000 uninfected mosquitoes.

Demonstration of acute post-viral myelitis with magnetic resonance imaging.

Post-infectious transverse myelitis is an important clinical entity in childhood, but the diagnosis is difficult to confirm by conventional radiologic investigations. We report a patient with acute, cervical, post-infectious transverse myelitis whose diagnosis had been established by the distribution and evolution of lesions on serial magnetic resonance imaging (MRI). Autopsy results confirmed the diagnosis of post-infectious transverse myelitis and the presence of MRI-identified lesions. To our knowledge, this patient is the first child reported with MRI findings in post-infectious transverse myelitis and suggests that MRI may be valuable in the diagnosis and management of children with this syndrome.

West Nile viral encephalitis.

West Nile virus (WNV) has emerged in recent years in temperate regions of Europe and North America, presenting a threat to both public and animal health. The most serious manifestation of infection is fatal encephalitis in humans and horses, as well as mortality in certain domestic and wild birds. A recent development in the epizootiology of this mosquito-borne flavivirus was the occurrence of a severe outbreak in New York City and surrounding areas. During this outbreak, mortality was observed in humans, horses, a cat and numerous species of wild birds, particularly members of the family Corvidae (crows). The author reviews basic information and summarises recent developments in the epidemiology and epizootiology of WNV.

Response to and efficacy of vaccination against eastern equine encephalomyelitis virus in emus.

OBJECTIVE: To evaluate humoral immune responses of emus vaccinated with commercially available equine polyvalent or experimental monovalent eastern equine encephalomyelitis (EEE) virus and western equine encephalomyelitis (WEE) virus vaccines and to determine whether vaccinated emus were protected against challenge with EEE virus. DESIGN: Cohort study. ANIMALS: 25 emus. PROCEDURE: Birds were randomly assigned to groups (n = 5/group) and vaccinated with 1 of 2 commercially available polyvalent equine vaccines, a monovalent EEE virus vaccine, or a monovalent WEE virus vaccine or were not vaccinated. Neutralizing antibody responses against EEE and WEE viruses were examined at regular intervals for up to 9 months. All emus vaccinated with the equine vaccines and 2 unvaccinated control birds were challenged with EEE virus. An additional unvaccinated bird was housed with the control birds to assess the possibility of contact transmission. RESULTS: All 4 vaccines induced detectable neutralizing antibody titers, and all birds vaccinated with the equine vaccines were fully protected against an otherwise lethal dose of EEE virus. Unvaccinated challenged birds developed viremia (> 10(9) plaque-forming units/ml of blood) and shed virus in feces, oral secretions, and regurgitated material. The unvaccinated pen-mate became infected in the absence of mosquito vectors, presumably as a result of direct virus transmission between birds. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that emus infected with EEE virus develop a high-titer viremia and suggest that they may serve as important virus reservoirs. Infected emus shed EEE virus in secretions and excretions, making them a direct hazard to pen-mates and attending humans. Commercially available polyvalent equine vaccines protect emus against EEE virus infection.

A nestable fiber pot for sampling resting mosquitoes.

To enhance the effectiveness of an arbovirus monitoring program, we evaluated a commercially available device for sampling resting vector mosquitoes. Diverse Anopheles, Culiseta, and Culex mosquitoes were taken in these nestable fiber pots. The pots sample about as many Culiseta melanura mosquitoes per device as do conventional resting boxes, but fewer than do boxes fitted with expanded frames. More Cs. melanura, and more bloodfed mosquitoes, but fewer species of mosquitoes are harvested with fiber pots than with CDC light traps. Fiber pots are more readily used, transported, and stored and are less expensive than conventional resting box devices or CDC light traps. A monitoring program based on the use of fiber pots, therefore, expends fewer resources than one using conventional resting boxes and collects about as many vector mosquitoes.