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Currently, there are increasing concerns about the possibility of a new epidemic due to emerging reports of Mayaro virus (MAYV) fever outbreaks in areas of South and Central America. Haemagogus mosquitoes, the primary sylvan vectors of MAYV are poorly characterized and a better understanding of the mosquito's viral transmission dynamics and interactions with MAYV and other microorganisms would be important in devising effective control strategies. In this study, a metatranscriptomic based approach was utilized to determine the prevalence of RNA viruses in field-caught mosquitoes morphologically identified as Haemagogus janthinomys from twelve (12) forest locations in Trinidad, West Indies. Known insect specific viruses including the Phasi Charoen-like and Humaiata-Tubiacanga virus dominated the virome of the mosquitoes throughout sampling locations while other viruses such as the avian leukosis virus, MAYV and several unclassified viruses had a narrower distribution. Additionally, assembled contigs from the Ecclesville location suggests the presence of a unique uncharacterized picorna-like virus. Mapping of RNA sequencing reads to reference mitochondrial sequences of potential feeding host animals showed hits against avian and rodent sequences, which putatively adds to the growing body of evidence of a potentially wide feeding host-range for the Haemagogus mosquito vector., (© 2021. The Author(s).)

Mayaro virus (MAYV), which causes mayaro fever, is endemic to limited regions of South America that may expand due to the possible involvement of Aedes spp. mosquitoes in its transmission. Its effective control will require the accurate identification of infected individuals, which has been restricted to nucleic acid-based tests due to similarities with other emerging members of the Alphavirus genus of the Togaviridae family; both in structure and clinical symptoms. Serological tests have a more significant potential to expand testing at a reasonable cost, and their performance primarily reflects that of the antigen utilized to capture pathogen-specific antibodies. Here, we describe the assembly of a synthetic gene encoding multiple copies of antigenic determinants mapped from the nsP1, nsP2, E1, and E2 proteins of MAYV that readily expressed as a stable chimeric protein in bacteria. Its serological performance as the target in ELISAs revealed a high accuracy for detecting anti-MAYV IgM antibodies. No cross-reactivity was observed with serum from seropositive individuals for dengue, chikungunya, yellow fever, Zika, and other infectious diseases as well as healthy individuals. Our data suggest that this bioengineered antigen could be used to develop high-performance serological tests for MAYV infections., (© 2021. The Author(s).)

The objective of this study was to characterize the prevalence of viral encephalitis due to arbovirus infection of the Togaviridae and Flaviviridae families in São Paulo, Brazil. A total of 500 cerebrospinal fluid (CSF) samples collected between August 2012 and January 2013, from patients with symptoms of acute encephalitis were analyzed. Findings suggestive of viral encephalitis-elevations in cell concentration, glucose and total protein-were observed in 234 (46.8%) samples, designated as Group 1. The remaining 266 samples comprised Group 2. All samples were tested for Flaviviruses (dengue virus 1, 2, 3 and 4, yellow fever virus and West Nile virus), Alphavirus (NS5 region) and enterovirus by RT- PCR and for herpesviruses and enteroviruses using CLART-Entherpex. A presumptive viral etiological agent was detected in 26 samples (5.2%), 18 (8.0%) in Group 1 and 8 (3.0%) in Group 2. In Group 1 human herpesviruses were detected in 9 cases, enteroviruses in 7 cases, dengue viruses (DENV) in 2 CSFs and St. Louis encephalitis virus (SLEV) in one case. In Group 2 there were 3 CSFs positive for human herpesviruses, 2 for enteroviruses, 2 for DENV and 1 for SLEV. Detection of arboviruses, even though present in a minority of infected patients, identifies these viruses as a probable etiological agent of encephalitis. This is of special concern in regions where this class of viruses is endemic and has been linked to other recent epidemics., Competing Interests: The authors have declared that no competing interests exist.

Rio Negro virophage (RNV) was co-isolated with a strain of mimivirus named sambavirus, from Brazilian Amazon. We report the near complete genome sequence of RNV, the first virophage isolated in Brazil. We also present new microscopical data demonstrating that RNV particles have similar dimensions to that described to sputnik virophages., (Copyright © 2018 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.)

Pancreas disease (PD) in salmonid fish is caused by an infection with Salmonid alphavirus (SAV) and remains as one of the major health problems in the European fish farming industry. Sequence studies have revealed a genetic diversity among viral strains. A subtype of SAV (SAV3) is causing an epizootic in farmed salmonids in Norway. Here we evaluate efficacy and safety of an inactivated virus vaccine based on ALV405, a strain of SAV3 that was isolated from Norwegian salmon. The vaccine provided an average relative percent survival (RPS) of 98.5 in an intraperitoneal challenge model, and induced nearly total protection against PD in a cohabitant challenge model. It provided significant protection against SAV-induced mortality also in a field trial under industrial conditions. Local reactions seen as melanization and adhesions in the visceral cavity were less severe than those induced by two commercial vaccines. Finally, we demonstrated that the protection is not impaired when the ALV405 antigen is combined with other viral or bacterial antigens in a polyvalent vaccine. The results confirm that efficient and safe protection against SAV infection and development of PD is possible using an inactivated virus vaccine, both alone and as a component in a polyvalent vaccine., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Wild birds are rarely found with active arbovirus infections, and relatively little is known about the patterns of viremia they exhibit under field conditions or how infection varies with date, bird age, or other factors that potentially affect transmission dynamics. Buggy Creek virus (BCRV; Togaviridae, Alphavirus) is an arbovirus associated with colonially nesting Cliff Swallows (Petrochelidon pyrrhonota) and transmitted by its vector, the hematophagous swallow bug (Oeciacus vicarius), an ectoparasite of the Cliff Swallow. Introduced House Sparrows (Passer domesticus) that have occupied swallow nests at colony sites in peridomestic settings are also exposed to BCRV when fed upon by swallow bugs. We used data from 882 nestling House Sparrows in western Nebraska from 2006 to 2008 to examine seasonal variation and age-related correlates of virus infection in the field. Over 17% of nestling House Sparrows had active infections. Prevalence was higher in 2007 than in 2008 when birds from all colony sites were analyzed, but there was no significant difference between years for sites sampled in both seasons. Buggy Creek virus prevalence was similar in early and late summer, with a peak in midsummer, coinciding with the greatest swallow bug abundance. Nestlings 10 days of age and younger were most commonly infected, and the likelihood of BCRV infection declined for older nestlings. Average viremia titers also declined with age (but did not vary with date) and were high enough at all nestling ages to likely infect blood-feeding arthropods (swallow bugs). Length of viremia for nestlings in the field was ≥4 days, in agreement with an earlier study of BCRV. Nestling birds offer many advantages for field studies of arbovirus amplification and transmission.

In February 2008, a Mayaro fever virus (MAYV) outbreak occurred in a settlement in Santa Barbara municipality, northern Brazil. Patients had rash, fever, and severe arthralgia lasting up to 7 days. Immunoglobulin M against MAYV was detected by ELISA in 36 persons; 3 MAYV isolates sequenced were characterized as genotype D.

The number of mosquito-borne viruses ('moboviruses') occurring in Europe since the twentieth century now stands at ten; they belong to three families-Togaviridae (Sindbis, Chikungunya), Flaviviridae (West Nile, Usutu, Dengue), and Bunyaviridae (Batai, Tahyna, Snowshoe hare, Inkoo, Lednice). Several of them play a definite role in human or animal pathology (Sindbis, Chikungunya, Dengue, West Nile, Tahyna). Mobovirus outbreaks are strictly determined by the presence and/or import of particular competent vectors of the disease. Ecological variables affect moboviruses considerably; the main factors are population density of mosquito vectors and their vertebrate hosts, intense summer precipitations or floods, summer temperatures and drought, and presence of appropriate habitats, e.g., wetlands, small water pools, or intravillan sewage systems. A surveillance for moboviruses and the diseases they cause in Europe is recommendable, because the cases may often pass unnoticed or misdiagnosed not only in free-living vertebrates but also in domestic animals and even in humans.

Buggy Creek virus (BCRV) is an unusual arbovirus within the western equine encephalitis complex of alphaviruses. Associated with cimicid swallow bugs (Oeciacus vicarius) as its vector and the cliff swallow (Petrochelidon pyrrhonota) and house sparrow (Passer domesticus) as its amplifying hosts, this virus is found primarily in the western Great Plains of North America at spatially discrete swallow nesting colonies. For 342 isolates collected in Oklahoma, Nebraska, Colorado and North Dakota, from 1974 to 2007, we sequenced a 2076 bp region of the 26S subgenomic RNA structural glycoprotein coding region, and analysed phylogenetic relationships, rates of evolution, demographical histories and temporal genetic structure of the two BCRV lineages found in the Great Plains. The two lineages showed distinct phylogeographical structure: one lineage was found in the southern Great Plains and the other in the northern Great Plains, and both occurred in Nebraska and Colorado. Within each lineage, there was additional latitudinal division into three distinct sublineages. One lineage is showing a long-term population decline. In comparing sequences taken from the same sites 8-30 years apart, in one case one lineage had been replaced by the other, and in the other cases there was little evidence of the same haplotypes persisting over time. The evolutionary rate of BCRV is in the order of 1.6-3.6x10(-4) substitutions per site per year, similar to that estimated for other temperate-latitude alphaviruses. The phylogeography and evolution of BCRV could be better understood once we determine the nature of the ecological differences between the lineages.

Atlantic salmon Salmo salar pre-smolt, smolt and post-smolt, with clinical signs of haemorrhagic smolt syndrome (HSS) have been found in several locations along the Norwegian coast (Rogaland to Troms). Affected fish had pale gills and bleeding at the fin bases, but seemed to be in good physical condition with no obvious weight loss. The internal organs and body cavity showed distinct bleedings. Petechiae were found on the gastrointestinal tract, swim bladder and peritoneum, visceral adipose tissue, heart and somatic musculature. The liver was bright yellow and sometimes mottled with petechiae and ecchymoses. Acitic fluid was found in the visceral cavity and fluid was also present in the pericardial cavity. Histological examination revealed haemorrhage in most organs. The glomeruli were degenerated and the renal tubules were filled with erythrocytes. The aims of this study were to describe the pathology and discover, if possible, the aetiology of the HSS. Tissues were collected for light and transmission electron microscopy (TEM), immunofluorescence (IFAT), reverse transcription (RT)-PCR diagnostics (screening for infectious salmon anaemia virus [ISAV], viral haemorrhagic septicaemia virus [VHSV], salmon pancreas disease virus [SPDV], sleeping disease virus [SDV] and infectious haematopoetic necrosis virus [IHNV]), and tissue homogenates (heart, liver, kidney and spleen) were sterile-filtered and inoculated into cell cultures. Homogenates made from several tissues were also injected intraperitoneally into salmon and rainbow trout Oncorhynchus mykiss. The diagnostic tests revealed no consistent findings of any pathogens, with the exception of TEM which showed 2 types of virus-like particles: Type I was 50 to 60 nm in diameter and Type II about 50 nm in diameter. These virus-like particles were found in salmon from all farms affected by HSS and screened by TEM. Several different cells, blood vessel endothelial cells, endocardial cells, heart myofibres, and leukocytes were associated with the 2 virus-like particles. The Type I particle seems to be an infectious pancreatic necrosis (IPN)-like virus, while (based on the number of target cells, particle morphology, budding and uptake into target cells) Type II particle could be a togavirus.

Two viruses, infectious salmon anaemia (ISA) virus and a novel togavirus-like virus, were isolated from ISA disease outbreaks that were first reported as a new syndrome, haemorrhagic kidney syndrome (HKS) affecting farmed Atlantic salmon Salmo salar L. on the East coast of Canada. Laboratory confirmation of ISA diagnosis was initially complicated by isolation of only the togavirus-like agent using the CHSE-214 cell line. Here we demonstrate that a clinical sample from a disease outbreak of ISA contained a mixture of ISA virus and togavirus-like virus. Reverse transcriptase-polymerase chain reaction (RT-PCR) confirmed the presence of both viruses during serial passage of cultures in SHK-1 and CHSE-214 cells. Virus harvested at passage level 3 in both cell lines caused high mortalities and severe gross pathology consistent with ISA virus infection in experimentally inoculated Atlantic salmon parr (approximately 35 g) in freshwater, beginning 12 d post inoculation. ISA virus was detected by virus isolation from kidney and liver tissues of all dead or moribund fish tested. A comparison of virus isolation, 1-step procedure RT-PCR and RNA dot-blot hybridization for detection of ISA virus (ISAV) in fish tissues showed virus isolation to have 100% sensitivity, followed by RT-PCR (66 and 28% sensitivity in kidney and liver, respectively), with RNA dot-blot hybridization as the least sensitive method (20 and 10% sensitivity in kidney and liver, respectively). No togavirus-like virus was detected in these samples by virus isolation. Moreover, another togavirus-like virus isolate grown in CHSE-214 cells in the absence of any other detectable pathogen was non-pathogenic in experimentally inoculated fish. This study confirms that the original ISA outbreaks in New Brunswick, Canada, were caused solely by ISAV.

Objective: To isolate arbovirus from mosquitoes caught from Yantai., Methods: The isolated viruses were tested for their physico- chemical properties, examined by electron microscopy and specific immuno-reactivity., Results: Fifteen strains of virus were isolated from mosquitoes in 1994 from Yantai. Three of them were further assayed. The results showed that the viruses could multiply on C6/36 cell and produce typical cytopathogenic effect. The viruses couldn't cause regular sickness and death of suckling mice by intracerebral inoculation. The viruses were sensitive to ether, but resistant to 5, -Idu. One sample was examined with electron microscope, and spherical virus particles were observed. The diameter of the virus particles is about 55 +/- 2. 3nm. It did not react with the JBE virus and Bunyaviridae group specific immuno-ascitic fluids but cross-reacted with the group A Togaviridae specific immuno-ascitic fluid., Conclusions: The results showed that the viruses isolated from Yantai belong to alphavirus of togaviridae.

A selected number of PCR protocols were evaluated to determine if they could serve as a universal protocol for detecting and identifying all arboviruses. In this study, four parameters that affect the efficacy of RT-PCR (RNA extraction method, choice of reverse transcriptase, choice of DNA polymerase and thermocycling program) were evaluated in combination. The most optimal combination of those parameters employed use of silica gel membrane spin column, RAV-2 reverse transcriptase, Tth DNA polymerase, and a simple modification of a published thermocycling program. By this modified protocol, viral RNA could be amplified satisfactorily with more than 50 pairs of primers designed for diagnosis of arboviruses representing five families. The sensitivity and specificity obtained by this universal protocol were comparable to those obtained by the original protocol for each primer pair tested; and for some primers, improved sensitivity was observed. It was also found that a simple modification of a suggested protocol of a commercial RT-PCR kit could produce nearly identical results and serve as another universal protocol. With the use of a universal diagnostic reverse transcriptase-polymerase chain reaction (RT-PCR) protocol, simultaneous screening of clinical or biological specimens against a large number of RNA viruses belonging to many families can be performed more efficiently for etiologic determination in the situations complicated by the difficulty of differential diagnosis. Furthermore, such a universal protocol facilitates reducing the cost of PCR-based diagnostic operation and standardizing the qualities of PCR-based diagnosis within an institution or among collaborating institutions. A logical strategy is to conduct diagnosis in two stages by using broadly group-reactive primers in the first stage to narrow the range of possible etiologic agents and using virus-specific primers in the second stage for identification. Before such a strategy is employed, however, more group-reactive primers for a large number of arboviruses, for which no such primers currently exist, must be made available. Furthermore, the best pair or pairs of primers need to be selected for each virus for the second stage of the strategy.

A virus inducing a cytopathic effect on porcine alveolar macrophages was isolated from the lungs of a pig with respiratory problems and lesions of proliferative and necrotizing pneumonia. The isolate was found to react with porcine reproductive and respiratory syndrome virus (PRRSV) monoclonal antibodies by indirect immunofluorescence and was designated LHVA-93-3. The virus could also be propagated on the MARC-145 cell line. The LHVA-93-3 macrophage-passaged isolate was inoculated orally or intranasally in four-week-old specific pathogen-free pigs. Histologically, focal to multifocal lesions of proliferative, necrotizing and interstitial pneumonia could be observed in the lungs of pigs inoculated orally or intranasally, 6 and 10 days post-inoculation. Virus could be reisolated from essentially the same tissues including serum following both routes of infection. The distribution of PRRSV antigens in fixed tissues as determined by immunogold silver staining (IGSS) was similar in orally or intranasally inoculated pigs. The results of this experimental transmission study indicate that pigs may become infected by PRRSV following oral as well as intranasal exposure.

A sensitive assay based on the polymerase chain reaction for the detection of Ockelbo virus RNA was developed. Two primer pairs from the gene coding for the E2 glycoprotein were chosen. By use of a nested strategy for the primers, as few as 1 to 10 PFU could be detected. The amplified products were visualized as bands of appropriate size on ethidium bromide-stained agarose gels. The primer pairs allowed amplification of several Ockelbo and Sindbis virus isolates but discriminated between these and other alphaviruses. Ockelbo virus RNA was detected in 4 of 10 skin biopsy specimens collected during the acute stage of the disease. The identities of the amplified products were confirmed by restriction endonuclease cleavage. Acute- and convalescent-phase sera as well as lymphocytes collected during the convalescent phase were negative by the polymerase chain reaction. No infectious virus could be recovered from any of the specimens.

An indirect enzyme-linked immunosorbent assay was developed for rapid detection of serum antibodies against the virus responsible for the porcine reproductive and respiratory syndrome (PRRS). This test is more sensitive than the reference technique (immunoperoxidase test applied to cultures of alveolar macrophages), particularly for detecting animals at the stage of seroconversion. It is also very specific for PRRS virus, because all specific hyperimmune sera against other porcine viruses, and all serum samples taken from herds before the disease appeared in western France were negative. The test has been used for routine diagnosis of PRRS. The results obtained during nine months from over 21,000 samples have confirmed the value of the test for diagnosis and epidemiological surveillance of the disease.

Toga virus-like particles (typically 60-70 nm: enveloped with small surface spikes) were detected in the native hepatectomy specimens in 7 of 18 patients grafted for acute liver failure attributed to sporadic non-A, non-B hepatitis and in 2 patients grafted for fulminant hepatitis attributed to anti-epileptic drug hepatotoxicity. These particles were not detected in the hepatectomies from 12 other patients grafted for other causes of acute liver failure, 12 for various chronic liver diseases, and 2 histologically normal livers. Acute hepatic failure, characterized histologically by severe haemorrhagic necrosis, developed 7 days after grafting in 5 patients, all in the non-A, non-B group with toga virus-like particles in native liver. Similar virus-like particles were detected in all grafts and were in greater abundance than in the native livers. The agent may be novel because pre- and post-grafting sera were negative for antibodies against representative panels of arboviruses and in first and second generation antibody tests for hepatitis C virus.

During a 1986 natural lethal outbreak of fulminating disease in guinea poult flocks in southwestern France, enveloped virus particles were consistently observed in the gut contents of infected birds. For the present study, a protocol was developed for the purification of these particles. Sucrose-banded virus obtained from birds infected experimentally with virus from the outbreak was found to have a buoyant density of 1.18 g/ml. The purified virus showed hemagglutinating activity, was shown by electron microscopy to have a togavirus-like morphology, and also was shown to be transmissible and pathogenic through oral ingestion. In addition, other enveloped particles have been occasionally detected in gut contents of both infected and uninfected birds; the improbability of the viral nature of these interfering particles is discussed.

We describe the first known occurrence in France of the porcine reproductive and respiratory syndrome (PRRS), a new porcine disease that first appeared in Germany in November 1990. Outbreaks of the disease appeared in November 1991 in Brittany (France), with comparable clinical features to those observed in other countries, were definitively confirmed by a serological analysis from the affected animals using an immunoperoxidase monolayer assay with PRRS-virus infected alveolar macrophages. Furthermore, we report the isolation from one serum of a filtrable agent that, in view of its cytopathic effect for porcine alveolar macrophages and of the serologic reactivity of the infected cells, appears similar to the Lelystad virus that has been implicated in the etiology of the disease.

The peroxidase-antiperoxidase technique was used to determine the cellular localization of Border Disease (BD) virus in cryostat sections of fetal and newborn lamb spinal cord following experimental infection by maternal inoculation in early gestation. Viraemic fetuses and lambs with hypomyelinogenesis showed BD viral antigen in neurons, glia, ependymal cells, vascular endothelial cells and fibrocytes within the dura mater. Double immunolabelling demonstrated co-expression of BD viral antigen and glial fibrillary acidic protein (GFAP) or myelin basic protein (MBP) in both fetal and newborn lamb glia. In fetal lambs there was a pia-associated population of glia in which viral antigen was also co-expressed with GFAP or MBP. The results suggest that BD virus infects myelinating oligodendroglia, astroglia and probably also transitional cells and pluripotential glioblasts. The relationship between infection of specific cell types and hypomyelinogenesis was not resolved but infection of transitional cells and oligodendroglia may affect oligodendroglial function and permit morphologically inapparent perturbations leading to hypomyelinogenesis. A single nonviraemic lamb with a precolostral antibody titre to BD virus and cystic cerebral cavities but no hypomyelinogenesis showed BD viral antigen confined to glia of the spinal cord white matter. This suggests that oligodendroglia may require to be infected before a critical period in their development or factors additional to oligodendroglia infection are necessary for hypomyelinogenesis.

Fourteen sheep persistently infected with border disease virus were investigated to examine the effects of persistent viraemia on lymphocyte subpopulations in peripheral blood and their responses to the mitogen phytohaemagglutinin (PHA) in vitro. Persistently infected sheep had significantly more CD8+ (cytotoxic-suppressor) T-lymphocytes than uninfected sheep of the same age (P less than 0.001). The total number of CD4+ (helper) T-lymphocytes were not significantly different but there were more T-lymphocytes (CD5+) which were CD4- and CD8- in normal sheep than persistently infected sheep (P less than 0.001). Peripheral lymphocytes obtained from persistently infected sheep showed significantly reduced blastogenesis induced by PHA than those obtained from normal sheep (P less than 0.001).

C6/36 Aedes albopictus cells were infected with Brazilian arbovirus from the families Togaviridae, Flaviviridae, Bunyaviridae and Rhabdoviridae. Replication was obtained with all the studied viruses and cytopathic effect was observed with some. Viral antigen was assayed in C6/36 cell cultures for antigen was assayed in C6/36 cells by an indirect immunofluorescence test using specific mouse immune ascitic fluid. Antigen production was detected in C6/36 cells infected with all the studied viruses. The author recommends the inoculation of C6/36 cell cultures for isolation of virus from the four studied families. The immunofluorescence technique is an important tool for classification and identification of virus growing in C6/36 cells.

Over 30 arboviruses belonging to the families Togaviridae (2), Rhabdoviridae (2), Reoviridae (more than 10), Bunyaviridae (17) and non-classified ones (about 4) were isolated from midges. The distribution area of some viruses, which are of great medical and veterinary importance, is located in immediate proximity to southern borders of the USSR. Biological and ecological peculiarities of midges in addition to the above data on the isolation of viruses define the perspectives of virusological examination of midges, primarily in Transcaucasia, Middle Asia, southern regions of Siberia and Far East of the USSR.

Virus-like particles (60-70 nm) with spiked surfaces budding into cell vacuoles and rod-shaped inclusions were detected in nuclei of hepatocytes from a British patient transplanted for sporadic non-A, non-B fulminant hepatitis (NANB-FHF), probably contracted in Kenya. Identical particles were seen in two successive grafts (days 2 and 10) at regrafting for recurrent FHF. Ultrastructural features resembled those of the RNA-containing arbovirus, Rift Valley fever virus, but serological markers against a representative panel for arboviruses (Togaviruses) and transmission in mice proved negative. The particles shared features with the different arboviruses seen in the hepatectomy specimen of a second patient with NANB-FHF, and in both patients an insect vector was implicated in the clinical history. The particles were identical in size to those of a third patient with NANB-FHF, who had remained in the United Kingdom. These findings, together with the recent report of isolation of an RNA-containing virus resembling the Togaviridae, in parenteral NANB, suggest that several exotic virus-like agents resembling the arboviruses may be involved in the aetiology of NANB, including in the sporadic forms of FHF in the United Kingdom.

Yellow fever, dengue (types 1, 2 and 4), Chikungunya, Rift Valley fever, Ebola, Marburg, and Lassa viruses were inoculated into susceptible cell cultures and daily investigated by indirect immunofluorescence (IFA) and electron microscopy (EM) with a view to achieve an early detection-identification of these agents. Compared to the other cell lines tested (Vero, BHK-21 and Aedes albopictus), CV-1 cells were found to be more sensitive. Viral antigens were detected by IFA from a few hours post inoculation (CHIK and RVF) to a maximum of 3 days (YF and EBO). For most of the viruses studied, the cytopathic effect (CPE) commenced 2-3 days after the detection of viral antigens. Virus particles were detected by EM only in the case of EBO, MBG and LAS, before any CPE was observed in cell cultures.

Lambs congenitally infected with border disease (BD) virus and sheep exposed to BD virus as adults were studied for one year to determine the pathogenesis of congenital exposure compared with adult exposure to the virus. Persistent BD virus was isolated in tissue culture and detected by immunofluorescence of the peripheral white blood cells, urine, and cerebrospinal fluid of lambs with congenital BD up to one year of age. These animals had no detectable serum neutralizing antibody response to the virus for the same interval. BD virus antigen was also detected by immunofluorescence in many central nervous system tissues of these lambs with congenital BD. Dysmyelination and glial proliferation in the central nervous system and microencephaly were noted in the lambs with congenital BD, and these lesions appeared to remain the same over a 12-month period.

The successful application of the immunoperoxidase method of (IP) with light microscopy for rapid detection of certain togaviruses and orthopoxviruses and antibodies to them is described. A comparative evaluation of the IP and immunofluorescent (IF) method for these purposes was performed. The results indicate practically similar sensitivity and specificity of both methods. The detection of viral antigen directly depended upon the infective dose and time of examinations (within 4--12 hours postinoculation). In evaluating the IP method, however, the account should be taken of the fact that it does not require expensive equipment and therefore may be used in any practical laboratory or even under field conditions. Rapid detection of both viral antigens and antiviral antibody by this method opens approaches for its wide application in the future.

Negative staining electron microscopy was used to identify viruses in 166 normal and 62 diarrhoeal faecal samples from 208 cats admitted to an animal shelter during a 16-month period (March 1984 to June 1985). On the basis of size and shape 7 distinct viral types were detected: 24 nm parvovirus-like particles, 30 nm astrovirus, 30 nm picornavirus-like particles, reovirus, rotavirus, coronavirus and a 75 nm "togavirus-like" particle. The incidence of these particles in the 208 cats was 11%, 7%, 6%, 0.4%, 5%, 1% and 1% respectively. Virus isolation studies using 40 of the faecal samples succeeded in isolating reovirus 1 in 2 cases. Immune electron microscope studies demonstrated the presence of antibody in a human serum to cat astrovirus, but failed to clarify the identity of the parvovirus-like particles and picornavirus-like particles, other than showing that some of the parvovirus-like particles were not related to feline panleukopenia virus. It was found that parvovirus-like particles, astrovirus, picornavirus-like particles, reovirus and rotavirus could be excreted by cats with normal faeces as well as cats with diarrhoeal faeces. Parvovirus-like particles, astrovirus, picornavirus-like particles and rotavirus could be excreted in high concentration in normal faeces. There was no simple relationship between age and diarrhoea in the population of cats studied. Age was not a critical factor in the excretion of parvovirus-like particles, astrovirus, picornavirus-like particles and rotavirus. The incidence of diarrhoea was not clearly associated with the seasons.

We studied the transmission of Fort Morgan (FM) virus within colonies of nesting Cliff Swallows and House Sparrows under three bridges in Morgan County, Colorado during 1976. Nests were examined, and blood or brain specimens were collected from nestlings once or twice a week. Flying birds and small mammals were also studied. We analyzed nesting activity, virus isolations from nestlings of both species, fledging success, multiple infections within a brood of nestlings, infection frequency by age of nestlings, nestling mortality, and infection frequencies by avian species and bridge site. Fort Morgan virus was isolated from 7% (80/1, 156) of the blood and brain samples collected from nestlings. The duration of viremia for nestling House Sparrows was at least 3-4 days based on virus isolation from sequential blood samples. Viremia of nestling Cliff Swallows and House Sparrows did not reduce fledging success, nor were young nestling sparrows viremic more frequently than older nestling sparrows. Nest destruction (by falling down) was a more important cause of nestling mortality than FM virus infection. All age groups of nestling sparrows were viremic at equal rates, but younger nestlings (less than or equal to 7 days old) were more likely than older nestlings (greater than 7 days old) to develop an encephalitic infection. Among nestling House Sparrows, FM virus infections were clustered in time and space. Nestling House Sparrows with FM virus-infected nest-mates were infected more often than conspecifics whose nest-mates were not infected. We concluded that nestling Cliff Swallows and symbiotic House Sparrows that reside in swallow nesting colonies are the principal vertebrate hosts for the maintenance and amplification of FM virus.

Virus-like particles (VLP), morphologically similar to togavirus particles, were detected in the sera of chimpanzees and humans with chronic non-A, non-B (NANB) hepatitis. Particles resembling the virus were also found in the cytoplasm of hepatocytes obtained from NANB hepatitis infected chimpanzees. Furthermore, two chimpanzees inoculated with materials containing the VLP developed NANB hepatitis. These results strongly suggest that the VLP may be closely associated with the agent of NANB hepatitis.

The prevalence of border disease virus in a flock of Targhee sheep in southern Idaho was determined by serologic and virologic techniques. Of 249 ewes, 73 (29%) were seropositive to the antigenically related togavirus, bovine seropositive to the antigenically related togavirus, bovine viral diarrhea virus (BVDV). Concurrent testing of sera from 337 neonatal lambs indicated that 172 (51%) were seropositive for BVDV. It was observed that a high percentage (35%) of the BVDV-seropositive lambs were derived from seronegative ewes. Immunotolerance of the ewe was considered as the explanation for the lack of serologic relationship between seropositive lambs and their seronegative dams. Eleven cytopathogenic viruses were isolated from nasal and vaginal specimens obtained from 80 clinically healthy ewes within 24 hours of parturition. The viral isolates were demonstrated to be antigenically related to BVDV by direct immunofluorescence. Six of the 11 isolates were obtained from BVDV-seronegative ewes.

A new strain of simian haemorrhagic fever (SHF) virus was isolated from chronically infected patas monkey no. 248 (P-248) in USU-104 cells. The P-248 isolate had the same size, morphology and cytoplasmic site of replication as the prototype LVR strain. However, the P-248 isolate caused a persistent infection without noticeable cytopathology in USU-104 cells rather than the strongly lytic infection produced by prototype LVR virus. The capacity of P-248 virus to produce a persistent, non-lytic infection of USU-104 cells was a very stable characteristic of the isolate. Extensive serial passage of this isolate through USU-104 cells (over 50 passages) and rhesus monkeys (six passages) failed to unmask virus with lytic properties for USU-104 cells. Culture medium from persistently infected cultures assayed in rhesus monkey peritoneal mononuclear phagocytes, where measurable cytopathology occurs, was found to contain about 10(5) to 10(6) TCID50/ml of cell-free P-248 virus. Immunolabelling techniques showed only a low percentage of infected cells in persistently infected cultures. The mechanism of persistence of the P-248 isolate in USU-104 cells has not been determined but evidence suggests it does not involve interferon or defective interfering particles.

The authors make a review of literature on viruses isolated from chiroptera's tissues. They study especially two of these viruses: a true arbovirus, the Japanese B encephalitis virus and a virus of the same genus but not arthropod-borne, the Rio Bravo virus. They consider successively for each virus: isolations from bats, serosurveys in bat and in man, and experimental studies with these viruses, allowing to estimate the role of chiroptera in the diffusion and the maintenance of some arboviruses. They also report some personal results of serosurveys in bats from Tunisia and Spain.