Detection of antibody to avian viruses in human populations.

The ability of three avian viruses to elicit antibody response in humans was surveyed for the purpose of identifying zoonotic diseases. Antibody levels in people associated with poultry were compared to those in people having limited poultry association. Antibody levels to three avian viruses: infectious bursal disease virus, a birnavirus; Newcastle disease virus, a paramyxovirus; and avian infectious bronchitis virus, a coronavirus were determined by enzyme-linked immunosorbent assays (ELISA). Differences between the two study groups were evident: people having a known association with poultry showed significantly higher levels of antibodies to Newcastle disease and avian infectious bronchitis virus. Antibodies detected may be due to virus exposure rather than zoonoses.

Use of egg yolk to determine antibody levels in chickens inoculated with a hemagglutinating duck adenovirus (adenovirus 127-like).

Chickens were experimentally infected with a duck adenovirus that has been shown to be serologically indistinguishable from Adenovirus 127. Sera and eggs were collected at intervals after exposure for antibody determination by the hemagglutination-inhibition (HI) test, the enzyme-linked immunosorbent assay (ELISA), and the immunodiffusion (ID) test. Egg yolks were processed for use in the serological tests by (a) dilution in phosphate-buffered saline (PBS), (b) extraction of the water-soluble fraction with chloroform, or (c) freezing and thawing PBS-diluted yolks and testing the supernatant fluid. HI antibody titers from serum and extracted yolk were similar except during the initial 2 weeks, when yolk antibody levels were low or absent. Chloroform-extracted yolks were suitable material for the HI, ELISA, and ID tests. Heat inactivation of the chloroform-extracted yolk had no effect on titers.

Use of egg yolk in serological tests (ELISA and HI) to detect antibody to Newcastle disease, infectious bronchitis, and Mycoplasma gallisepticum.

Serum and yolks from commercial flocks and from hens exposed to Newcastle disease virus (NDV), infectious bronchitis virus (IBV), and Mycoplasma gallisepticum (MG) were tested for immunoglobulin G antibody by the enzyme-linked immunosorbent assay (ELISA) and the hemagglutination-inhibition (HI) test. Yolks prepared by chloroform extraction and low-speed centrifugation performed well in the serological tests used and were a suitable alternative to serum for antibody determination by the ELISA for NDV, IBV, and MG and by HI test for NDV.

Evidence of exposure of waterfowl and other aquatic birds to the hemagglutinating duck adenovirus identical to EDS-76 virus.

Serum and fecal samples from 12 species of aquatic birds were studied for evidence of exposure to a hemagglutinating duck adenovirus (DAV). DAV is serologically indistinguishable from egg-drop syndrome-76 virus. A total of 285 serum samples were tested by the hemagglutination-inhibition (HI) test. Forty-two percent of the birds had HI antibodies, with titers ranging from 8 to 256. Wild ducks showed the highest frequency of antibodies (56%) while in coots and grebes, antibody was less frequent, 33% and 26%, respectively. Attempted virus isolations from 79 fecal samples were unsuccessful. The data support the hypothesis that DAV is indigenous in wild duck populations and suggest that infection and viremia are limited in time and occur at a very early age.

Comparison of enzyme-linked immunosorbent assay with hemagglutination-inhibition and immunodiffusion tests for detection of antibodies to a hemagglutinating duck adenovirus in chickens.

An indirect enzyme-linked immunosorbent assay (ELISA) for the detection of antibody to a hemagglutinating duck adenovirus is described. Optimum conditions for the test were determined, and the system was compared with procedures currently used. Experimentally infected chickens were assayed for specific antibody by ELISA, hemagglutination inhibition (HI), and immunodiffusion (ID). The ELISA was a sensitive and reliable method for detecting antibody, although positive titers were not always in agreement with HI and ID results at 1 week postinoculation, probably reflecting the different classes of antibody being detected.

Is avian adeno-associated virus an endogenous virus of chicken cells?

The adeno-associated viruses (AAV) are defective parvoviruses which produce infective progeny only in cells co-infected with a 'helper' adenovirus (Ad). Both human and simian AAV have been recovered from human and simian primary cell cultures following their inoculation with 'AAV-free' Ad. Whereas some studies have suggested that AAV exists in a latent state in these cells, others have indicated that the AAV genome is capable of establishing and maintaining a latent state in defined laboratory conditions which mimic the situation proposed for the 'latent' AAV recovered from human and simian tissues. Here, avian adeno-associated virus (AAAV) was consistently recovered from limiting dilutions of purified and unpurified avian Ad stocks propagated in embryonating chicken eggs derived from two independently raised flocks of White Leghorn (WL) chickens but not when these Ad stocks were propagated in duck cells. These observations suggest that AAAV is a latent endogenous virus of at least some flocks of WL chickens.

The effect of incubation temperature on the propagation of duck adenovirus (virus 127-like) in duck and chicken cells.

Duck adenovirus (Cornell strain) was propagated in duck and chicken embryo cells at 37.5 C and at 40 C. In duck cells, virus levels, as indicated by HA titers, peaked earlier at 40 C than at 37.5 C. High titers were eventually observed in duck cells at both temperatures. In chicken embryo fibroblasts, no titers were observed at 37.5 C, whereas low titers were observed at 40 C. Evidence of virus propagation was not detected in chicken embryo liver and kidney cells.

Serologic evidence of avian adeno-associated virus infection in an unselected human population and among poultry workers.

Six (6.0%) of 100 serum samples from an unselected adult population were positive for antibody to avian adeno-associated virus (A-AV) by agar gel precipitation (AGP), and 10 (15.6%) of 64 were positive by virus neutralization (VN). Three (14.3%) of 21 samples from poultry workers (industry or research) were positive for avian A-AV antibody by AGP and 14 (66.7%) of 21 were positive by VN. All sera positive by AGP were also positive by VN. Twenty-two of 244 sera positive for antibody to avian A-AV by VN were positive for human adenovirus antibody. None of the sera were positive for avian adenoviral antibody by AGP. No cross reaction was noted by AGP when antiserum to avian A-AV was reacted against primate antigens of serotypes 1-4 or when antiserum to A-AV serotypes 1-14 were reacted against avian A-AV antigen. Antiserum prepared against primate A-AV serotypes 1-4 did not neutralize the avian A-AV. These results suggest that avian A-AV infections are not restricted to avian species but are found in the human adult population.

Neutralizing antibodies to CELO and avian adenovirus-associated viruses in the albumen of chicken eggs.

Neutralizing antibodies to CELO virus and to avian adenovirus-associated virus (A-AV) were detected in the albumen of eggs from four hens inoculated with these viruses. The antibody concentrations of serum, yolk, and albumen were determined before inoculation and at various times postinoculation (PI) by enzyme-linked immunosorbent assay (ELISA) and virus-neutralization (VN) tests. The antibody concentration in albumen was 0.3% to 1.0% of that detected in serum and yolk. Uninoculated hens showed no detectable antibody in serum, yolk, or albumen. It is suggested that the presence of antibody in the egg albumen may play a role in egg-transmission of viruses.

Enhancement and inhibition of CELO virus pathogenicity in quail by avian adenovirus-associated virus.

Dual infection of 12 day-old quail (Colinus virginianus) with 10(6) plaque forming units of CELO virus and low doses of avian adeno-associated virus (A-AV), resulted in significant enhancement of CELO virus-induced mortality, whereas dual infections with high doses of A-AV resulted in a delay in mortality. A-AV induced enhancement and inhibition of CELO virus pathogenicity could be blocked by the addition of A-AV antiserum prior to infection.

An enzyme-linked immunosorbent assay for detection of antibodies to avian adenovirus and avian adenovirus-associated virus in chickens.

An enzyme-linked immunosorbent assay system (ELISA) was adapted for the detection of antibodies to avian adenovirus (AV) and avian adenovirus-associated virus (A-AV). Both before and after exposure, sera from chickens undergoing natural and experimental infections were assayed by ELISA, virus neutralization (VN), and immunodiffusion (ID) for antibody to both CELO virus and A-AV. The ELISA system was found to be comparable to VN for determining antibody concentrations to CELO virus and A-AV. In many cases, ELISA was found to be more sensitive than ID.

Populations of infectious virus produced during avian adenovirus-associated virus infection.

Multiple rounds of infection in vitro or in vivo with avian adenovirus-associated virus (AvA-AV) and avian adenovirus (AvAV) result in production of both heavy (H) and light (L) infectious forms. In this study, the infectious AvA-AV progeny produced at different hours during recombined dual infection of cells with either H or L AvA-AV and AvAV was determined by equilibrium CSCl centrifugation and infectivity assay. In both types of infection, H virions were found early, both intra- and extracellularly, whereas L virions were found late. The data iondicate an H to L particle density shift during infection. Virus-specified cell-dependent factors mediated the process extracellularly, as activity was detected in infected cell-conditioned medium and in lysates of infected cells but not in medium or components of uninfected cells. The shift in density was accompanied by a conversion in particle type. H and L virions differed in size and conformation as evidenced by differences in serological cross-reactivity and physical-chemical stability during heat inactivation, ultrasonic disruption and DNA extraction.

Egg transmission of avian adenovirus-associated virus and CELO virus during a naturally occurring infection.

Both avian adenovirus-associated virus (A-AV) and CELO virus were isolated from the embryonating eggs of 25-week-old black sex-linked hens during a naturally occurring infection. In the first 7 days of egg collection, A-AV was isolated from 10 of 43 (23.2%) embryonating eggs, and CELO virus was isolated from 8 of 43 (18.6%) embryonating eggs. Both viruses were isolated from six eggs. In the next 16 days of egg collection, A-AV and CELO virus were coisolated from 1 of 127 (0.8%) eggs; all other samples were negative for both viruses. All six hens transmitting A-AV to eggs and 5 of 6 hens transmitting CELO virus showed seroconversions (fourfold increase in antibody concentrations). Viruses were not isolated from eggs after the hens showed a fourfold increase in antibody concentrations.

Inhibition and enhancement of avian adenovirus plaque production by heavy and light avian adenovirus-associated viral particles.

The effect of heavy and light avian adenovirus-associated viral (A-AV) particles on the replication of several adenovirus serotypes was studied in chicken embryo kidney cells. There was a significant decrease (P less than 0.05) in the number and size of adenovirus-induced plaques at A-AV multiplicities of infection greater than 40. Enhancement of plaque production was observed when A-AV multiplicities of infection were 1 to 40. There was a significant increase in the number and size of infective centers. Analysis of cellular yields indicated an increase in the number of adenoviruses produced per cell. Heavy A-AV particles of buoyant density 1.42 g/cm3 in CsCl were found to enhance plaque production more than light particles (1.38 g/cm3). Conversely, light particles showed greater inhibition of plaque production. Adenovirus serotypes varied in their response to enhancement or inhibition by A-AV particles of different density.

Effect of avian adeno-associated virus on pathogenicity of Tipton virus in chicks.

The avian adeno-associated virus (A-AV) reduced the pathogenicity of an adenovirus infection in vivo. Groups of chicks were infected with Tipton virus alone or in combination with high or low doses of A-AV. In both trials, the associated virus delayed and reduced chick mortality. This effect was dose-dependent and significant at the higher dose level.

An in vivo and in vitro study of infectious laryngotracheitis virus in chicken leukocytes.

Infectious laryngotracheitis (ILT) virus, an avian herpesvirus, caused an infection in chickens that was followed by leukopenia due to a decrease in the number of circulating lymphocytes. Viral synthesis in leukocytes in cell cultures was evident by specific and progressive viral antigens in the nuclei of infected leukocytes as shown by fluorescent-antibody technique and by the formation of multinucleated giant cells, typical of the herpesviruses. In addition, viral multiplication was observed in the leukocyte cell cultures as shown by viral assay. It is suggested that the leukocytes in chicken are participating in the production of ILT virus by serving as host cells for viral multiplication. This is followed by a cytopathogenic effect, with the production of multinucleated giant cells. That may result in the destruction of cells, causing a leukopenia.