Duck Embryo Fibroblast Cell Cultures Research Articles

Isolation of Poxvirus from Debilitating Cutaneous Lesions on Four Immature Grackles (Quiscalus sp.)

Poxvirus was isolated from nodules on four immature grackles (Quiscalus sp.) collected in two residential areas of Victoria, Texas. All of the birds were emaciated and had nodules on the eyelids, bill, legs, toes, and areas of the skin on the wings, neck, and ventral abdomen. These pox nodules were extensive and probably interfered with both sight and flight. The preliminary diagnosis was confirmed by virus isolation, histopathology, and electron microscopy. Poxvirus was isolated on the chorioallantoic membrane of embryonated hen's eggs and in Muscovy duck embryo fibroblast cell culture. Phaenicia calliphoridae (blowfly) larvae were found in one of the pox nodules, raising the possibility of mechanical transmission of the virus by contaminated adult blowflies.

Venezuelan equine encephalomyelitis virus: Concentration, partial purification, inactivation and immunogenicity

Venezuelan equine encephalomyelitis (VEE) TC-84 vaccinal virus, from 10-1. quantities of infected duck embryo fibroblast cell culture fluids, was isolated by combined continuous-flow centrifugation with isopycnic banding in sucrose. Most of the recovered infectivity and hemagglutinating activity were in a single band at a buoyant density (ϱ) of 1.2. About 90% of the total input protein (450–520 mg) was removed with the effluent, whereas most of the remaining 10% also banded at a ϱ of 1.2. Infectivity was inactivated with formalin at a final concentration of 0.05% at 37°C for 24 hr. Formalin-inactivated virus retained its immunogenicity and induced VEE virus-specific antibody in horses and guinea pigs. The horses and those guinea pigs that received equivalent doses of vaccine survived after a challenge of their immunity with virulent VEE virus.

Duck Plague Virus Replication in Muscovy Duck Fibroblast Cells

Duck embryo fibroblast cell cultures from seven species of ducks were compared for virus yield, plaque quality, and sensitivity to infection by the duck plague herpesvirus (duck virus enteritis). Muscovy duck and wood duck cells gave the best results for virus yield and plaque quality, but muscovies were considered superior because they are more available than wood ducks. Pintails and lesser scaup gave the poorest results, and pekin duck, black duck, and redhead duck were intermediate. A growth curve for the virus, determined in muscovy cells, had a latent period of six hours and a maximum new virus titer reached at 36 hours. Because of their superior plaque production and ability to replicate the virus, muscovy embryo fibroblasts (MCE) are recommended for diagnostic and research work.

TRANSMISSION OF AVIAN POX FROM STARLINGS TO ROTHCHILD'S MYNAHS

Several wild species of birds, including starlings (Sturnus vulgaris), house sparrows (Passer domesticus) and pigeons (Columba livia) gained access to an aviary housing Rothchild's mynahs (Leucospar rothchildii) and over 100 additional birds representing a variety of species. Six of approximately 15 mynahs became infected with avian pox and all of them died. None of the other birds in the aviary developed lesions. Pox virus was isolated from mynah facial lesions on chicken chorioallantoic membrane and in duck embryo fibroblast cell culture. It did not produce lesions in white Leghorn chickens, but did produce lesions in 4 of 11 wild starlings captured outside the aviary. Results indicated the agent was an indigenous starling pox capable of infecting and producing disease in mynah birds. Destruction of the captive starlings and isolation of the remaining mynahs immediately stopped the mortality.

Differentiation between strains of Marek's disease virus and turkey herpesvirus by immunofluorescence assays

Two serological types of Marek's disease virus and a herpesvirus of turkeys have been differentiated by indirect immunofluorescence tests as (1) pathogenic strains of Marek's disease virus (MDV) and their attenuated variants: HPRS-16, HPRS-16/att, HPRS-B14, JM, JM/att, GA, VC and 'Oldenburg', a recent field isolate; (2) apathogenic strains HPRS-24 and HPRS-27 of MDV; (3) herpesvirus of turkeys strain FC126 and its HVT(A-) variant. Virus strains could not be distinguished on the basis of qualitative differences in immunofluorescent staining of intracellular virus-induced antigens. Results were similar whether chicken kidney, chicken embryo fibroblast or duck embryo fibroblast cell cultures were used. Fluorescence of virus-induced antigens was stronger with homologous than with heterologous antisera. Using the direct immunofluorescence technique Marek's disease virus and turkey herpesvirus infections could be distinguished. There were never any significant differences in the appearance and distribution of antigen in infected cells treated with homologous or heterologous antisera at dilutions of comparable activity using the indirect immunofluorescence technique. Antibody titres of antisera were 4 to 8-fold higher in the indirect immunofluorescence test against the homologous virus-induced antigens than against heterologous antigens. Cross-reactions between the 3 serological types could be prevented by absorption of antisera with the appropriate antigens. Cross-reactions could also be prevented by the appropriate dilution of antisera before use in the indirect immunofluorescence test.

Differentiation between strains of Marek's disease virus and Turkey herpesvirus by immunofluorescence assays

Summary Two serological types of Marek's disease virus and a herpesvirus of turkeys have been differentiated by indirect immunofluorescence tests as (1) pathogenic strains of Marek's disease virus (MDV) and their attenuated variants: HPRS‐16, HPRS‐16/att, HPRS‐B14, JM, JM/att, GA, VC and ‘Oldenburg’, a recent field isolate; (2) apathogenic strains HPRS‐24 and HPRS‐27 of MDV; (3) herpesvirus of turkeys strain FC126 and its HVT(A‐ ) variant. Virus strains could not be distinguished on the basis of qualitative differences in immunofluorescent staining of intracellular virus‐induced antigens. Results were similar whether chicken kidney, chicken embryo fibroblast or duck embryo fibroblast cell cultures were used. Fluorescence of virus‐induced antigens was stronger with homologous than with heterologous antisera. Using the direct immunofluorescence technique Marek's disease virus and turkey herpesvirus infections could be distinguished. There were never any significant differences in the appearance and distribution of antigen in infected cells treated with homologous or heterologous antisera at dilutions of comparable activity using the indirect immunofluorescence technique. Antibody titres of antisera were 4 to 8‐fold higher in the indirect immunofluorescence test against the homologous virus‐induced antigens than against heterologous antigens. Cross‐reactions between the 3 serological types could be prevented by absorption of antisera with the appropriate antigens. Cross‐reactions could also be prevented by the appropriate dilution of antisera before use in the indirect immunofluorescence test.

Characterization of an avian reovirus isolated in Queensland

An avian reovirus, RAM-1, produced cytopathic changes in chicken kidney, chick embryo fibroblast and duck embryo fibroblast cell cultures. The virus could also be titrated in chicken embryos, but this method was much less sensitive than cell culture. Following yolk sac inoculation, deaths occurred in two to four days. Inoculation onto the chorio-allantoic membrane resulted in necrotic plaques, with about 10 per cent. mortality. Surviving embryos had green discoloration of the liver, enlarged spleen and sometimes pericarditis. Features of the histopathological response were ectodermal hyperplasia and necrosis in the chorio-allantoic membrane, and accumulations of eosinophils in the liver, the spleen and the pericardium. Pericarditis developed in a few chickens inoculated at one day of age. The virus was not pathogenic for suckling mice. Virions typical of reoviruses could be demonstrated by electron microscopic examination of infected cell cultures by negative staining, or thin section techniques. The virus was stable on exposure to chloroform and pH 3·0, and was partially inactivated by exposure to 56 °C. for 30 min. Neutralizing antibody could be detected in cell cultures, or in embryonated eggs inoculated onto the chorio-allantoic membrane, but yolk sac inoculation gave unsatisfactory results. Precipitating antibody in hyperimmune rabbit serum was demonstrated with antigen derived from cell cultures and from chorio-allantoic membrane. Convalescent serum from chickens reacted only with the antigen derived from chorio-allantoic membrane.

Detecting Marek's Disease Herpesvirus: Evaluation of Biological Systems

Systems were evaluated for isolating and identifying Marek's disease herpesvirus (MDHV) from purposely contaminated samples of poultry virus vaccines. The sensitivity and specificity of duck embryo fibroblasts (DEF) and chick kidney (CK) cell cultures for detecting MDHV were compared with the inoculation of embryonated eggs and day-old chicks. Duck embryo fibroblast cell cultures were more sensitive for detecting MDHV and less susceptible to the cytopathogenic effect (CPE) of vaccine viruses than either CK cell cultures or embryonated eggs. The inoculation of day-old chicks appeared less sensitive than DEF cell cultures when infection was determined by culture of kidneys or by use of a radial diffusion test for MDHV feather follicle antigen at 14 days postinoculation. At 21 days postinoculation, chicks appeared as sensitive as DEF cell cultures when infection was determined by kidney culture and more sensitive than cell cultures when the radial diffusion test was employed to determine infected birds. Inoculation of chicks was the only satisfactory method for isolating MDHV from samples of avian pox virus vaccines. Fluorescent-antibody (FA) staining confirmed the identity of MDHV isolates made in DEF cell cultures. The FA procedure was used to examine 28 samples of commercial virus vaccines for MDHV. This virus was not detected as a contaminant in any of the samples.

Horizontal Transmission of Turkey Herpesvirus to Chickens: 2. Some Factors Affecting Transmission

The likelihood of horizontal spread of turkey herpesvirus (HVT) increased as the dose of the cell-associated HVT given to donor chickens (8-week-old) was increased from 7,400 to 14,800 and to 74,000 plaque-forming-units (PFU). However, further increase in virus dose to 370,000 PFU per donor did not result in spread.Similar increase in the horizontal spread of HVT resulted with the cell-free HVT when the virus dose was increased from 1,400 to 11,280 PFU per donor bird. With further increase to 490,000 PFU per donor, the virus srpead poorly.Cell-associated HVT appeared to spread more effectively than cell-free virus.HVT grown in duck embryo fibroblast cell cultures spread readily when a small (5,800 PFU) or large dose (435,000 PFU) of the virus was given to donor birds, and seemed to spread more effectively than HVT grown in chicken embryo fibroblast cell cultures.Marek’s disease virus maternal antibodies present in chicks (10-day-old) at the time of HVT inoculation had no suppressive effect on the horizontal transmission of HVT to contact cagemates.