Comparative sensitivities of oviduct and tracheal organ cultures and chicken embryo kidney cell cultures to infectious bronchitis virus

In vitro studies with organ (oviduct and trachea) and chicken embryo kidney cell cultures were attempted to assess the pathogenicity of locally isolated infectious bronchitis virus (IBV-P:120) initially isolated from the oviduct of young chicks. In oviduct cultures infected with IBV, ciliary movements decreased as early as 24 hours postinoculation (PI), and on the 6th day ciliary movements ceased completely. Cytopathic changes were also noticed. Immunofluorescent antigen was detected from 1 to 6 days PI, the maximum being on the 3rd day. The characteristic microscopic changes in the oviduct explants were reduced by 24 hours PI and had completely ceased on the 5th day. Cytopathic effect and immunofluorescent antigen were present from 1 to 8 days PI, being maximum on the 5th day. Histological changes marked by loss of cilia, rounding of the epithelial cells, degeneration, and sloughing were detected from 2 to 8 days PI. Low-embryo-passaged (EP-7) IBV did not produce cytopathic effect on the chicken embryo kidney cell cultures. On the contrary, high-embryo-passaged (EP-14) virus produced cytopathic effect at the third tissue-culture-passage level.     

Bilateral subcutaneous fibrosarcomas in a cat following feline parvo-, herpes- and calicivirus vaccination

A crossbred cat developed a subcutaneous fibrosarcoma on the left side of the thorax at the site of previous administration of a feline parvo-, herpes- and calicivirus vaccine. A few months later the cat developed a second mass on the right side of the thorax after a booster vaccine had been administered at this site. This unique case of bilateral fibrosarcomas in a cat shortly after vaccination with parvo-, herpes- and caliciviruses suggests an individual disposition for the development of vaccine-associated sarcomas and a possible triggering of this type of pathological response which could have precipitated the development of the second tumour. To the authors' knowledge, this is the first case of vaccine-induced fibrosarcomas occurring bilaterally after injection of a feline parvo-, herpes- and calicivirus containing vaccine at different sides of the thorax.     

Characteristics of a feline mammary carcinoma cell line

The establishment of an epithelial cell line, JM, from a feline mammary adenocarcinoma is described. In vitro, the morphological and cultural properties of these cells, their surface features such as lectin binding and their response to hormones were ascertained. In vivo, they are tumorigenic in athymic nude mice inducing tumour nodules composed of epithelia-like cells.     

Isolation of feline herpesvirus 1 from a young kitten

Five kittens exposed neonatally to older cats exhibiting fever with nasal and ocular discharge became ill and died within 10 days of the onset of illness. One kitten which died at 16 days old was examined post mortem and feline herpesvirus 1 was isolated from the brain, liver, lung and spleen.     

Genetic analysis of feline caliciviruses associated with a hemorrhagic-like disease.

Feline calicivirus (FCV) is 1 of the most common causes of upper respiratory tract disease in cats. Other disease syndromes associated with FCV infection have been reported. Recently, calicivirus infection associated with a hemorrhagic-like disease leading to significant mortality in cats has been reported. The clinical signs are similar to those observed with the calicivirus of rabbit hemorrhagic disease. This study characterized 2 FCV isolates associated with hemorrhagic-like disease. Nucleotide sequencing of the complete genome has been done for these 2 isolates as well as for 4 additional isolates representing other disease syndromes. Previously reported sequence data for the entire genome of classical FCV (6 isolates) and a portion of the capsid gene for hemorrhagic-like FCV (3 isolates), isolated in different regions of United States were used in the genetic analysis. Sequence data were used to determine relationships among the isolates and any correlation with phenotype. Nucleotide sequence comparisons of the entire genome and individual open reading frames revealed high homology among all isolates. Data suggest that the virulence may have genetic determinants on the basis of phylogenetic clustering of the isolates associated with hemorrhagic-like disease.     

Induction of feline immunodeficiency virus from a chronically infected feline T-lymphocyte cell line

The infection of the feline T-lymphocyte cell line FeT-J with the feline immunodeficiency virus (FIV) Petaluma strain led to the establishment of nonvirus-producing cells. One clone (C15) obtained by limiting dilution was found to express FIV in response to chemical inducers of retroviruses. The chemical treatment of C15 cells led to not only FIV protein synthesis but also an augmentation of viral production. Examination of the C15 cell derivatives obtained by recloning revealed that 10-40% of treated cells constitutively expressed FIV antigens, whereas 100% with expressed FIV antigen in response to the inducer. Chemical induction resulted in more than a 100-fold increase in infectious viral production. The results suggest that a majority of FeT-J cells that are infected with FIV exist in a non-productive state. Establishing a cell line that can be non-productively infected by FIV may help determine the mechanisms of FIV latency.     

Studies of enteric coronaviruses in a feline cell line

Development is reported of a feline cell line which can support the growth of coronaviruses from canine (CCV), feline (FIPV) and porcine (TGEV) species. The cell culture has been serially transferred over 100 times and has retained its initial growth requirements, proliferative capacity and morphologic features. Each virus had specific growth characteristics in this cell culture although all produced a similar CPE and plaques under agar. Cross neutralization studies demonstrated a two-way relationship between TGEV and CCV and between TGEV and FIPV, whereas a one-way relationship was demonstrated between CCV and FIPV.     

Susceptibility of cell line cultures of bovine kidney origin to infectious bovine rhinotracheitis and parainfluenza-3 viruses

A comparative study was carried out on the susceptibility of primary bovine embryo kidney (PBEK) cell cultures, and that of AUBEK and MDBK cell lines to infectious bovine rhinotracheitis (IBR) and Parainfluenza-3 (PI-3) viruses.

The cytopathic effects induced by the two viruses were rather inconsistent, based on observations of unstained preparations. On the other hand, there was no significant difference between the susceptibility of the PBEK cultures and the cell line cultures to infection with either virus on the basis of the lesions detected in stained preparations, and of the growth curve patterns.

It is concluded that PBEK cell cultures are more sensitive for isolating IBR or PI-3 viruses than are the AUBEK and MDBK cell lines. However, the latter appear to be satisfactory for studies of these two viruses.     

Full-length ORF2 sequence-based genetic and phylogenetic characterization of Korean feline caliciviruses

Feline calicivirus (FCV) is a highly infectious pathogen in cats and widely distributed worldwide with high genetic variation. Full-length open reading frame 2 of 5 from recently isolated Korean FCV isolates were sequenced and compared with those of global isolates. The results of phylogenetic analysis supported dividing global FCV isolates into two genogroups (type I and II) and demonstrated the presence of genogroup II in Korea, indicating their geographic spread in East Asia. High sequence variations in region E of the FCV isolates emphasizes that a novel vaccine needs to be developed to induce protective immunity against various FCV strains.     

Propagation of infectious bovine rhinotracheitis (bovine herpes-1) virus in murine primary cell cultures

Virus synthesis in BALB/C mouse lung and kidney primary cultures infected with infectious bovine rhinotracheitis (IBR) virus started between 6 and 8 hours after virus inoculation and reached a maximum titer of 5.5 log₁₀ plaque forming units (PFU) at 48 hours postinfection (PI). Cytopathic effect (CPE) in cell cultures occurred at 8–10 hours and over 90% of the cells had CPE by 48 to 72 hours PI. The bulk of the newly replicated virus (60–80%) was cell-associated as determined by plaque assay of extracellular and intracellular virus. Pulse-chase experiments demonstrated incorporation of radioactive precursors into viral DNA and protein macromolecules. Viral DNA synthesis was initiated between 2 and 4 hours PI and was maximum at 4 to 6 hours. Viral proteins were detected at 4 hours and peaked between 6 and 8 hours PI. Enzyme-linked immunosorbant assay (ELISA) confirmed synthesis of specific viral proteins, which gradually increased during the virus growth cycle. Abstract in French is given at the end of the article.

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Resume La synthèse du virus de la rhinotrachéite infectieuse bovine (virus IBR) sur des cultures primaires de poumon et de reins de souris (souche Balb/C) commençait entre 6 et 8 heures après inoculation du virus et atteignait un titre maximal de 5.5 log PFU (Plaque Forming Unit) 48 heures après infection. L'effet cytopathogenic (CPE) sur ces cultures cellulaires apparaissaient en 8 à 10 heures et plus de 90% des cellules montraient un CPE en 48 à 72 heures après infection. La détermination par la méthode des plages sous agarose, du virus extracellulaire et du virus intracellulaire, indiquait que 60–80% du virus synthétisé était associé avec les cellules. Les expériences de pulsechase demontraient l'incorporation des précurseurs radioactifs dans les protèines et ADN viraux. La synthèse virale d'ADN était initiée entre 2 et 4 heures après infection et était maximale en 4 à 6 heures. Les protéines virales étaient détectées en 4 heures avec un maximum de synthèse entre 6 et 8 heures après infection. Enzyme-linked immunosorbant assay (ELISA) confirmait la synthèse de protéines virales spécifiques qui montrait une augmentation graduelle pendant le cycle de replication du virus.

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Supported in part by grants 0831 from the Cooperative State Research Service of the US Department of Agriculture and published as contribution 85-160-J, Kansas Agricultural Experiment Station.     

Masitinib mesylate does not enhance sensitivity to radiation in three feline injection-site sarcoma cell lines under normal growth conditions

Masitinib, a selective tyrosine kinase inhibitor, was investigated as a radiosensitizer in three primary feline injection-site sarcoma (ISS) cell lines. Sensitivity to masitinib was previously assessed via cell growth inhibition assays with mean IC₅₀ values of 5.5–8.6 μM. Clonogenic assays were performed to determine the effect of masitinib and radiation on cell survival. Single dose radiation (0–12 Gy) experiments were carried out under normal growth conditions in control ISS cells and in cells incubated with 1 or 6 μM masitinib for 72 h prior to irradiation. Radiation administered either alone or in combination with masitinib induced a dose-dependent reduction in clonogenic survival. Survival from the combined masitinib and radiation treatment was not significantly different from that of radiation alone. Results suggest that masitinib does not directly enhance ISS cell radiosensitivity under normal in vitro conditions, although this does not preclude the utility of further investigations to assess sensitization properties under altered conditions.