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An antibody-capture enzyme-linked immunosorbent assay (ELISA) for detection of antibody to turkey coronavirus (TCV) utilizing infectious bronchitis virus (IBV) antigen was developed. Anti-TCV hyperimmune turkey serum and normal turkey serum were used as positive or negative control serum for optimization of the ELISA system. Goat anti-turkey immunoglobulin G (light plus heavy chains) conjugated with horseradish peroxidase was used as detector antibody. The performance of the ELISA system was evaluated with 45 normal turkey sera and 325 turkey sera from the field and the cutoff point was determined. Serum samples of turkeys experimentally infected with TCV collected sequentially from 1 to 63 days postinfection were applied to the established antibody-capture ELISA using IBV antigens. The optimum conditions for differentiation between anti-TCV hyperimmune serum and normal turkey serum were serum dilution at 1:40 and conjugate dilution at 1:1600. Of the 325 sera from the field, 175 were positive for TCV by immunofluorescent antibody (IFA) assay. The sensitivity and specificity of the ELISA relative to IFA test were 93.1% and 96.7%, respectively, based on the results of serum samples from the field turkey flocks using the optimum cutoff point of 0.18 as determined by the logistic regression method. The ELISA values of all 45 normal turkey sera were completely separated from that of IFA-positive sera. The ELISA results of serum samples collected from turkeys experimentally infected with TCV were comparable to that of the IFA assay. Reactivity of anti-rotavirus, anti-reovirus, anti-adenovirus, or anti-enterovirus antibodies with the IBV antigens coated in the commercially available ELISA plates coated with IBV antigens could be utilized for detection of antibodies to TCV in antibody-capture ELISA.

Epithelial damage in infectious bronchitis occurs early in the disease process. Heterophil infiltration into the tracheal mucosa is greatest at that time. To determine the contribution of heterophils to tracheal epithelial damage of infectious bronchitis, eight 3-wk-old specific-pathogen-free chickens were made heteropenic by four daily intramuscular injections of cyclophosphamide at 75 mg/kg body weight. Infection with Massachusetts 41 infectious bronchitis virus was timed to coordinate heteropenia with peak tracheal epithelial damage. Heteropenia was monitored by total leukocyte and differential cell counts of peripheral blood. Tissue damage and heterophil infiltrate were monitored by histopathology of tissues taken at termination of the study. Heteropenic birds had lower peripheral blood and tracheal heterophil numbers than nonheteropenic birds. No difference was found in epithelial damage of heteropenic and nonheteropenic birds. Epithelial damage in infectious bronchitis is most likely due to damage by the virus and not due to the infiltrated heterophils.

A sudden death syndrome was induced in chicks and poults fed diets containing Fusarium fujikuroi, formulated to contain 0-330 mg/kg moniliformin (M) with or without the maximum recommended therapeutic concentration of monensin. Lesions of monensin toxicosis were not observed. Clinical signs were referable to cardiac dysfunction (sudden death, dyspnea, cyanosis, depression). Poults and chicks dying early in the study had no gross lesions or had lesions of right ventricular dilation. Treated poults and chicks dying late in the study or euthanatized at termination of the study had lesions of bilateral myocardial hypertrophy, usually concentric. Absolute heart weights and relative heart weights, expressed as a percentage of body weight, were significantly greater in treated birds than controls (P < 0.05), whereas body weights were significantly less (P < 0.05). Microscopically, lesions progressed from acute myocardial degeneration to necrosis, fibrosis, and hypertrophy. Ultrastructural findings were consistent with the gross and microscopic lesions. Serum pyruvate concentrations were a useful indicator of M-induced cardiotoxicosis. Concentrations of serum pyruvate increased with increased concentration of dietary M, but were not affected by addition of monensin to the diet. In chicks ingesting 40-300 mg/kg M, serum pyruvate concentrations were significantly greater (P > 0.05) than those in controls (controls, 0.28 +/- 0.08 mmol/liter; exposed 0.38 +/- 0.11-0.55 +/- 0.13 mmol/liter). Poults ingesting 80-330 mg/kg M had significantly greater serum pyruvate concentrations than controls (controls 0.33 +/- 0.09 mmol/liter; exposed 0.43 +/- 0.13-1.00 +/- 0.006 mmol/liter). The Vetronics System was used to evaluate electrocardiographic alterations in a limited number of chicks and poults surviving to the end of the feeding trial. Electrocardiographic alterations in poults and chicks fed diets containing > or = 40 mg/kg and > or = 160 mg/kg M, respectively, were consistent with ventricular hypertrophy, myocardial injury, and hypoxia. Electrocardiographic alterations were more striking in poults than in chicks. Altered myocardial metabolism due to M toxicosis, in conjunction with the unusual susceptibility of domestic poultry to altered cardiac metabolism, is believed to be the cause of the organ-specific lesions in these birds. These findings suggest that cardiac injury with subsequent alterations in cardiac electrical conductance may be a cause of the sudden deaths observed in poultry chronically intoxicated with dietary M.

Cyclophosphamide (Cytoxan) was given at 75 mg/kg body weight via daily intramuscular injections for 4 days to 3-week-old specific-pathogen-free (SPF) chickens in an attempt to determine if heteropenia could be induced in chickens. Control birds were given a like quantity of phosphate-buffered saline, the diluent for Cytoxan. Peripheral blood heterophil numbers were determined and monitored by total leukocyte and differential cell counts. Birds were grouped in pairs on day 0 based on total leukocyte count. The number of heterophils each bird had on day 0 served as a baseline heterophil count for that bird. Thereafter heterophil numbers were determined on the last day of drug treatment and every other day until blood heterophil numbers were 20% of that bird's baseline heterophil count (heteropenia). The effects of Cytoxan on trachea, lung, liver, kidney, bursa of Fabricius, bone marrow, spleen, and thymus were determined by microscopic examination of those tissues collected the day following heteropenia. Cytoxan had no effect on trachea, lung, liver, kidney, and thymus. Bursa of Fabricius and spleen had decreased amounts of lymphoid aggregates. Bone marrow of Cytoxan-treated chickens was hypocellular. The study was then repeated to determine the reversibility of Cytoxan-induced heteropenia. Cytoxan-treated birds were allowed to recover until blood heterophil numbers equaled or exceeded those of control birds. Cytoxan, through bone marrow suppression, induced a reversible heteropenia that developed between treatment days 10 and 12. In addition, Cytoxan induced a reversible lymphocytopenia between days 4 and 10 and a regenerative anemia between days 8 and 10. The ability to produce heteropenia in SPF chickens will allow the use of a heteropenic model for further study of the heterophil's contribution to the inflammatory response.

Cellular response of chickens to infection with infectious bronchitis virus (IBV) was investigated by lavage of the respiratory tract of five 2-week-old specific-pathogen-free (SPF) chickens at 2, 8, 24, 48, 72, and 96 hours postinfection (PI) with either Massachusetts 41 (IBV-M41) or Australian T (IBV-T) IBV. Tissue response was monitored by microscopic examination of trachea and lung from five non-lavaged infected chickens collected at the same intervals. The total number of cells recovered by lavage from IBV-M41-infected chickens was dramatically higher than the total number recovered from IBV-T-infected chickens and uninfected controls. By contrast, the total number of cells recovered from IBV-T-infected chickens was no higher than that of the uninfected chickens. Heterophils constituted the majority of inflammatory cells recovered from both IBV-M41-infected and IBV-T-infected chickens. Heterophil numbers in IBV-M41-infected chickens paralleled total cell-number recovery, whereas heterophil numbers in IBV-T-infected birds were no higher than those in uninfected chickens. The number of lymphocytes recovered from IBV-M41-infected chickens increased 72 hours PI and continued to increase for the duration of the study. Lymphocyte numbers in IBV-T-infected chickens exceeded those in uninfected chickens only at 96 hours PI. The number of lavage macrophages in IBV-M41-infected chickens increased earlier than the number of lymphocytes but later reached a plateau. IBV-T macrophage numbers did not exceed those of uninfected chickens. Tissue damage occurred most consistently in the trachea and occurred when lavage heterophil numbers were rising or at their peak. Lavage cell recovery and composition reflected tracheal mucosa inflammatory cell infiltrate.

Severe hepatic trematodiasis due to Platynosomum proxillicens was diagnosed as the cause of death in two sulfur-crested cockatoos (Cacatua sulfurea). Gross pathologic alterations included hepatomegaly in one bird and hepatic and generalized cyanosis in the other. Microscopic changes included multifocal hepatic necrosis, dilated fibrous bile ducts containing trematode, parasites, bile duct hyperplasia, and hepatic inflammation.

Two commercial White Leghorn flocks of 41,000 and 57,000 25-week-old hens experienced death losses of 676 and 1,089 birds in a week. Six birds from each flock were presented for necropsy to the Animal Disease Diagnostic Laboratory at Purdue University with a clinical history of excessive deaths, unsatisfactory production, and dehydration. Gross lesions were present on the tongue and on oral and pharyngeal mucosa as uniformly pale or white areas with interspersed multifocal sites of ulceration. Other changes were cyanotic, shriveled combs and wattles and ruptured ovarian follicles with concurrent egg yolk peritonitis. Microscopic examination revealed large areas of coagulative necrosis with multifocal epithelial ulceration of the oral and pharyngeal mucosae. There was an intense inflammatory, heterophilic response at the periphery of affected areas. Alterations in the abdominal viscera were limited to egg yolk peritonitis. The high death rate and oral lesions were attributed to accidental use of excessive levels of a quaternary ammonium disinfectant in the drinking water.

A flock of 7-week-old broiler chickens in northwestern Arkansas showed signs of respiratory distress. Gross pathologic alterations were excessive exudate in the tracheas and congestion of the nasal turbinates. An adenovirus was isolated from fresh trachea specimens. Microscopically, alterations in tracheal sections were thickened mucosa with inflammatory-cell lamina-proprial infiltrate and epithelial squamous metaplasia. Protozoan parasites present on the epithelial surface were identified by electron microscopy as Cryptosporidium spp. These organisms were attached to the tracheal mucosa epithelium and the luminal surface of mucous glands. The significance of the intercurrent adenovirus infection was not determined.

Experiments were conducted to characterize renal lesions in chickens induced by four strains of infectious bronchitis virus (IBV); each has been described as nephropathogenic. Those strains were also compared in vaccinated and unvaccinated older chickens for nephropathogenicity. The younger birds were much more susceptible to the nephritogenic effects of the strains. All four strains produced acute renal changes consisting of tubular damage and interstitial inflammatory cell infiltration and edema. Although both cortex and medulla were involved, the latter was generally affected more severely. The Holte strain proved to be the least pathogenic, followed by the more pathogenic Gray and Italian strains and finally by the Australian strain. All four strains produced similar chronic renal changes in unvaccinated birds, with no correlation to the severity of lesions seen at the acute phase. Chronic active and inactive types of interstitial nephritis were seen at the chronic phase. Vaccinated birds challenged with the Australian strain had the highest prevalence of the chronic active type of interstitial nephritis. The implication of renal viral persistence in the development of chronic active interstitial nephritis is discussed.

An outbreak of histomonad infection of an apparent atypical nature was diagnosed in a flock of about 850 Bobwhite quail. Mortality was 95% over a 3-week period. The most prominent gross pathologic lesions were in the livers: disseminated white foci of necrosis, 1 to 2 mm in diameter, and subcapsular multifocal splenic necrosis was seen occasionally; lower intestinal lesions were infrequent. Histologic examination of liver and spleen sections revealed focal necrosis associated with variable numbers of protozoal organisms identified as a Histomonad spp. Identification of the protoza was ascertained by electron microscopy. Histmonads were isolated from affected quail livers and propagated in specific-pathogen-free chicken embryos. Lesions produced in embryos were evaluated. Isolates of the organism were used to reproduce the disease in young Bobwhite quail.

Visceral toxoplasmosis was diagnosed in 23 mynahs (Acridotheres) imported from Mexico, which resulted in the death of all birds in the shipment. The affected birds were inappetent and depressed and had ruffled feathers before death. Gross alterations included mottled, enlarged livers with multiple diffuse subcapsular white foci, congested lungs, mottled spleens, and thickened air sacs. Microscopic alterations were multifocal granulomatous hepatitis, pneumonitis, diffuse splenitis, focal mononuclear myocarditis, and airsacculitis. Foamy macrophages in affected areas of the livers contained 8-10 protozoan organisms 1-3 nm in diameter. Similar organisms were present in affected spleens. The organisms were identified with special stains and electron microscopy as Toxoplasma gondii. Human infection was diagnosed in the caretaker by an attending physician. He was treated with pyrimethamine and apparently recovered.

Two epornitics of erysipelas were diagnosed in domestic White Pekin ducks. The first was in 2,000 18-day-old ducklings with a 30% mortality. The other was in 2,400 breeder hens with losses of four or five birds a day. Gross pathologic lesions in birds necropsied from both instances were enlarged swollen livers with occasional subcapsular hemorrhages. Pure isolates of Erysipelothrix insidiosa only were cultured from the livers. Histologic examination revealed congestion and multifocal necrosis in the liver and spleen with associated reticuloendothelial cell proliferation. Special stains demonstrated numerous organisms within the reticuloendothelial cells.

Disseminated acute focal hepatic coagulation necrosis was present in 9 turkeys submitted from 5 outbreaks of hemorrhagic enteritis. The lesion was unaccompanied by inflammatory cell infiltrate, biliary hyperplasia, or pancreatic necrosis, all of which tentatively distinguish this lesion from that of turkey viral hepatitis. No inclusion bodies were found.

The 2512 infectious bursal disease virus (IBDV) strain maintained in the authors' laboratory was compared with apparent 2512 IBDV isolates designated I-2512, P-2512, and H-2512. The latter three viruses were obtained from different sources and, ostensibly, had their origin in the Purdue laboratory. Their effects on immunogenicity, transmissibility, pathogenicity, and cell cultures varied. One of these isolates was said to be only 2 embryo passages higher than the original seed virus in our laboratory. Included in the study, also for comparison, was a cell-cultured-adapted 2512-cloned attenuated virus. The findings emphasize changes that may occur in the identity of a virus from manipulation, mutation, storage, errors in labeling, or other factors. These characteristics should be identified if the virus is to be used as a vaccine, in research, or for other purposes. The need for well-characterized reference strains in repositories is discussed relative to their importance in potential vaccine research and development.

An outbreak of fatal encephalitis in four-week-old chickens is described. The causal agent was shown to be Baylisascaris procyonis, the common roundworm of raccoons.

Ten strains of adenovirus representing 10 serotypes were administered intratracheally to two groups of 3-week-old specific-pathogen-free chickens. Birds in group 1 were given only adenovirus. Birds in group 2 were inoculated with a virulent infectious bursal disease virus (IBDV) by eye-drop at one day of age as well as with the adenovirus at 3 weeks. The chicks were examined daily. Respiratory rales were observed in some birds with the dual infection. Gross pathologic alterations were minimal and limited to multiple scattered, pale foci in the lungs of an occasional bird in various groups. Histopathologic changes in the lungs were those of multifocal interstitial and, occasionally, diffuse pneumonia. The pneumonic lesions were more severe in the chickens given IBDV. Tracheitis was seen with two of the serotypes.

Eight strains of infectious bursal disease virus (IBDV) were characterized by the criteria of immunity and pathogenicity engendered in young chickens. Some strains have been used commercially, and the others are potential candidates for vaccines. They were administered by drinking water, eyedrop, vent drop, and subcutaneous and aerosol routes. The viruses varied widely in pathogenicity in terms of bursal damage, morbidity, and mortality. Immunity induced with different routes of administration also differed markedly when measured by challenge, virus serum-neutralizing antibody titers, and the agargel precipitin test. Maternally derived immunity interfered with stimulation of active immunity by some of the less pathogenic strains. The more virulent strains, however, were able to provide protection under similar circumstances, although bursal damage was frequently severe. Two strains proved relatively virulent by the criteria employed. Characteristics desirable for IBDV strains to be given as vaccines under field conditions to young chickens are discussed briefly.