Pathogenicity of embryo-adapted serotype 2 OH strain of infectious bursal disease virus in chickens and turkeys

The pathogenicity of serotype 2 OH strain of infectious bursal disease virus (IBDV) to specific-pathogen-free (SPF) chicken embryos and 2-wk-old SPF chickens and turkey poults was investigated. The virus was pathogenic for chicken embryos after five passages as evidenced by pathologic changes in inoculated embryos. The embryo-adapted virus was not pathogenic for 2-wk-old SPF chickens and turkey poults as indicated by lack of clinical signs, gross or microscopic lesions in the bursa of Fabricius of inoculated birds. Bursa-to-body-weight ratios of the inoculated chickens and turkey poults were not significantly different from those of uninoculated controls. Virus-neutralizing antibodies to serotype 2 IBDV were detected in inoculated chickens and turkeys. Results of this study indicated that the embryo-adapted serotype 2 OH IBDV isolate that is pathogenic for chicken embryos is infectious but not pathogenic in chickens and turkeys.     

Viral agents associated with poult enteritis and mortality syndrome: the role of a small round virus and a turkey coronavirus

Intestinal samples from turkey poults affected with poult enteritis and mortality syndrome (PEMS) were examined for viruses by immune electron microscopy and double-stranded RNA virus genome electropherotyping. Turkey coronavirus (TCV), avian rotaviruses, reovirus, and a yet undefined small round virus (SRV) were detected. The SRV and TCV were isolated and propagated in turkey embryos. Challenge of specific-pathogen-free turkey poults with SRV, TCV, or both resulted in mortality and clinical responses similar to those of natural PEMS. Our experiments indicate that SRV and TCV are possibly important agents in the etiology of PEMS and the combination of these infections might result in outbreaks with high mortality. The severity of clinical signs and mortality of PEMS are postulated to be partly related to the virus agents involved in individual outbreaks.     

Enteric disease in specific-pathogen-free turkey poults inoculated with a small round turkey-origin enteric virus

Four- and 5-day-old specific-pathogen-free turkey poults were inoculated orally or by contact exposure to a small round turkey-origin enteric virus. At days 4 and 8 postinoculation (PI), the orally inoculated poults had significantly lower body weight gains than control poults. Poults at day 4 (orally inoculated) and 5 (contact-exposed) PI had watery droppings, dilated thin-walled ceca filled with yellow foamy fluid, catarrhal small intestinal secretions, pale intestinal serosa, and mild lymphocytic enteritis. In addition, at day 4 PI, poults were lymphopenic, had intracytoplasmic crystalline arrays of 17.1 +/- 1.1 nm viral particles in the jejunal villar enterocytes, and had an 18-to-24-nm virus in intestinal contents. Analysis of morphometric data revealed mild shortening of villi in the duodenum and elongation of crypts in the duodenum and ileum during the late stage of the syndrome (day 8 PI). These findings suggest that the 18-to-24-nm virus can produce an enteric disease syndrome and that the acute clinical manifestation of this syndrome is not the result of morphologic change such as intestinal villus atrophy. The definitive identity of this 18-to-24-nm virus is not known; however, based on size and intracytoplasmic arrays of virus, it is most probably an enterovirus.     

Field studies with convalescent serum and infectious bursal disease vaccine to control turkey coryza

Convalescent serum given to 1-day-old poults delayed clinical signs of turkey coryza by several days and reduced mortality on infected farms. Turkey breeders immunized with cell-culture-adapted infectious bursal disease virus (IBDV) or turkey infectious bursal disease virus (TIBDV) had a marked increase in virus-neutralization (VN) antibody titers. The VN antibody titer was significantly higher in progeny poults than in poults from unimmunized breeders. Clinical turkey coryza and mortality was considerably less in poults from IBD- or TIBD-vaccinated breeders than in control poults. They also responded more favorably to hemorrhagic enteritis and fowl cholera vaccination.     

Propagation of virulent and avirulent turkey hemorrhagic enteritis virus in cell culture

Virulent and apathogenic isolates of turkey hemorrhagic enteritis virus (HEV) were successfully propagated in lymphoblastoid cell lines of turkey origin, whereas spleen and kidney cell cultures from HEV-infected turkeys failed to replicate the virus. The lymphoblastoid cell lines used were MDTC-RP16 and MDTC-RP19, which were previously established from tumors induced by Marek's disease virus in turkeys. Virus replication followed co-cultivation of lymphoblastoid cells with spleen cells from HEV-infected turkeys. Virus replication was demonstrated by immunofluorescence, by agar-gel-precipitin tests, and by electron microscopy. Supernatant fluid of cultures infected with virulent HEV caused death and specific lesions in turkey poults. Poults vaccinated with apathogenic HEV were protected against death and lesions after challenge with pathogenic HEV, which was recovered from infected cultures. The MDTC-RP19 cell line appeared far more susceptible than the MDTC-RP16 cell line to infection with HEV.     

The effect of infectious bursal disease virus on the immune system of turkeys

In 1979 it was reported that an infectious bursal disease virus (IBDV) isolated from a case of respiratory disease of turkeys differed antigenically from the chicken isolates of this virus. We injected turkey poults with the turkey-originating TY89 and chicken-originating BD/6 isolates of IBDV and studied their effects on antibody production to the virus, serum immunoglobulin G (IgG), antibody response to sheep erythrocytes, in vitro response of peripheral blood lymphocytes to mitogens, and microscopic structure of the bursa of Fabricius. The chicken isolate BD/6 caused a significant decrease in the response to sheep erythrocytes, lower serum IgG, transient decrease in the response of lymphocytes to PHA, and mild microscopic lesions in the bursa of Fabricius. The turkey isolate TY89, however, caused no obvious damage to the immune system of the infected poults. We suggest that a partial and transient functional disorder of the immune system of poults can occur after infection with IBDV originating from chickens, even if the poults exhibited no clinical signs.     

Infectious bursal disease virus and Alcaligenes faecalis infections in turkeys

To determine if infectious bursal disease virus (IBDV) augments alcaligenes rhinotracheitis (ART), turkey poults were exposed to IBDV, Alcaligenes faecalis, or both IBDV and A. faecalis. In five experiments, poults exposed to IBDV alone exhibited neither signs of disease nor histopathologic lesions. Serum antibodies to IBDV were detected in poults exposed to this virus by inoculation and by direct contact with inoculated birds. Signs of ART were observed 4 to 6 days following exposure to A. faecalis. Clinical signs of ART and histopathologic lesions in the upper respiratory tract of poults exposed to both IBDV and A. faecalis were similar to those observed in poults exposed to A. faecalis alone.     

Turkey origin reovirus-induced immune dysfunction in specific pathogen free and commercial turkey poults

Recently, pathogenesis studies, using genetically distinct turkey-origin reoviruses (TRVs), revealed that poults infected with certain TRV isolates had moderate to severe bursal atrophy, suggesting virus-induced immune dysfunction. In order to characterize the effect of TRV infection on the turkey immune system, classical assays were undertaken to quantify the humoral and cell-mediated immune responses in small Beltsville and broad-breasted white poults infected with the TRV isolate NC/SEP-R44/03. A marked effect on the cutaneous basophil hypersensitivity response, and on the antibody response to Newcastle disease virus (NDV) exposure, was noted in commercial and specific pathogen free (SPF) poults inoculated with NC/SEP-R44/03 at three days of age. Moderate to severe bursal atrophy, similar to that noted previously in SPF poults, occurred in commercial poults inoculated at three days of age. This immune dysfunction and bursal atrophy was not present in commercial poults inoculated at three weeks of age.     

Immunomodular effect of fusion gene DNA vaccine of avian metapneumoviruses

The objective of this work was to develop and evaluate the immunomodular effect of a DNA vaccine based on the fusion (F) gene of avian metapneumoviruses (aMPV) and to study its protection against field virus challenge, as this will help to better control the disease in turkeys. In this study, the F protein of the Egyptian isolate (Giza-turkey rhinotracheitis-4) of the B-subtype of aMPV isolated in 2009 was expressed from a DNA plasmid in Vero cells. After 1 i.m. injection of turkey poults with this plasmid, the antibody response was detected by ELISA. The turkey poults inoculated with locally prepared DNA aMPV vaccine had highly significant phagocytic activity, as measured by phagocytic percent and index of macrophage activation, in comparison to those inoculated with inactivated and live attenuated vaccines and with the noninfected control group. Intratracheal challenge of turkey poults at 21 d postvaccination by a dose of 100 uL of field Egyptian Giza-turkey rhinotracheitis-4 virus of a titer 6 log₁₀ tissue culture infective dose 50 resulted in 100% protection in poults that received locally prepared DNA aMPV vaccine, whereas those that received commercial aMPV vaccines experienced 80 and 90% protection; typical clinical signs of aMPV infection were seen in control nonvaccinated poults. Therefore, a high success rate was noted when using F gene DNA plasmid vaccine by the induction of a potent immunomodular effect for both cell-mediated and humoral immune response. The use of the F gene DNA plasmid vaccine developed in this study provided 100% protection in vaccinated poults, which can help in controlling aMPV infections in turkeys.     

Blood glucose and tissue glycogen levels in turkey poults with spontaneous round heart disease and furazolidone-induced cardiomyopathy.

Furazolidone (FZ) at 700 ppm was added to feed mixtures fed turkey poults two weeks posthatching to induce acute experimental cardiomyopathy. Poults in the control pen received the same ration but without FZ. Four of the control poults developed spontaneous round heart disease. From EKG data and blood samples obtained at weekly intervals, poults were selected for sacrifice at 5 weeks of age. Tissue samples from the left myocardial wall, liver, and pectoralis major and tibialis anterior muscles were analyzed for glycogen by biochemical assay. Blood glucose was determined with the Technicon autoanalyzer. Deposition of glycogen increased significantly (p less than 0.05) in the myocardium of all affected poults and in the liver of all FZ-treated poults. Glycogen levels of the pectoralis major and tibialis anterior muscles were not affected by FZ, but a significant increase (p less than 0.05) was apparent in the pectoralis major muscle of spontaneous round heart poults. It was concluded that FZ influences glycogen metabolism, probably by enzyme inhibition, and that it tends to magnify effects seen in the spontaneous round heart syndrome. Glycogen infiltration of tissues such as the heart and white skeletal muscle suggests that the round heart syndrome may be a manifestation of the glycogen storage disease, idiopathic generalized glycogenosis. Lack of significant differences in the blood serum glucose levels of all poults indicates that these levels are not a reliable clinical parameter for monitoring development of the round heart syndrome.     

Isolation of a reovirus from poult enteritis and mortality syndrome and its pathogenicity in turkey poults

Poult enteritis and mortality syndrome (PEMS) is an acute, infectious intestinal disease of turkey poults, characterized by high mortality and 100% morbidity, that decimated the turkey industry in the mid-1990s. The etiology of PEMS is not completely understood. This report describes the testing of various filtrates of fecal material from control and PEMS-affected poults by oral inoculation into poults under experimental conditions, the subsequent isolation of a reovirus, ARV-CU98, from one of the PEMS fecal filtrates, and in vivo and in vitro studies conducted to determine the pathogenicity of ARV-CU98 in turkey poults. In order to identify a filtrate fraction of fecal material containing a putative etiologic agent, poults were challenged in two independent experiments with 220- and 100-nm filtrates of fecal material from PEMS-negative and PEMS-positive poults. The 100-nm filtrate was chosen for further evaluation because poults inoculated with this filtrate exhibited mortality and significantly lower (P < or = 0.05) body weight and relative bursa weight, three clinical signs associated with PEMS. These results were confirmed in a third experiment with 100-nm fecal filtrates from a separate batch of PEMS fecal material. In Experiment 3, body weight and relative bursa and thymus weights were significantly lower (P < or = 0.05) in poults inoculated with 100-nm filtrate of PEMS fecal material as compared with poults inoculated with 100-nm filtrate of control fecal material. Subsequently, a virus was isolated from the 100-nm PEMS fecal filtrate and propagated in liver cells. This virus was identified as a reovirus on the basis of cross-reaction with antisera against avian reovirus (FDO strain) as well as by electrophoretic analysis and was designated ARV-CU98. When inoculated orally into poults reared under controlled environmental conditions in isolators, ARV-CU98 was associated with a higher incidence of thymic hemorrhaging and gaseous intestines. In addition, relative bursa and liver weights were significantly lower (P < or = 0.05) in virus-inoculated poults as compared with controls. Virus was successfully reisolated from virus-challenged poults but not from control birds. Furthermore, viral antigen was detected by immunofluorescence in liver sections from virus-challenged poults at 3 and 6 days postinfection and virus was isolated from liver at 6 days postinfection, suggesting that ARV-CU98 replicates in the liver. In addition to a decrease in liver weight, there was a functional degeneration as indicated by altered plasma alanine aminotransferase and aspartate aminotransferase activities in virus poults as compared with controls. Although this reovirus does not induce fulminating PEMS, our results demonstrated that ARV-CU98 does cause some of the clinical signs in PEMS, including intestinal alterations and significantly lower relative bursa and liver weights. ARV-CU98 may contribute directly to PEMS by affecting the intestine, bursa, and liver and may contribute indirectly by increasing susceptibility to opportunistic pathogens that facilitate development of clinical PEMS.     

Isolation and characterization of avian rotaviruses

Rotaviruses were isolated from intestinal contents obtained from flocks of turkey poults and pheasant poults with diarrhea and from different age groups of chickens showing various signs of intestinal disorders. The incorporation of 5 micrograms trypsin/ml in the inoculum and medium was essential for virus isolation in chicken kidney cells. All isolates were identified as rotaviruses by fluorescent-antibody technique using a National Institutes of Health reference rotavirus antiserum against human rotavirus strain "D," Type 2. Negative-staining and transmission electron microscopy showed the presence of rotavirus particles. Furthermore, polyacrylamide gel electrophoresis pattern of viral RNA segments of our isolates confirmed that they are rotaviruses. Seven-day-old turkey poults could be infected with turkey and chicken rotavirus isolates; in contrast, chicks of the same age were refractory.     

Electron microscopic identification of viruses associated with poult enteritis in turkeys grown in California 1993-2003

Poult enteritis (PE) is one of the most common diseases seen in young turkey flocks. Since 1993, more than 1800 cases of suspected PE have been submitted for examination by negative stain electron microscopy; this has involved more than 2400 individual results, because in many cases more than one virus was identified; at least 1500 individual results were positive for viruses. Viruses have been identified in poults as young as 3 days and up to 9 wk of age. The most commonly found viruses are rotavirus-like viruses and small round viruses ranging from 15 nm to 30 nm, either alone or in combination. Reovirus, birnavirus, and adenovirus have also been detected. There has been no evidence to suggest the presence of coronaviruses. This report summarizes our findings.     

Protection of turkey poults from Bordetella avium infection and disease by pili and bacterins

The role of pili in protection against Bordetella avium infection in turkey poults was studied. An isolate that produced the largest number of pili under growth conditions developed in our laboratory was used for preparation of pili and bacterin and for challenge. The pili were isolated, purified, examined by electron microscopy, and tested for purity by gel electrophoresis. Poults were vaccinated with oil-adjuvant pili, formaldehyde- or merthiolate-inactivated bacterins, or a commercial bacterin. Poults were vaccinated once or twice subcutaneously at different ages and challenged intranasally with a pathogenic B. avium isolate 5 days following the last vaccination. A few vaccinated birds had very mild clinical signs. B. avium was isolated from the sinuses of a few vaccinated birds, and growth was scanty. The mean colony counts from tracheal sections was significantly higher (P less than 0.1) in unvaccinated challenged poults than in vaccinated challenged poults. It is postulated that B. avium pili are important immunogens in turkey poults.     

Viral agents associated with outbreaks of diarrhea in turkey flocks in Quebec.

The relative importance of various enteric viruses associated with diarrhea of turkey poults was investigated by an evaluation of specimens received since 1982. Specimens originated from one to eight week old turkey poults, with mild to severe diarrhea, from 114 flocks in 42 commercial operations located in southern Quebec. The acute phase of enteritis occurred usually in poults between two and four weeks of age. Clarified intestinal contents were examined by direct electron microscopy and enzyme immunoassays. Enzyme-linked immunosorbent assays were performed with antisera to bovine rotavirus group antigen, avian reovirus types 1 to 5, and the prototype strain of the turkey enteric coronavirus. The presence of viruses could be demonstrated by electron microscopy in 55.3% of the specimens, and at least five different viruses were incriminated either alone or in combination. The coronavirus was by far the most common enteric virus with a prevalence of 47.5%. By enzyme-linked immunosorbent assay, rotavirus, reovirus and turkey coronavirus were detected in 14.5%, 18.1% and 61.4% of the specimens, respectively. By electron microscopy, 56.6% of these cases were positive for at least one virus.     

Pathogenicity of turkey coronavirus in turkeys and chickens

We designed this study to compare the replication potential of turkey coronavirus (TCV) and its effect in chickens and turkeys and to study the effect of singleand combined infection of turkey poults with TCV and astrovirus. We studied the pathogenicity of TCV in experimentally inoculated turkey poults and chickens by observing the dinical signs and gross lesions. Two trials were conducted with 1-day-old and 4-wk-old specific-pathogen-free turkey poults and chickens. One-day-old turkey poults developed diarrhea at 48 hr postinoculation. Poults euthanatized at 3, 5, and 7 days postinoculation had flaccid, pale, and thin-walled intestines with watery contents. The 4-wk-old turkeys had no clinical signs or gross lesions. One-day-old and 4-wk-old chicks developed no clinical signs or gross lesions although the TCV was detected in gut contents of the birds throughout the experimental period (14 days). In another experiment, mean plasma D-xylose concentrations in 3-day-old turkey poults inoculated with TCV, turkey astrovirus, or a combination of both viruses were significantly lower than in the uninoculated controls.     

Turkey coryza: lack of correlation between plasmids and pathogenicity of Bordetella avium

Plasmids were removed from pathogenic Bordetella avium using a variety of treatments. The plasmid-cure rates depended on the treatment and isolate. Pathogenicity of B. avium in turkey poults was not altered by removal of plasmids.