Scanning electron, light, and immunofluorescent microscopy of coronaviral enteritis of turkeys (Bluecomb).

Intestinal sections from both experimental and field cases of turkey coronaviral enteritis (TCE) were examined by scanning electron microscopy and light microscopy through 10 days after inoculation and by a direct fluorescent antibody test for TCE through 12 days. Serums were collected for an indirect fluorescent antibody test for TCE through 160 days after inoculation. Lesions observed with the scanning electron microscopy were catarrhal enteritis with hemorrhage per diapedesis, epithelial desquamation, and villous atrophy which developed and regressed within 6 days after inoculation. Light microscopy demonstrated similar lesions, except that villus-to-crypt ratios remained depressed 10 days. The direct fluorescent antibody test demonstrated the presence of coronaviral antigen throughout the sampling period, and serum antibodies to TCE were present until at least 160 days, when the experiment was terminated.     

Serologic evidence of infectious bursal disease virus serotype II infection in Minnesota turkeys

Serum samples from seven randomly selected Minnesota turkey flocks were tested for antibodies to infectious bursal disease virus serotype I (Lukert strain, isolated from chickens, and North Carolina strain, isolated from turkeys) using a virus-neutralization (VN) test. All flocks were found to have low antibody titers to both Lukert and North Carolina strains. Five out of the seven flocks had high VN titers to the Missouri strain, a serotype II virus isolated from turkeys.     

Secretory antibodies against turkey coronaviral enteritis.

Studies on local immunity to transmissible coronaviral enteritis of turkeys (bluecomb) was made. Intestinal secretions and bile from affected birds contained secretory immunoglobulins against coronaviral antigen throughout the 6 months' duration of the experiment. Attempts to purify and to characterize the globulins in intestinal secretions and bile of the affected birds were made, using the techniques of gel filtration, DEAE chromatography, and immunoelectrophoresis. Class-specific anti-turkey IgA antiserum in the agar gel precipitin test further established the presence of IgA in the intestinal secretions and bile.     

Development of a competitive enzyme-linked immunosorbent assay for detection of turkey coronavirus antibodies

A competitive enzyme-linked immunosorbent assay (cELISA) was developed for detection of turkey coronavirus (TCV) antibodies. The cELISA utilized a recombinant baculovirus (Autographa californica nuclear polyhedrosis virus)-expressed TCV nucleocapsid (N) protein and biotin-labeled TCV N protein-specific monoclonal antibody. Sensitivity and specificity of the cELISA for detection of TCV antibodies were determined by comparison with the indirect fluorescent antibody test (IFAT) with 1269 reference, experimentally derived, and field-origin sera. Sera with discordant cELISA and IFAT results were further evaluated by western immunoblot analyses. The cELISA detected antibodies specific for TCV and infectious bronchitis virus, a closely related coronavirus, but did not detect antibodies specific for other avian viruses. A high degree of concordance was observed between the cELISA and IFAT; sensitivity and specificity of the cELISA relative to IFAT were 92.9% and 96.2%, respectively. Western immunoblot analyses provided additional evidence of cELISA specificity. The findings indicate that the cELISA is a rapid, sensitive, and specific serologic test for detection of TCV antibodies in turkeys.     

Incidence of histomonosis in turkeys in France since the bans of dimetridazole and nifursol

Between April 2003 and March 2005, 113 outbreaks of histomonosis were recorded in standard turkey farms in France, and 15 cases were recorded in turkey breeding centres. Most of the cases were in north-west France, the principal farming area for turkeys. The majority of the cases occurred during the hottest months, from April to September. Large numbers of cases occurred among birds from four to eight weeks of age, but there were some cases in three-week-old birds and some in birds up to 17 weeks of age. In most of the standard turkey flocks the mortality was less than 10 per cent, but it was above 30 per cent in nearly 20 per cent of the outbreaks. In the breeding flocks, the average mortality was 60.2 per cent. The size of the flocks, the sex of the birds and the age at which the first clinical signs appeared did not seem to influence the mortality.     

Hemorrhagic enteritis of turkeys in California: serologic study of hemorrhagic enteritis virus antibody with an enzyme-linked immunosorbent assay

The incidence of hemorrhagic enteritis (HE) infection in California turkeys was studied by testing 2220 turkey blood samples from 173 flocks for HE virus (HEV) antibody by the enzyme-linked immunosorbent assay (ELISA). Maternal antibody was detected at 1 day of age in all flocks tested, and it vanished after 3 weeks. Acquired HEV antibody appeared at 8 to 10 weeks, and 100% of the meat and breeder turkey flocks were positive after 11 weeks of age. HEV infection occurred earlier in the meat flocks than in the breeder flocks, and it also occurred earlier during summer than during the fall and winter months.     

Infection of turkeys with Ornithobacterium rhinotracheale and Mycoplasma synoviae

Within 1 mo, two separate outbreaks of respiratory disease occurred in two flocks on the multiage market turkey farm in Slovenia. More severe dinical signs and higher mortality were observed in male birds. Ornithobacterium rhinotracheale (ORT) was isolated in pure culture from tracheas of the affected birds in both outbreaks. Commercial enzyme-linked immunosorbent assay test showed the presence of antibodies to ORT in sera of birds from both clinically affected flocks and also in two flocks of younger birds without clinical sings. Immunoblotting with ORT culture isolated during the outbreak as an antigen confirmed the presence of antibodies to ORT in sera of turkeys of all four flocks examined. In addition, three different serologic assays also detected antibodies to Mycoplasma synoviae (MS) in three out of four flocks. The concomitant infection with MS did not show an obvious effect on mortality rates nor on the antibody response against ORT. Younger birds appeared to be less susceptible to ORT pathogenicity because in those flocks the infection was subclinical.     

Prevalence of enteropathogenic Escherichia coli in naturally occurring cases of poult enteritis-mortality syndrome

Enteropathogenic Escherichia coli (EPEC) previously were identified in poult enteritis-mortality syndrome (PEMS)-affected turkeys and associated as a cause of this disease. In the present study, the prevalence of EPEC in PEMS-affected turkeys was examined retrospectively with archived tissues and intestinal contents collected from 12 PEMS-affected turkey flocks in 1998. Formalin-fixed intestinal tissues were examined by light and electron microscopy for attaching and effacing (AE) lesions characteristic of EPEC, and frozen (-75 C) intestinal contents were examined for presence of EPEC. Escherichia coli isolates were characterized on the basis of epithelial cell attachment, fluorescent actin staining (FAS) test, and presence of E. coli attaching/effacing (EAE), shigalike toxin (SLT) type I, SLT II, and bundle-forming pilus (BFP) genes by polymerase chain reaction procedures. EPEC isolates were examined for pathogenicity and ability to induce AE lesions in experimentally inoculated young turkeys. AE lesions were identified by light microscopy in Giemsa-stained intestines from 7 of 12 PEMS-affected turkey flocks. Lesions consisted of bacterial microcolonies attached to epithelial surfaces with epithelial degeneration at sites of attachment and inflammatory infiltration of the lamina propria. Electron microscopy confirmed the identity of AE lesions in six of seven flocks determined to have AE lesions by light microscopy. EPEC were identified in 4 of 12 flocks on the basis of the presence of EAE genes a nd absence of SLT I and SLT II genes; all isolates lacked BFP genes. EPEC isolates produced AE lesions and variable mortality in turkeys coinfected with turkey coronavirus. In total, EPEC were associated with 10 of 12 (83%) naturally occurring PEMS cases on the basis of identification of AE lesions and/or EPEC isolates. These findings provide additional evidence suggesting a possible role for EPEC in the pathogenesis of PEMS.     

Investigation of field outbreaks of turkey haemorrhagic enteritis in Hungary

Epidemiological, pathological, serological and virological investigations are reported on turkey haemorrhagic enteritis virus (THEV) infection in Hungarian turkey flocks. The pathogenesis of infection in experimentally infected turkeys and chickens, as well as the usefulness of polymerase chain reaction (PCR)/sequencing method for epidemiological investigation and for the differentiation of vaccine and field strains of THEV was also studied. Since the first recognition of the disease in Hungary in the late 1970s, until recently the disease has been diagnosed sporadically in its mild form. In the last few years (2000-2005), however, the number of outbreaks and the severity of the disease increased (9-23 affected flocks/year). Most of the outbreaks occurred at the age of 6 to 8 weeks and was complicated with Escherichia coli infection. The antibody levels to THEV in turkey flocks gradually declined till 5-7 weeks of age, and then they increased sharply due to natural infection with THEV. The immune response to vaccination (at 5 weeks of age) showed no significant antibody level increase one week postvaccination, but four weeks later the antibody level reached high values and then remained at this high level. The agar gel immunodiffusion (AGID) test to detect turkey adenovirus A (TAdV-A) antigen and PCR methods for THEV-specific DNA gave similarly positive results if spleens with pathognomonic lesions were tested; however, PCR proved to be more sensitive in cases with less characteristic pathological lesions. Nucleotide sequence alignment of PCR products amplified from Hungarian field strains and the Domermuth vaccine strain and that of the published THEV hexon sequences in GenBank database revealed slight differences between the sequences.     

Identification and phylogenetic diversity of parvovirus circulating in commercial chicken and turkey flocks in Croatia

Phylogenetic diversity of parvovirus detected in commercial chicken and turkey flocks is described. Nine chicken and six turkey flocks from Croatian farms were tested for parvovirus presence. Intestinal samples from one turkey and seven chicken flocks were found positive, and were sequenced. Natural parvovirus infection was more frequently detected in chickens than in turkeys examined in this study. Sequence analysis of 400 nucleotide fragments of the nonstructural gene (NS) showed that our sequences had more similarity with chicken parvovirus (ChPV) (92.3%-99.7%) than turkey parvovirus (TuPV) (89.5%-98.9%) strains. Phylogenetic analysis grouped our sequences in two clades. Also, the higher prevalence of ChPV than TuPV in tested flocks was defined. The necropsy findings suggested a malabsorption syndrome followed by a preascitic condition. Further research of parvovirus infection, pathogenesis, and the possibility of its association with poult enteritis and mortality syndrome (PEMS) and runting and stunting syndrome (RSS) is needed to clarify its significance as an agent of enteric disease.     

Serologic evidence of infectious bursal disease virus infection in Iowa turkeys

Two infectious bursal disease viruses (IBDVs)--a cell-culture-adapted chicken strain designated Edgar strain and a recent isolate from turkeys in Missouri--were used to assay sera from 10 Iowa turkey flocks collected between 1 and 16 weeks of age. The two viruses were serologically distinct in cross-neutralization tests. For all flocks, a similar serologic pattern was found consisting of (1) low maternal antibody titers to turkey IBDV and occasionally to Edgar strain IBDV between 1 and 3 weeks of age, (2) a period of very low or no detectable titers between 3 and 7 weeks of age, and (3) sharply rising high titers to turkey IBDV with low titers to Edgar IBDV beginning at 5 to 8 weeks of age. These findings indicate that infection with IBDV of the serotype represented in this study by the turkey isolate is common in Iowa turkey flocks, whereas infection with IBDV represented by Edgar strain is uncommon. Infection occurred between 3 and 7 weeks of age during the late brooding period or after birds had been moved to an intermediate growing facility. All flocks developed complicated respiratory disease with excessive mortality caused by Escherichia coli septicemia, typically between 3 and 6 weeks of age. Although there was a temporal relationship between IBDV infection and respiratory disease, the possible role of IBDV in the process is unknown.     

Experimental infection of chickens and turkeys with Mycoplasma gallisepticum reference strain S6 and North Carolina field isolate RAPD type B

During an epidemic of mycoplasmosis in chicken and turkey flocks in North Carolina between 1999 and 2001, isolates of Mycoplasma gallisepticum (MG) from affected flocks were characterized by random amplification of polymorphic DNA (RAPD), and eight distinct RAPD types were identified. MG RAPD type B accounted for more than 90% of the isolates and was associated with moderate-to-severe clinical signs and mortality. The virulence of MG RAPD type B for chickens and turkeys was compared with sham-inoculated negative controls and MG S6 (a virulent strain)-inoculated positive controls. Clinical signs occurred in chickens and turkeys inoculated with either MG RAPD type B or MG S6. However, they were not as frequent or severe as those seen in naturally affected flocks, and there was no mortality in the experimental groups. Based on gross and microscopic findings, MG RAPD type B was equal to or more virulent than MG S6. All MG-inoculated birds were culture and PCR positive at 7 and 14 days postinoculation (PI). Among serological tests, the serum plate agglutination test was positive for the majority of chickens and turkeys (58%-100%) infected with either strain of MG at both 7 and 14 days PI. The hemagglutination inhibition test was negative for all birds at 7 days PI and positive for a few chickens (8%-17%) and several turkey sera (40%-60%) at 14 days PI. Only a single serum was positive by enzyme-linked immunosorbent assay (an MG S6-infected turkey) at 14 days PI.     

Detection of antibody to turkey coronavirus by antibody-capture enzyme-linked immunosorbent assay utilizing infectious bronchitis virus antigen

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.     

Prevalence and risk factors for Salmonella spp. and Campylobacter spp. caecal colonization in broiler chicken and turkey flocks slaughtered in Quebec, Canada

We conducted an observational study to estimate prevalence and risk factors for Salmonella spp. and Campylobacter spp. caecal colonization in poultry. Eighty-one broiler chicken and 59 turkey flocks selected among flocks slaughtered in the province of Quebec, Canada, were included in the study. Flock status was evaluated by culturing pooled caecal contents from about 30 birds per flock. Exposure to potential risk factors was evaluated with a questionnaire. Odds ratios were computed using multivariable logistic regression.

The prevalence of Salmonella-positive flocks was 50% (95% CI: 37, 64) for chickens and 54% (95% CI: 39, 70) for turkeys, respectively. Odds of Salmonella colonization were 2.6 times greater for chicken flocks which failed to lock the chicken house permanently. In turkeys, odds of Salmonella colonization were 4.8–7.7 times greater for flocks which failed to be raised by ≤2 producers with no other visitors allowed onto the premises, or origin from a hatchery.

The prevalence of Campylobacter-positive flocks was 35% (95% CI: 22, 49) for chickens and 46% (95% CI: 30, 62) for turkeys. Odds of colonization were 4.1 times higher for chicken flocks raised on farms with professional rodent control and 5.2 times higher for flocks with manure heap >200 m from the poultry house, and also increased with the number of birds raised per year on the farm and with the age at slaughter. For turkeys, odds of Campylobacter flock colonization were 3.2 times greater in flocks having a manure heap at ≤200 m from poultry house and 4.2 times greater in flocks drinking unchlorinated water.     

Drug use and antimicrobial resistance among Escherichia coli and Enterococcus spp. isolates from chicken and turkey flocks slaughtered in Quebec,Canada

An observational study was conducted of chicken and turkey flocks slaughtered at federal processing plants in the province of Quebec, Canada. The objectives were to estimate prevalence of drug use at hatchery and on farm and to identify antimicrobial resistance (AMR) in cecal Escherichia coli and Enterococcus spp. isolates and factors associated with AMR. Eighty-two chicken flocks and 59 turkey flocks were sampled. At the hatchery, the most used antimicrobial was ceftiofur in chickens (76% of flocks) and spectinomycin in turkeys (42% of flocks). Virginiamycin was the antimicrobial most frequently added to the feed in both chicken and turkey flocks. At least 1 E. coli isolate resistant to third-generation cephalosporins was present in all chicken flocks and in a third of turkey flocks. Resistance to tetracycline, streptomycin, and sulfisoxazole was detected in > 90% of flocks for E. coli isolates. Antimicrobial resistance (AMR) was observed to bacitracin, erythromycin, lincomycin, quinupristin-dalfopristin, and tetracycline in both chicken and turkey flocks for Enterococcus spp. isolates. No resistance to vancomycin was observed. The use of ceftiofur at hatchery was significantly associated with the proportion of ceftiofur-resistant E. coli isolates in chicken flocks. In turkey flocks, ceftiofur resistance was more frequent when turkeys were placed on litter previously used by chickens. Associations between drug use and resistance were observed with tetracycline (turkey) in E. coli isolates and with bacitracin (chicken and turkey), gentamicin (turkey), and tylosin (chicken) in Enterococcus spp. isolates. Further studies are needed to provide producers and veterinarians with alternative management practices and tools in order to reduce the use of antimicrobial feed additives in poultry.     

Serological survey of infectious bursal disease virus serotypes 1 and 2 in California turkeys

The prevalence of two infectious bursal disease (IBD) viruses--serotype 1, of chicken origin, and serotype 2, of turkey origin--was studied in California turkeys. Serum samples were collected from 15 turkey flocks representing nine counties. The virus-neutralization test was used. Overall, only 15% of 342 samples were positive for serotype 1, whereas 89% were positive for serotype 2. However, 12 out of 15 flocks had a least one sample positive for serotype 1, and 14 out of 15 flocks had at least one sample positive for serotype 2. Flocks with antibodies to serotype 1 had low geometric mean titers (GMTs) (1.5 to 8.8) and low-to-medium prevalence rates (5 to 53.3%). Flocks with antibodies to serotype 2 had high GMTs (36.8 to 2048) and high prevalence rates (60 to 100%). Results of this study lead us to question the efficacy of IBD vaccination in turkeys.