Field evaluation of maternal antibody transfer from breeder turkey hens to egg yolks, egg whites, and poults.

Immunoglobulins, which are passed vertically from hens to their progeny, are first present in the eggs but with time also in the developing embryos and eventually in the serum of hatching chicks, and have protective function during embryogenesis and in the first few weeks of birds' life, before the immune system becomes fully efficient. Considering the above fact, the aim of this study was to determine total levels of IgM and IgY as well as specific IgY antibody titers against selected pathogens in the serum of breeder turkeys and their progeny, as well as in egg yolks and egg whites. Study results demonstrated that the level of IgY antibodies in the serum of turkey breeder hens reached 22.04 mg/mL on average in the whole egg laying cycle. In addition, the mean transfer percentage of IgY antibodies from turkey layers to their progeny reached approximately 31.4%, but the level of this transfer differed depending on pathogen character and accounted for 33.2%, 51.9%, 45.1%, and 44.3% in the case of antibodies against avian metapneumoviruses, Newcastle disease virus, Ornithobacterium rhinortacheale, and Pasteurella multocida, respectively. Antibody percentage transfer differed also as affected by the stage of the egg production cycle. Study results confirmed the earlier observed dependency concerning the class of antibodies transferred to eggs from laying hens, and while the IgY were mainly detected in the egg yolk extracts, the IgM were found only in egg white extracts; in comparison to IgY, the IgM antibodies were not transferred to the serum of turkey poults. To our best knowledge, this is the first study that describes in detail the phenomenon of maternal antibody transfer in turkeys.

Comparative pathomorphological, bacteriological and serological examination of broiler breeders and pheasants experimentally infected with Ornithobacterium rhinotracheale.

The aim of the investigations was to determine the influence of Ornithobacterium rhinotracheale (ORT) on the development of pathomorphological lesions in the respiratory organs and on the health status of experimentally infected broiler breeders and pheasants from the rearing stage. There was no evidence of clinical signs in infected broiler breeder hens nor in the group of infected pheasants except for one bird in the latter group which exhibited slower movement and gasping. The frequency and intensity of pathomorphological lesions were higher in pheasants. The gross pathology findings were characterized mainly by redness of the mucosa of the upper respiratory tract and accumulation of mucous content in the nasal cavities, infraorbital sinuses, larynx and trachea. Histopathology confirmed the presence of inflammation of the upper respiratory tract. Lesions in the lungs included hyperaemia, granulomatous and fibrinous pneumonia. ORT was reisolated only from the group of infected pheasants. Reisolation was successful from the respiratory organs (trachea, larynx, infraorbital sinuses, and lungs) of eight out of 10 infected birds. The serological response in both species was characterized by rapid production of specific antibodies that reached a maximum level in the blood in the first week after experimental infection. The antibody titres decreased gradually and were maintained at a stable level until the 12th week after inoculation. Fourteen weeks post-inoculation specific antibodies could not be detected by enzyme-linked immunosorbent assay.

Conditions that induce biofilm production by Ornithobacterium rhinotracheale.

Ornithobacterium rhinotracheale (ORT) is a Gram-negative bacillus that causes respiratory disease in birds, and directly affects the poultry industry. The mechanisms behind these infections are not completely known. Currently, its capacity to form biofilms on inert surfaces has been reported; however, the conditions for biofilm development have not been described yet. The present work was aimed at identifying the conditions that enhance in vitro biofilm formation and development by ORT. For this, serovars A-E were analysed to assess their ability to induce biofilm development on 96-well flat-bottom polystyrene microtitre plates under diverse conditions: temperature, incubation time, and CO2 concentration. The results obtained showed not only that all serovars have the ability to produce in vitro biofilms, but also that the optimal conditions for biofilm density were 40°C after 72 h at an elevated CO2 concentration. In conclusion, ORT biofilm formation depends on the environmental conditions and may contribute to the persistence of this microorganism.

Biochemical and immunological responses of young turkeys to vaccination against Ornithobacterium rhinotraheale and different levels of dietary methionine.

The objective of this study was to verify the hypothesis that increasing levels of dietary methionine can stimulate the mechanisms of cell-mediated and humoral immunity in young turkeys. The blood and organs involved in cell-mediated and humoral immune responses were analyzed in 8-week-old turkeys that had been vaccinated against Ornithobacterium rhinotraheale (ORT) infection (on days 17 and 48). The birds were fed diets with a low (LM), medium (MM) and high (HM) methionine content (0.45 and 0.40%, 0.60 and 0.51%, 0.71 and 0.57% in weeks 1 - 4 and 5 - 8, respectively). Dietary methionine supplementation led to a significant increase in body weights of turkeys at 56 days of age, from 3532 g in group LM to 3720 g in group MM and 3760 g in group HM (p=0.001). A significant increase in vaccine-induced antibody titers against ORT was noted in group HM relative to group LM (p=0.006). Increasing levels of methionine had no significant effect on total serum IgG nor IgM levels and most serum biochemical parameters, TP, ALB, GLOB, GLU, AST, ALP, P and Ca. In comparison with group LM, group HM turkeys were characterized by a lower percentage of IgM⁺ B cell subpopulation in the blood and bursa of Fabricius. The percentages of CD4⁺ and CD8⁺ T cell subpopulations in the bursa of Fabricius in group HM were significantly different from those found in groups LM and MM. The highest percentages of CD4⁺ T cells and CD8⁺ T cells in the spleen were observed in groups LM (p<0.001) and HM (p=0.04), respectively. The differences were statistically significant relative to the remaining groups. Turkeys of group LM were characterized by a lower CD4⁺ T cell percentage in the thymus (p<0.001) and a lower CD8⁺ T cell percentage in the cecal tonsils (CTs) (p<0.01). Vaccination against ORT resulted in a significant increase in the percentage of CD4⁺CD8⁺ T cell subpopulation and a decrease in the percentage of CD8⁺ T cell subset in the spleen.

Immunity to bacterial infection in the chicken.

Bacterial infections remain important to the poultry industry both in terms of animal and public health, the latter due to the importance of poultry as a source of foodborne bacterial zoonoses such as Salmonella and Campylobacter. As such, much focus of research to the immune response to bacterial infection has been to Salmonella. In this review we will focus on how research on avian salmonellosis has developed our understanding of immunity to bacteria in the chicken from understanding the role of TLRs in recognition of bacterial pathogens, through the role of heterophils, macrophages and γδ lymphocytes in innate immunity and activation of adaptive responses to the role of cellular and humoral immunity in immune clearance and protection. What is known of the immune response to other bacterial infections and in particular infections that have emerged recently as major problems in poultry production including Campylobacter jejuni, Avian Pathogenic Escherichia coli, Ornithobacterium rhinotracheale and Clostridium perfringens are discussed.

Improved broiler performance associated with Ornithobacterium rhinotracheale vaccination in breeders.

During a field study in 2010 the daily growth, feed conversion, first-week mortality, broiler loss due to mortality and slaughterhouse condemnation, and production index were monitored in 100 broiler flocks derived from four breeder farms vaccinated with Nobilis OR inac and four Ornithobacterium rhinotracheale-unvaccinated breeder farms of the same organization in Belgium. Other parameters related to the broiler flocks, such as flock size, season, age of the breeders, and corresponding breeder farms, were also noted. All gathered data were examined with ANOVA, linear correlation, and linear regression analyses. Results demonstrated a significant 22.3% lower broiler loss and a significant 3.9% higher production index in the broiler flocks derived from breeders vaccinated with Nobilis OR inac. These results confirm field observations obtained in 1999, thereby providing further evidence for an effect of O. rhinotracheale vaccination in breeders with regard to the improved performance of broilers.

The effect of vaccination of pullets against Ornithobacterium rhinotracheale infection.

The effect of vaccination of chickens with different inactivated vaccines against experimental Ornithobacterium rhinotracheale challenge was investigated. Eight different vaccines, with different inactivating substances (Formalin and thiomersal) and with or without adjuvant (mineral oil, alum and aluminium hydroxide gel), were produced. Following vaccination of experimental chickens at week 8 with formalin-inactivated mineral oil adjuvant bacterin, the mean O. rhinotracheale antibody titres rose to 5.88 2log 21 days after primary vaccination and enhanced to a titre of 6.59 2log 21 days after booster vaccination. The bacterin in mineral oil adjuvant induced the highest serologic response and a significant decrease of lesions such as air sacculitis and pneumonia in vaccinated birds compared with the unvaccinated challenge control birds. The bacterin in either alum or aluminium hydroxide gel adjuvant induced a moderate serologic response and a decrease of lesions compared with the unvaccinated challenge controls. The study showed that vaccination of layer chicken at the eighth week followed by a booster dose at the 12th week of age can effectively protect against O. rhinotracheale infections.

Seroprevalence of Ornithobacterium rhinotracheale infection in broiler and broiler breeder chickens in West Azerbaijan Province, Iran.

Ornithobacterium rhinotracheale is a pleomorphic Gram-negative rod shaped bacterium of the rRNA superfamily V that is associated with respiratory disease in poultry. This study was conducted to determine the seroprevalence of O. rhinotracheale infection in broiler and broiler breeder chickens in West Azerbaijan (Urmia lake region) by using a commercial enzyme-linked immunosorbent assay. In this study, 463 serum samples were obtained from 50 broiler flocks and 472 blood sera from 42 broiler breeder flocks. Results showed that 41 broiler flocks (82%) and 39 broiler breeder flocks (92.8%) were positive. Ornithobacterium rhinotracheale antibodies were detected in 205 (44.2%) of the 463 broiler serum samples. Of the 472 blood sera examined from broiler breeder, 340 (72%) were positive. The results of this study indicated that the prevalence of O. rhinotracheale antibodies is high in the broiler and broiler breeder flocks in West Azerbaijan.

Vaccine potential of recombinant Ornithobacterium rhinotracheale antigens.

Ornithobacterium rhinotracheale is a pathogen involved in respiratory infection and systemic disease in poultry. Previously, eight potential vaccine candidates were identified that induced cross-protective immunity when administered to chickens as a multi-component vaccine. In this study, we analyzed the immunogenicity of these eight recombinant proteins by subunit vaccination, and characterized the different proteins and corresponding genes more thoroughly by sequencing, in vitro expression analysis, and cellular localization experiments. We found, that all genes encoding the eight antigens were highly conserved among different O. rhinotracheale serotypes, but the different antigens were not expressed by all serotypes. Cellular fractionation experiments indicated that the majority of the antigens are predominantly located in the outer membrane fraction. Vaccination of chickens with single-antigen vaccines demonstrated that the Or77 antigen was protective against serotypes that expressed Or77 in vitro, suggesting that the protein has strong potential as a vaccine antigen. Furthermore, immunization with four-component subunit vaccines indicated the existence of immunogenic synergism between the candidate vaccine antigens.

Successful selection of cross-protective vaccine candidates for Ornithobacterium rhinotracheale infection.

Ornithobacterium rhinotracheale is a bacterial pathogen known for causing respiratory disease in poultry. In this study, we demonstrate for the first time that cross-protective immunity against different O. rhinotracheale serotypes can be induced by live vaccination. Sera from these live-vaccinated and cross-protected birds were used to identify new vaccine targets by screening an O. rhinotracheale expression library. Out of 20,000 screened plaques, a total of 30 cross-reactive clones were selected for further analysis. Western blot analysis and DNA sequencing identified eight different open reading frames. The genes encoding the eight cross-reactive antigens were amplified, cloned in an expression vector, and expressed in Escherichia coli. Purified recombinant proteins with a molecular mass ranging from 35.9 kDa to 62.9 kDa were mixed and tested as a subunit vaccine for (cross-)protection against challenge with homologous and heterologous O. rhinotracheale serotypes in chickens. Subunit vaccination resulted in the production of antibodies reactive to the recombinant proteins on Western blot, and this eight-valent vaccine conferred both homologous and heterologous protection against O. rhinotracheale challenge in chickens.

Key role of Chlamydophila psittaci on Belgian turkey farms in association with other respiratory pathogens.

Two hundred turkey sera from eight Belgian and two French farms were tested for the presence of antibodies against avian pneumovirus (APV), Ornithobacterium rhinotracheale (ORT), Mycoplasma gallisepticum, Mycoplasma meleagridis and Chlamydophila psittaci. At slaughter, C. psittaci, APV and ORT antibodies were detected in 94, 34 and 6.5% of the turkeys, respectively. No antibodies against M. gallisepticum or M. meleagridis were present. Additionally, turkeys on three Belgian farms were examined from production onset until slaughter using both serology and antigen or gene detection. All farms experienced two C. psittaci infection waves, at 3-6 and 8-12 weeks of age. Each first infection wave was closely followed by an ORT infection starting at the age of 6-8 weeks, which was still detectable when the second C. psittaci infection waves started. Animals on farm A were not vaccinated against APV leading to an APV subtype B outbreak accompanying the first C. psittaci infection wave. Despite subtype A APV vaccination on farms B and C, the second C. psittaci infection waves were accompanied (farm B) or followed (farm C) by a subtype B APV infection. On all farms respiratory signs always appeared together with a proven C. psittaci, APV and/or ORT infection. This study suggests an association between C. psittaci, APV and ORT, and indicates the multi-factorial aetiology of respiratory infections in commercial turkeys. All three pathogens should be considered when developing prevention strategies for respiratory disease.

Diagnosis of Ornithobacterium rhinotracheale infection in chickens by ELISA.

Specific Ornithobacterium rhinotracheale (ORT) antibodies were determined in serum samples of 24 clinically infected broiler flocks of different ages (1-42 d) and 11 broiler-breeding flocks (at ages between 26-56 w) by ELISA. Two commercially available kits were separately assessed. The BioCheck ELISA kit was used for testing 363 serum samples representing 12 broiler flocks, where 74 samples (20.3 %) were found to be positive and 49 (13.5 %) were suspected. The IDEXX ELISA kit was used for testing 148 serum samples representing different 12 broiler flocks, where 115 samples (77.7 %) were positive. Testing of additional 70 serum samples from 5 broiler- breeder flocks, associated with drop in egg production (1-4.5 %) at different ages, by BioCheck ELISA kit revealed that 78.5 % of the samples were positive and 21.4% were suspected. On the other hand, 338 serum samples representing 6 broiler-breeder flocks, associated also with egg drop, showed a 84.6 % rate of positive reaction, when tested by IDEXX ELISA kit. Positive serology correlated well with the clinical manifestations and isolation of the organism, which substantiates the reliability of the used kits in diagnosis of the disease.

Studies of the transmission routes of Ornithobacterium rhinotracheale and immunoprophylaxis to prevent infection in young meat turkeys.

The importance and prevention of the horizontal as well as the vertical transmission of Ornithobacterium rhinotracheale were investigated. In our first experiment we observed that specific-pathogen-free broiler chickens that were placed in hatching incubators at a commercial turkey hatchery during hatch showed respiratory tract lesions at postmortem examination that were positive for O. rhinotracheale by bacteriology and immunohistology. It appeared that vertical transmission occurred and that horizontal transmission of O. rhinotracheale is possible. In a second experiment, the turkeys derived from vaccinated parents showed significantly fewer respiratory tract lesions at postmortem examination at 16 days of age than the birds derived from nonvaccinated parents. In a third experiment, all vaccinated young birds, regardless of the vaccination state of their parents, showed significantly fewer respiratory tract lesions at 6 wk of age. We concluded that vaccination of the breeders reduces vertical transmission and that vaccination of the progeny is needed to resist challenge at 6 wk of age.

The use of a bacterin vaccine in broiler breeders for the control of Ornithobacterium rhinotracheale in commercial broilers.

Ornithobacterium rhinotracheale (ORT) is a recently identified bacterial pathogen of poultry, linked to the respiratory disease complex of broilers and the economic losses associated with that disease complex. Present control measures applied for the disease include the continuous use of in-feed antibiotics. A recently developed bacterin vaccine that is applied to broiler-breeder hens to pass on protective immunity to their broiler progeny was tested under large-scale commercial conditions in South Africa. An indirect ELISA test for antibodies to ORT, optimised for use in South Africa, was used to determine antibody levels in breeders and broilers. ELISA test results showed that the vaccine stimulated the development of high antibody titre levels in broiler breeders. The efficacy of the vaccine in protecting the progeny of these birds from ORT challenge could not be determined during the trial, although the progeny of vaccinated hens appeared to perform slightly better under commercial conditions than the progeny of unvaccinated hens.

Prevalence of antibodies against Ornithobacterium rhinotracheale in broilers and breeders in Southern Brazil.

In this investigation, we determined the prevalence of the Ornithobacerium rhinotracheale (ORT) infection in broilers and broiler breeders in southern Brazil. We also correlated the presence of antibodies in broilers with performance. Sera from 1550 broilers from 50 flocks were collected during the slaughter time in nine companies with federal veterinary inspection of the state of Rio Grande do Sul. Sera from 480 meat-type breeders of 40 flocks from 14 companies in southern Brazil were also analyzed by enzyme-linked immunosorbent assay, and the prevalence of antibodies was determined. The prevalence of ORT antibodies in broiler flocks was 63.83%, but in each individual flock only 6.52% of the birds were positive. The prevalence in broiler breeder flocks was 100.00%, and in each individual flock 94.62% of the birds were positive. There was a positive correlation between the presence of antibodies to ORT and decreased body weight in broilers. There was no significant correlation between presence of antibodies to ORT and age, lineage, efficiency index, feed conversion, and mortality. There was a positive correlation between the presence of respiratory signs and antibodies to ORT, although the reverse correlation was not significant. These results confirm that ORT is present and widespread in broilers and broiler breeders in southern Brazil.