Respiratory Disease In Turkeys Research Articles

A Study on Isolation and Molecular Identification of Bordetella avium from Iranian Commercial and Backyard Broiler Turkeys within 2016-2018.

Bordetellosis or turkey coryza, caused by Bordetella avium, has been an issue for turkey industry since its first description in 1967 when it was reported for the first time. Bordetella avium causes a highly contagious upper respiratory disease in turkeys. Therefore, this study aimed to isolate and characterize this species from commercial and backyard turkeys in Tehran, Isfahan, and Northern provinces of Iran. For the purpose of the study, 625 tracheal swabs were taken from 425 commercial poults and 200 backyard poults aged 2-6 weeks from September 2016 to September 2018. The swabs were immediately plated on MacConkey and blood agar plates and then pooled (5 swabs/pool) in tubes, containing 2 mL distilled water, to perform direct polymerase chain reaction (PCR) for the identification of B. avium. A total of 17 swab pools were found to be positive for B. avium. A subset of seven positive samples were sequenced for the flanking region of piuA gene. The analysis of the sequences indicated that the sequences were 98%, 96%, and 98% similar to B. avium 197N (AM167904.1), 4142 (AY925058.1), and 4156 (AY925068.1) sequences, respectively. To the best of our knowledge, the current study is the first attempt toward the molecular detection and characterization of B. avium in Iran. It is highly recommended to perform further studies to isolate, characterize, and differentiate the regional isolates in order to help the developing turkey industry of Iran meet the increasing demands for protein in the diet of the citizenry.

Draft Genome Sequences of Six Bordetella hinzii Isolates Acquired from Avian and Mammalian Hosts.

Bordetella hinzii is a Gram-negative bacterium known to infect poultry, humans, rabbits, and rodents. It is an opportunistic pathogen in immunocompromised humans, and some strains cause mild to moderate respiratory disease in turkeys. Little is known as to the degree of genetic diversity within the species or the genetic basis for virulence. Here, we report the genome sequences of six isolates of B. hinzii acquired from humans, rabbits, or turkeys. These data provide a framework for refining the population structure of the genus, establishing relationships among genetically distinct isolates, and developing an understanding of the possible virulence mechanisms of the bacterium.

The pathogenicity of avian metapneumovirus subtype C wild bird isolates in domestic turkeys

BackgroundAvian metapneumovirus subtype C (aMPV/C) causes severe upper respiratory disease in turkeys. Previous report revealed the presence of aMPV/C in wild birds in the southeast regions of the U.S.MethodsIn this study, aMPV/C positive oral swabs from American coots (AC) and Canada geese (CG) were passaged three times in the respiratory tract of specific pathogen free (SPF) turkeys and used as aMPV/C P3 virus isolates in subsequent studies.ResultsWild bird P3 isolates showed similar growth characteristics when compared to virulent aMPV/C in chicken embryo fibroblast ( CEF) cell cultures and their glycoprotein G gene sequence was closely related to the G gene of aMPV/C Colorado reference virus. Three-day-old commercial or SPF turkeys were inoculated oculonasally with wild bird aMPV/C P3 isolates. At 5 and 7 days post-inoculation (DPI), severe clinical signs were observed in both of the AC and CG virus-exposed groups. Viral RNA was detected in tracheal swabs by reverse transcriptase polymerase chain reaction (RT-PCR). In addition, immunohistochemistry showed virus replication in the nasal turbinate and trachea. All virus-exposed turkeys developed positive antibody response by 14 DPI.ConclusionsOur data demonstrate that aMPV/C wild bird isolates induced typical aMPV/C disease in the domestic turkeys.

Effects of Cyclosporin A induced T-lymphocyte depletion on the course of avian Metapneumovirus (aMPV) infection in turkeys

The avian Metapneumovirus (aMPV) causes an economically important acute respiratory disease in turkeys (turkey rhinotracheitis, TRT). While antibodies were shown to be insufficient for protection against aMPV-infection, the role of T-lymphocytes in the control of aMPV-infection is not clear. In this study we investigated the role of T-lymphocytes in aMPV-pathogenesis in a T-cell-suppression model in turkeys. T-cell-intact turkeys and turkeys partly depleted of functional CD4(+) and CD8(+) T-lymphocytes by Cyclosporin A (CsA) treatment were inoculated with the virulent aMPV subtype A strain BUT 8544. CsA-treatment resulted in a significant reduction of absolute numbers of circulating CD4(+) and CD8alpha(+) T-lymphocytes by up to 82 and 65%, respectively (P<0.05). Proportions of proliferating T-cells within mitogen-stimulated peripheral blood mononuclear cells were reduced by similar levels in CsA-treated birds compared to untreated controls (P<0.05). CsA-treated turkeys showed delayed recovery from aMPV-induced clinical signs and histopathological lesions and a prolonged detection of aMPV in choanal swabs. The results of this study show that T-lymphocytes play an important role in the control of primary aMPV-infection in turkeys.

Investigations on the protective role of passively transferred antibodies against avian metapneumovirus infection in turkeys

The avian metapneumovirus (aMPV) is the causative agent of an acute respiratory disease in turkeys, which causes considerable economic losses to the poultry industry. Currently attenuated live and inactivated vaccines are widely used to control the disease, but vaccine breaks are frequently observed. For improvement of current vaccination strategies it is necessary to gain enhanced knowledge of the immune mechanisms against aMPV infection. Field observations suggest that vaccine-induced aMPV-specific antibodies are not indicative for protection. In the present study we investigated the role of antibodies in protection of turkeys against aMPV. In two experiments, commercial turkey poults received aMPV-specific antibodies by intravenous injection. The antibody transfer resulted in increased antibody levels in the sera. Virus-specific antibodies were also detected on mucosal surfaces such as the trachea, conjunctivae and gall bladder. Turkeys were subsequently challenged with a virulent aMPV subtype A strain. Development of clinical signs, virus detection by polymerase chain reaction and histopathological changes of tracheal mucosa in challenged turkeys with and without passively transferred antibodies were comparable with each other. Our results suggest that humoral immunity does not provide sufficient protection against aMPV infection. Thus, the measurement of vaccine-induced aMPV antibody response may not be considered as an adequate indicator of vaccine efficacy. Further research on the protective role of cell-mediated immune mechanisms is necessary to improve current vaccine strategies.

Effect of multiple- and single-day enrofloxacin medications against dual experimental infection with avian pneumovirus and Escherichia coli in turkeys

Escherichia coli infections are a common cause of respiratory disease in turkeys. To control these respiratory infections in turkeys, drinking water treatment with enrofloxacin is widely used. The current treatment schedule advises a 5-d treatment at 10 mg/kg of BW. Because enrofloxacin exerts a concentration-dependent activity, it might be useful to provide this 50 mg/kg total dose in a single-day treatment regimen. Therefore, we assessed whether a single-day treatment schedule with 50 mg/kg of BW was clinically equivalent to the advised multiple-day treatment schedule with 10 mg/kg of BW for 5 d. For this purpose, 3 groups of seventeen 22-d-old turkeys were experimentally inoculated with avian metapneumovirus and 3 d later with E. coli. One group received 10 mg of enrofloxacin/kg of BW in the drinking water for 5 successive days. The second group received 50 mg of enrofloxacin/kg of BW for 20 h in the drinking water. The third group was enclosed as an untreated positive control group. Both the multiple- and the single-day enrofloxacin treatment regimens reduced E. coli multiplication in the respiratory tract tissues (turbinates, trachea and lung), but the 5-d treatment with 10 mg of enrofloxacin/kg in turkeys provided the best results by shortening the course of clinical disease, by eliminating E. coli from the respiratory tract without remultiplication, and by reducing macroscopic lesions. The efficacy of the single-day treatment did not equal that of the 5-d treatment, possibly by not eliminating E. coli from the respiratory organs, which made it possible for the remaining bacteria to reemerge in those organs. None of the used treatment regimens promoted the selection of bacterial clones with reduced susceptibility or resistance.

Recovery of avian metapneumovirus subgroup C from cDNA: cross-recognition of avian and human metapneumovirus support proteins.

Avian metapneumovirus (AMPV) causes an acute respiratory disease in turkeys and is associated with "swollen head syndrome" in chickens, contributing to significant economic losses for the U.S. poultry industry. With a long-term goal of developing a better vaccine for controlling AMPV in the United States, we established a reverse genetics system to produce infectious AMPV of subgroup C entirely from cDNA. A cDNA clone encoding the entire 14,150-nucleotide genome of AMPV subgroup C strain Colorado (AMPV/CO) was generated by assembling five cDNA fragments between the T7 RNA polymerase promoter and the autocatalytic hepatitis delta virus ribozyme of a transcription plasmid, pBR 322. Transfection of this plasmid, along with the expression plasmids encoding the N, P, M2-1, and L proteins of AMPV/CO, into cells stably expressing T7 RNA polymerase resulted in the recovery of infectious AMPV/CO. Characterization of the recombinant AMPV/CO showed that its growth properties in tissue culture were similar to those of the parental virus. The potential of AMPV/CO to serve as a viral vector was also assessed by generating another recombinant virus, rAMPV/CO-GFP, that expressed the enhanced green fluorescent protein (GFP) as a foreign protein. Interestingly, GFP-expressing AMPV and GFP-expressing human metapneumovirus (HMPV) could be recovered using the support plasmids of either virus, denoting that the genome promoters are conserved between the two metapneumoviruses and can be cross-recognized by the polymerase complex proteins of either virus. These results indicate a close functional relationship between AMPV/CO and HMPV.

Evaluation of the persistence and gene expression of an anti-Chlamydophila psittaci DNA vaccine in turkey muscle

BackgroundDNA vaccination has been shown to elicit specific cellular and humoral immune responses to many different agents in a broad variety of species. However, looking at a commercial use, the duration of the immune response against the vaccine is critical. Therefore the persistence of the DNA vaccine, as well as its expression, should be investigated. We conducted these investigations on a DNA vaccine against Chlamydophila psittaci, a Gram-negative intracellular bacterium which causes respiratory disease in turkeys and humans. Previous studies showed that the DNA vaccine confers partial protection against C. psittaci infection in turkeys. Turkeys were injected intramuscularly with the DNA vaccine : a eukaryotic expression vector (pcDNA1::MOMP) expressing the major outer membrane protein (MOMP) of an avian C. psittaci serovar D strain. Over a period of 11 weeks, cellular uptake of the DNA vaccine was examined by PCR, transcription of the insert by reverse transcript-PCR (RT-PCR) and mRNA translation by immunofluorescence staining of muscle biopsies.ResultsThe results indicate that the DNA vaccine persists in turkey muscle for at least 10 weeks. Moreover, during this period of time MOMP was continuously expressed, as evidenced by the immunofluorescence staining and RT-PCR.ConclusionSince C. psittaci infections occur at the age of 3 to 6 and 8 to 12 weeks, a vaccine persistence of 10 weeks seems adequate. Therefore, further research should concentrate on improving the elicited immune response, more specifically the cell-mediated immune response, rather than prolonging the lifespan of the plasmid.

Investigations on the clonality of isolates of Pasteurella gallinarum obtained from turkeys in Germany

Pasteurella gallinarum has been considered an opportunistic pathogen rather than a primary pathogen for chickens. As P. gallinarum has been found to have a high genotypic diversity, one would expect a polyclonal distribution among isolates from different farms if this organism is a secondary invader. The aims of this investigation were to genetically characterize isolates obtained from outbreaks affecting several turkey farms to confirm the existence of the infection in turkeys and to investigate the genetic relationship between isolates from affected farms. A total of 17 isolates from 14 outbreaks of respiratory disease in Germany were subjected to extended phenotypic and genotypic characterization. All isolates were of the same phenotype, typical of P. gallinarum. Ribotyping of three isolates using either HpaII or HindIII showed that they had identical profiles and indicated that the isolates all originated from the same clone. Comparison with HpaII ribotypes from a previous study showed that the pattern was identical to that obtained with isolates from a Zimbabwean outbreak in chickens during 1999 to 2000. Restriction endonuclease analysis typing of 14 isolates from all 14 farms showed that they had identical profiles but these differed from those obtained with isolates from the Zimbabwean outbreak. Sequencing of the 16S rRNA gene and sequence comparisons with other Pasteurellaceae confirmed their classification as P. gallinarum. Identification of the same clone of P. gallinarum from 14 outbreaks of acute respiratory disease in turkeys within a time period of 2 months suggests a common source of infection, and that P. gallinarum probably played a primary role rather than a secondary role in the outbreaks.

Pathogenesis of Avian Pneumovirus Infection in Turkeys

Avian pneumovirus (APV) is the cause of a respiratory disease of turkeys characterized by coughing, ocular and nasal discharge, and swelling of the infraorbital sinuses. Sixty turkey poults were reared in isolation conditions. At 3 weeks of age, serum samples were collected and determined to be free of antibodies against APV, avian influenza, hemorrhagic enteritis, Newcastle disease, Mycoplasma gallisepticum, Mycoplasma synoviae, Mycoplasma meleagridis, Ornithobacterium rhinotracheale, and Bordetella avium. When the poults were 4 weeks old, they were inoculated with cell culture-propagated APV (APV/Minnesota/turkey/2a/97) via the conjunctival spaces and nostrils. After inoculation, four poults were euthanatized every 2 days for 14 days, and blood, swabs, and tissues were collected. Clinical signs consisting of nasal discharge, swelling of the infraorbital sinuses, and frothy ocular discharge were evident by 2 days postinoculation (PI) and persisted until day 12 PI. Mild inflammation of the mucosa of the nasal turbinates and infraorbital sinuses was present between days 2 and 10 PI. Mild inflammatory changes were seen in tracheas of poults euthanatized between days 4 and 10 PI. Antibody to APV was detected by day 7 PI. The virus was detected in tissue preparations and swabs of nasal turbinates and infraorbital sinuses by reverse transcription polymerase chain reaction, virus isolation, and immunohistochemical staining methods between days 2 and 10 PI. Virus was detected in tracheal tissue and swabs between days 2 and 6 PI using the same methods. In this experiment, turkey poults inoculated with tissue culture-propagated APV developed clinical signs similar to those seen in field cases associated with infection with this virus.

Pathogenic and immunosuppressive effects of avian pneumovirus in turkeys.

Avian pneumovirus (APV) causes a respiratory disease in turkeys. The virus has been associated with morbidity and mortality due to secondary infections. Our objective was to determine if APV caused immunosuppression in the T-cell or B-cell compartments and to study the pathogenesis of the disease in APV maternal antibody-lacking 2-wk-old commercial turkeys. APV was administered by the eyedrop/intranasal route. Observations were made for gross lesions, viral genome, and T-cell mitogenesis and cytokine secretion at 3, 5, 7, 14, and 21 days postinoculation (DPI). During the acute phase of the disease that lasted for about 1 wk, the turkeys exposed to APV showed clinical signs characterized by nasal discharge and sinus swelling. Virus genome was detected by in situ hybridization in cells of turbinates and trachea at 3 and 5 DPI. At 3 and 5 DPI, spleen cells of the birds infected with APV markedly decreased proliferative response to concanavalin A (Con A). Con A and lipopolysaccharide stimulation of spleen cells from virus-exposed turkeys resulted in accumulation of nitric oxide-inducing factors (NOIF) in the culture fluid. NOIF were not detected in culture fluids of Con A-stimulated spleen cells of virus-free turkeys. APV did not compromise the antibody-producing ability of turkeys against several extraneous antigens such as Brucella abortus and tetanus toxoid.

Outer Membrane Proteins for Serologic Detection of Ornithobacterium rhinotracheale Infection in Turkeys

Ornithobacterium rhinotracheale (ORT) is a bacterium responsible for a respiratory disease in turkeys and chickens and has been identified as one of the emerging respiratory bacterial pathogens. The clinical signs and lesions caused by ORT are very similar to those caused by other respiratory infectious agents; therefore, an accurate diagnostic test is necessary to identify the infection. In this study, we have investigated the use of outer membrane proteins of ORT in an indirect enzyme-linked immunosorbent assay (ELISA) to detect the exposure to ORT infection. Outer membrane proteins of ORT were extracted and used as an antigen in ELISA to detect infection in turkeys exposed to different serotypes of ORT. The ELISA results were compared with the conventional serum plate agglutination test. The agglutination test detected specific antibodies for ORT in 65% of experimentally infected turkeys during the first 2 wk of infection. The ELISA detected up to 100% of infected birds for 8 wk postinfection. The results suggest that ELISA is able to detect the exposure to ORT in later stages of the infection and this assay can be used in serologic surveillance of ORT infection for poultry in the field.

Appearance of type B avian Pneumovirus in great Britain

Summary A survey in Britain in 1994/95 has shown that in all cases where avian pneumovirus (APV), was detected in turkeys of over 4 weeks of age, the virus was of type B. Oesophageal swabs were collected twice‐weekly for up to 14 weeks in seven longitudinal surveys. RNA was extracted and analysed by reverse transcription‐nested polymerase chain reaction designed to discriminate between type A and B APV. Virus of the same type as the vaccinal strain applied (type A or B) was detected from 7 and up to 35 days post‐vaccination. Subsequently, type B virus only was detected when the poults were 32 to 67 days old, although in any one growing unit the period of detection was maximally one week. Episodes of increased mortality were usually associated with the detection of this type B virus a few days previously. In a single experiment inoculation of poults with virulent type A or B APV induced cross‐protection although that protection was incomplete. Our results confirm that outbreaks of respiratory disease in turkeys in Britain during 1994 to 1995 were still associated with APV and that the virus was type B. In contrast, between 1985 and 1990 only APV antigenically similar to type A and B strains had been detected in Britain and in continental Europe respectively.

Turkey rhinotracheitis virus: in vivo and in vitro polypeptide synthesis.

Turkey rhinotracheitis (TRT) virus is a paramyxovirus associated with recent outbreaks of acute respiratory disease in turkeys. On morphological criteria it resembles the pneumoviruses more than the other members of the paramyxovirus family. In this communication we report the identification of five virus-induced polypeptides in TRT virus-infected BS-C-1 cells. The Mr of these polypeptides were estimated as 38K, 35K, 30K, 19K and 15K from their electrophoretic mobility in SDS-polyacrylamide 6 to 15% gradient gels. The virus specificity of four (the 38K, 35K, 30K and 19K Mr polypeptides) of these five was confirmed by their predominance as products of in vitro translation of mRNA from TRT virus-infected BS-C-1 cells. An additional virus-specific polypeptide with an estimated Mr of 22K was revealed by in vitro translation and may be either a non-structural polypeptide or a minor structural protein. All six polypeptides were immunoprecipitated by a murine antiserum. Three other polypeptides (129K, 57K and 45K) were also immunoprecipitated. The 57K, 45K and 15K Mr polypeptides were glycosylated, and the 57K glycopolypeptide may be a disulphide-bonded dimer of the 45K and 15K glycopolypeptides. Another major glycopolypeptide of 83K Mr was observed but its virus-specificity could not be confirmed. Taken together our results tentatively identify eight or nine TRT virus-specified polypeptides and are consistent with the classification of TRT virus as a new member of the genus Pneumovirus.

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.

Negative Findings concerning Alcaligenes faecalis as an Etiologic Agent in Acute Respiratory Disease of Turkeys

An acute respiratory disease of turkeys in Israel was first reported in November 1978. Alcaligenes faecalis was isolated from sick turkeys and from chickens not affected by the disease. Plate agglutination tests with A. faecalis antigen of 1,067 turkey and 494 chicken serum samples gave variable results: healthy turkeys gave positive reactions and sick turkeys sometimes gave negative ones. All isolated strains were highly sensitive in vitro drug sensitivity tests, but chemotherapy failed in the field. Pathogenicity trials with A. faecalis, given alone or in combination with Yucaipa virus to 8-day-old turkey poults, failed to reproduce the disease.

Transmission of Acute Respiratory Disease (Rhinotracheitis) of Turkeys

Clinical signs of acute respiratory disease of turkeys were transmitted to susceptible day-old poults by direct contact, litter contact, and drinking water. Attempts failed to transmit the disease by air (cage-to-cage) or by oral and nasal inoculation with feces, nasal exudates, or nasal turbinate extracts. The disease was not transmitted by inoculation with white blood cells. Chicks were not affected by the disease, but young quail developed signs of respiratory disease when exposed to contaminated drinking water. Thus, acute respiratory disease in turkeys appears to be of an infectious nature, and the infectious agent(s) probably exist in the heavily contaminated environment used to house young commercial turkey poults.

Isolation of an Etiologic Agent of Acute Respiratory Disease (Rhinotracheitis) of Turkey Poults

A small gram-negative motile bacillus was isolated from laboratory poults affected by acute respiratory disease (rhinotracheitis) of turkeys. The bacterium was inoculated intranasally into susceptible day-old poults; the poults developed typical clinical signs of acute respiratory disease, and the bacterium was reisolated. This same bacterium was isolated from commercial poults with typical signs of acute respiratory disease but not from poults of similar age which were clinically normal. The bacterium has not been identified taxonomically. We conclude that it is a primary etiologic agent for acute respiratory disease of turkey poults.

Early Lesions in Chicken and Turkey Embryos Inoculated with Mycoplasma gallisepticum Strain S6 at Various Stages of Incubation

Much of the variability reported in earlier studies of the lesions induced in chick embryos by mycoplasma is probably referrable to the use of poorly characterized organisms. Another important discrepancy results from the use of dead and chilled embryos, which would also tend to cloud the histopathologic results. This study sought only to define the early lesions of Mycoplasma gallisepticum (Strain S6) infections in chicken and turkey embryos at various stages of incubation. The only previous embryo work with this strain of Mycoplasma is that of Casorso (1), who reported the failure of this organism to produce lesions in 7-day-old chicken embryos. No histopathologic studies seem to have been made of this infection in turkey embryos. The S6 strain of M. gallisepticum is a neurotropic strain isolated from a turkey brain which readily produces a severe cerebral arteritis when inoculated intravenously in poults aged 2 to 7 weeks (4,5,6). By this route it is much less pathogenic to chicks. Thomas et al. (6) found only a single case of very mild arteritis among 8 inoculated 3-day-old hatched chicks. However, they demonstrated that this organism produced a toxin that was probably responsible for the vascular changes. This strain, in addition to its neurotropic tendencies, will incite severe respiratory disease in turkeys and chickens, depending on the route of inoculation (9).

Methaemoglobin formation by Mycoplasma gallisepticum: the role of hydrogen peroxide.

Mycoplasma gallisepticum, the agent responsible for common respiratory disease of turkeys, has been shown to agglutinate turkey erythrocytes1. During the course of investigations of a neurotropic strain of this organism (S6) which produces encephalitis associated with cerebral arteritis2, it was observed that incubation of a suspension of M. gallisepticum with washed turkey erythrocytes at 37° C caused the cells to become dark brown. Washed mycoplasmas enclosed in dialysis tubing also caused browning of erythrocyte suspensions and converted turkey and human haemoglobin to a brown solution which exhibited the spectral characteristics of methaemoglobin.