Virulence of recent isolates of Mycoplasma synoviae in turkeys

Systemic Mycoplasma synoviae (MS) infection was induced experimentally in commercial turkeys with recent MS isolates (K4822D and K4774J) from turkey breeder flocks that exhibited no clinical signs typical of MS infection except for a low incidence of swollen footpads. The virulence of each strain was compared by evaluating gross and microscopic lesions, serologic responses, and MS isolation rates at 10 and 21 days postchallenge and by comparing these results with those obtained from a known virulent isolate (K1968), another previously characterized field isolate (K4463B), and unchallenged controls. All strains induced lesions typical of infectious synovitis but showed distinct differences in the extent of the gross and microscopic lesions and in the isolation rates from the tissues in turkeys. K1968 induced the most extensive lesions in hock and stifle joints and footpads, but strains K4822D, K4774J, and K4463B all induced synovitis and were similar in virulence for synovial tissues. Very mild respiratory lesions were induced by all of the strains studied. All strains yielded strong positive serologic responses. We concluded that these recent field isolates, although able to induce synovitis, are less virulent for turkeys than a known virulent strain. Nevertheless, under severe experimental challenge, these strains have the capability of causing lesions that may be incompatible with economical turkey production.     

Experimental evidence of indirect transmission of Mycoplasma synoviae

The aim of the study was to analyse experimental transmission of Mycoplasma synoviae, an avian pathogen. Three experiments using specific pathogen-free day-old chicks placed in isolators were conducted. In the first experiment, the birds were introduced in an isolator previously contaminated with a M. synoviae broth culture. After 34 days, these birds were eliminated and, for the second trial, the chicks were introduced in the same isolator without disinfecting. In the third assay, the chicks were placed in an isolator containing a mixture of food, feathers and dust collected less than an hour earlier from a M. synoviae infected laying hen flock. In the second and third experiments in order to exacerbate the M. synoviae infection, the birds were inoculated with infectious bronchitis (IB) virus. The presence of M. synoviae in the environment and in tracheal swabs was monitored by culture, a multiplex PCR (mPCR) detecting M. synoviae and Mycoplasma 16S rDNA and a multiplex RT-PCR (mRT-PCR) detecting the M. synoviae mRNA coding for a membrane protein and Mycoplasma 16S rRNA. In in vitro experimental conditions, M. synoviae mRNA and 16S rRNA were detected up to 20 min and 23 h respectively after mycoplasma death. In the first assay, the first infected bird was detected on the 13th day. In the second trial, culturable M. synoviae or viable M. synoviae were detected in the isolator for 3 or 4 to 5 days respectively after depopulation of the birds of the first assay whereas the first culture positive tracheal swabs were detected on the 33rd day, after IB inoculation. In the third experiment, the first infected birds were detected on the 54th day. Thus, the different assays showed that M. synoviae contaminated material (dust, feathers and food) can infect chicks, sometimes after remarkably long silent periods.     

The isolation of Mycoplasms synoviae from chickens with infectious synovitis and air-sacculitis in the Republic of South Africa.

Mycoplasma synoviae was isolated from the trachea of chickens showing either typical infectious synovitis lesions or air-sacculitis. M. synoviae was identified by means of the direct plate fluorescent antibody technique and its growth-dependence on nicotine-amide-adenine dinucleotide. This is the first documented report on the isolation and identification of M. synoviae in the Republic of South Africa.     

Experimentally induced synovitis of chickens with Mycoplasma synoviae: effects of dexamethasone treatment.

Specific - pathogen - free chickens were inoculated in the right tibiometatarsal joint with a synovitis-derived Mycoplasma synoviae strain before or during dexamethasone treatment. Development of synovitis in chickens inoculated during the drug treatment was apparently delayed in comparison with development of synovitis in non-treated chickens. Severity of clinical synovitis in chickens inoculated before the drug was given was apparently less than that in chickens not treated or in chickens treated with dexamethasone. Histopathologic changes in the early stage of the infection (1 to 2 weeks) were not modified by dexamethasone treatment, although those changes in the succeeding stage (6 to 7 weeks) were greatly lessened. A relationship was observed between the dosage of dexamethasone and the severity of synovitis, as well as the kinds of cells that infiltrated into the joint lesions. Although serum antibody titers in chickens treated with an excessive dose of dexamethasone were markedly lower, clinical, bacteriologic, and histopathologic observations in chickens treated with dexamethasone were similar to those previously found in surgically thymectomized chickens. These results may support the theory that multiple synovitis of chickens caused by M synoviae infection develops mainly because of an immune response, especially by thymus-dependent functions.     

Prevalence of antibodies to Mycoplasma synoviae in laying hens and possible effects on egg shell quality

Mycoplasma synoviae (M. synoviae) can cause respiratory disease, synovitis, or result in a silent infection in chickens and turkeys. The importance of M. synoviae is well established in broilers but only a few studies have been conducted in layers. In the present study, the prevalence of M. synoviae in commercial layer flocks was estimated using ELISA. For this study, 19 commercial layer flocks were selected randomly from New South Wales and Queensland region of Australia from producers who were willing to participate in the survey. Sixty eggs per flocks were randomly collected, out of these 30 eggs were used for ELISA and remaining 30 eggs were used to estimate various egg shell quality parameters. Subsequently, association between the serological status of eggs for M. synoviae and egg shell quality was studied. In the flocks under study, seroprevalence of M. synoviae was found to be high at 69% (95% confidence interval (CI)=41.3-89.0). Statistical analysis showed an association between serological status for M. synoviae and egg quality parameters such as translucency, shell breaking strength, % shell reflectivity and shell deformation. On the other hand, there was no significant association between serological status for M. synoviae and other egg quality parameters such egg weight, egg shell weight, % egg shell or shell thickness.     

The humoral immune response of chickens to Mycoplasma gallisepticum and Mycoplasma synoviae studied by immunoblotting

The humoral immune response over time of White Leghorn chickens experimentally infected with Mycoplasma gallisepticum or M. synoviae by an aerosol inoculation or a contact exposure were compared by immunoblotting. The response of chickens infected with M. gallisepticum were similar with respect to proteins recognized and intensity of response, regardless of mode of infection. On the other hand, chickens infected by aerosolization of M. synoviae responded to more proteins and with greater intensity than did M. synoviae contact-exposed birds. Chickens infected with M. gallisepticum responded with antibodies to over 20 proteins, while chickens infected with M. synoviae responded with antibodies to 12 proteins. Field sera from chickens naturally infected on commercial poultry farms with M. gallisepticum or M. synoviae were analyzed by immunoblotting and were found to react with a number of mycoplasma proteins. However, no correlation was seen when comparing intensity of immunoblot staining and hemagglutination-inhibition titer of the field sera. The experimental antisera were used to identify species-specific proteins of M. gallisepticum and M. synoviae. Six immunogenic species-specific proteins of M. gallisepticum with relative molecular masses of 82 (p82), 65-63 (p64), 56 (p56), 35 (p35), 26 (p26), and 24 (p24) kilodaltons (kDa) were identified. Two species-specific proteins of M. synoviae with relative molecular masses of 53 (p53) and 22 (p22) kDa were identified. Additionally, a highly immunogenic 41 (p41) kDa protein of M. synoviae was identified. Species-specific proteins identified in these mycoplasmas and the 41 kDa protein of M. synoviae were purified by preparative SDS-PAGE in amounts sufficient for further characterization and for use in serodiagnostic tests.     

Prevalence of mycoplasma antibodies in lesser prairie-chicken sera

Serologic testing by the serum plate agglutination (SPA) procedure was performed to detect the presence of cross-reacting antibodies to Mycoplasma meleagridis, Mycoplasma synoviae, and Mycoplasma gallisepticum in lesser prairie-chickens (Tympanuchus pallidicinctus) trapped over a 2-yr period in Finney and Kearny counties of southwestern Kansas. Sera examined from birds (n = 50) obtained in March-April 2000 tested positive for M meleagridis, M. synoviae, and M. gallisepticum at levels of 6%, 10%, and 10%, respectively, for the population examined. Mycoplasma meleagridis antibodies were detected in 3 samples (2.7%), M. synoviae antibodies in 2 samples (1.7%), and M. gallisepticum antibodies in 3 samples (2.7%) from birds (n = 112) collected in March-April 2001. Data obtained by SPA can result in false positives and should be verified by additional procedures such as the hemagglutination-inhibition test. Low amounts of sera prohibited this additional testing. Thus, the positive SPA results should be considered presumptive for the presence of Mycoplasma antibodies. Although Mycoplasma antibodies have been detected in wild turkeys (Meleagris gallopavo) from Kingman and Butler counties in Kansas, this report is the first of possible mycoplasmosis in Finney and Kearny counties, Kansas. All birds testing positive by this procedure should be considered as potential carriers of Mycoplasma and should not be used in relocation efforts.     

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.     

Cases of swollen head syndrome in broiler chickens in Greece

From 50 commercial broiler flocks included in a study concerning respiratory disease, signs of swollen head syndrome (SHS) were shown in eight. Postmortem examination was performed in eight birds showing signs of SHS from each flock. The trachea and head from each bird were collected for laboratory investigation. An enzyme-linked immunosorbent assay (ELISA) was used for the detection of viral and avian mycoplasma antigens in the trachea, and bacteriologic examinations were performed from the infraorbital sinuses of the infected birds. According to the ELISA results, the most frequently detected antigen in the trachea was Mycoplasma synoviae (six flocks, 75%), followed by infectious bronchitis virus (IBV) (five flocks, 62.5%), avian adenovirus (four flocks, 50%), avian reovirus (three flocks, 37.5%), Mycoplasma gallisepticum (one flock, 12.5%), and Newcastle disease virus (NDV) (one flock, 12.5%). Turkey rhinotracheitis (TRT), infectious laryngotracheitis, and avian influenza viral antigens were not detected. Experimental assays for characterization of NDV and IBV isolates showed that they were strains of low virulence (evidently vaccine strains). Bacteriologic examinations from the infraorbital sinuses of the affected birds resulted in the isolation of Escherichia coli (seven cases, 87.5%) and Staphylococcus spp. (one case, 12.5%). It is evident that TRT virus did not play a causal role in SHS in commercial broiler flocks in Greece, but in this condition, other viruses (IBV, NDV), mycoplasmas, or bacteria may be involved, and environmental conditions seem to be essential to the occurrence and severity of the disease.     

Cell-mediated and humoral immune response of chickens to Mycoplasma synoviae.

The leukocyte migration-inhibition test was employed to demonstrate the presence of cell-mediated immunity and to ascertain its relation to immunoglobulin production in Mycoplasma synoviae infection in chickens. With peripheral leukocytes and a preparation of M. synoviae used as antigen, good discrimination was obtained between naturally or experimentally infected birds and uninfected control birds. Only the infected groups showed significant inhibition. Positive migration inhibition values developed in the second week of infection, often before the appearance of hemagglutination-inhibition titers, and continued to accompany the production of immunoglobulins with some degree of correlation for at least 6 or 12 months.     

Survey of pathogens and blood parasites in free-living passerines.

To determine the disease prevalence of free-living passerines, 1709 passerines were sampled from 38 different field sites in Ohio. Choanal and cloacal swabs were collected from each bird and cultured for the presence of Pasteurella multocida, Salmonella spp., and Escherichia coli by standard microbiologic techniques. In addition, the serum from each bird was analyzed for the presence of antibodies to Mycoplasma gallisepticum, Mycoplasma synoviae, Newcastle disease virus, and avian influenza virus. A blood smear was also made to examine for the presence of blood parasites. Results indicated that the isolation of E. coli varied with bird species, with the European starling having a higher (21.4%) isolation of E. coli. Salmonella spp. were also isolated from these free-living passerines. Pasteurella multocida was not isolated from any of the sampled passerines. These birds did not have antibodies to M. gallisepticum, M. synoviae, Newcastle disease virus, or avian influenza virus. Blood parasites were not detected in any of the birds sampled.     

Persistence of Mycoplasma synoviae in hens after two enrofloxacin treatments and detection of mutations in the parC gene

The ability of Mycoplasma synoviae, an avian pathogen, to persist despite fluoroquinolone treatments was investigated in hens. Groups of Mycoplasma-free hens were experimentally infected with the M. synoviae 317 strain and treated twice with enrofloxacin at the therapeutic dose. The results show that the two treatments did not have any influence on this strain of M. synoviae recovery from tracheal swabs. Mycoplasmas were isolated from tracheal swab cultures, but not from inner organs such as the liver or spleen, suggesting that this strain of M. synoviae was not able to cross the mucosal barrier to disseminate throughout the host. A significant increase of the resistance level to enrofloxacin of five re-isolated mycoplasma clones, was observed after the second treatment. This increase was associated in two clones to a Ser81-->Pro substitution, found in the ParC quinolone-resistance determining region (QRDR) of DNA topoisomerase IV. This is the first time that a mutation in a gene coding for topoisomerase IV is described in M. synoviae after in vivo enrofloxacin treatments in experimentally infected hens.     

Poor serologic response to upper respiratory infection with Mycoplasma synoviae in turkeys

Mycoplasma synoviae (MS) was isolated from a flock of commercial tom turkeys in which a small percentage of the birds exhibited clinical signs and lesions typical of MS synovitis. However, serologic testing of such flocks revealed poor to inconsistent reactivity by agglutination, enzyme-linked immunosorbent assay (ELISA) or hemagglutination inhibition; isolation of MS from such flocks proved to be very difficult. Turkeys were challenged with one of the isolates (K4463B) either by aerosol or systemically by a combination of intravenous, foot pad, and eyedrop routes. Turkeys challenged by the systemic route responded normally to all serologic tests, whereas those challenged by aerosol either responded very poorly on all serologic tests or were seronegative up to 6 wk postchallenge even though they were positive for MS by tracheal culture. These results suggest that turkeys may harbor an upper respiratory infection with MS while remaining serologically negative.     

Survey of the health and husbandry of small poultry flocks in Great Britain

An investigation into the health and husbandry of 15 small poultry flocks was undertaken. Each flock was visited in July and a questionnaire on management practices and disease history was completed. The flocks were clinically examined and serological tests for Salmonella pullorum, Mycoplasma gallisepticum, M synoviae, M meleagridis, Newcastle disease, infectious bursal disease, infectious bronchitis, eggdrop syndrome 76, adenoviruses and reoviruses were carried out. Oesophageal and cloacal swabs were cultured for mycoplasma and pullorum reactors were cultured. M gallisepticum, M synoviae, M meleagridis and M gallinarum infections were detected and serological reactions for all the viral diseases, except egg drop syndrome 76, were found. Evidence of Newcastle disease and pullorum disease was encountered. Lice were present in five flocks and mites in four flocks. Welfare standards varied.     

Intraspecific variation in 16S rRNA gene of Mycoplasma synoviae determined by DNA sequencing

Mycoplasma synoviae (MS) is an important avian pathogen may cause both respiratory disease and joint inflammation synovitis in poultry, causing economic losses to the Brazilian poultry industry. The genotypic variation in 16S rRNA gene is unknown. Partial sequences of 16S rRNA gene of 19 strains of M. synoviae were sequenced and analyzed in order to obtain molecular characterization and evaluation of the genetic variability of strains from distinct Brazilian areas of poultry production. Different polymorphic patterns were observed. The number of polymorphic alterations in the studied strains ranged from 0 to 6. The nucleotide variations, including deletion, insertion and substitutions, ranged from 3 to 5. The genotypic diversity observed in this study may be explained by spontaneous mutations that may occur when a lineage remains in the same flock for long periods. The culling and reposition in poultry flocks may be responsible for the entry of new strains in different areas.     

An outbreak of erysipelas in 2-day-old poults

Systemic erysipelas infection was seen in 2-to-4-day-old poults from three separate ranches owned by the same company. The affected poults were all from the same breeder source; poults from other breeder sources were seemingly unaffected. Mortality increased on days 2 and 3, ranging from 2% to 8.5%. Birds submitted were dehydrated and very weak, with one half of the poults submitted having died during transport to the lab. Gross lesions included swollen, congested livers and spleens, as well as hemorrhagic breast muscle in one case. Toes were swollen and reddish-purple in color. The poults had been toe-trimmed during hatchery processing using a commercial microwave. Histologically, periportal inflammation with heterophilic infiltration in the liver was noted. Spleens showed hyalinization of arteries, lymphoid depletion, and necrosis. Toe joints showed purulent synovitis and cellulitis. Gram stains done on impression smears of liver and spleen showed rare to moderate numbers of small gram-positive rods. Erysipelothrix rhusiopathiae was isolated from 18 of 22 livers cultured, five of six toe joints cultured, and from the yolk sac in two birds.     

鸡滑液囊支原体的分离鉴定及病理组织学观察

为了解宁夏及其周边不同地区蛋鸡场中的鸡滑液囊支原体的种类及其致病力,采用改良Frey氏鸡滑液囊支原体培养基,从疑似感染鸡滑液囊支原体的不同蛋鸡群中分离出病原株,设计鸡滑液囊支原体vlhA基因特异性引物,对分离到的病原进行基因序列扩增并测序,使用MEGA6.0软件中的邻接法(Neighbor-joining,NJ)构建分离株系统发育树并进行遗传进化分析,采用从临床症状最明显的鸡体分离的菌株进行动物感染试验,制作石蜡切片,HE染色,病理组织学观察。结果表明,所分离到的病原菌均为鸡滑液囊支原体,部分菌株之间极其相似;鸡滑液囊支原体不仅可以使鸡只关节肿胀、生长缓慢,也可引起鸡只的肝脏轻微肿大,肝细胞部分坏死、间质增生;脾脏结缔组织增生,出血。     

Detection of Mycoplasma synoviae in clinically normal pheasants

Mycoplasma synoviae was isolated from the tracheas of seven clinically normal pheasants found in the vicinity of a chicken farm infected with M synoviae, but not from 120 pheasants and partridges with respiratory disease. When specimens were examined by the polymerase chain reaction only two additional pheasants infected with M synoviae were identified, one healthy and one diseased.     

Laboratory infection of house sparrows (Passer domesticus) with Mycoplasma gallisepticum and Mycoplasma synoviae

House sparrows were infected by aerosol with Mycoplasma gallisepticum (MG) or M. synoviae (MS). MG was reisolated from 5 to 11 sparrows 10 days postinfection, but infection appeared to be temporary. Mycoplasma-free chickens reared in the experimental house became infected with MG during the trial. MS was recovered from only one sparrow. Serological tests were unsatisfactory for diagnosing infected birds. The results suggest that house sparrows may be temporary biological carriers of MG.     

Identification of species-specific and interspecies-specific polypeptides of Mycoplasma gallisepticum and Mycoplasma synoviae

Temporal antisera (TA) prepared in susceptible Leg-horn-type chickens against Mycoplasma gallisepticum and M synoviae were evaluated to determine the extent of cross-reactivity in ELISA and hemagglutination inhibition tests. Species-specific and interspecies-specific polypeptides were identified after electrophoretic separation and protein immunoblotting with reference antisera, TA, and a monoclonal antibody specific for M gallisepticum. Mycoplasma gallisepticum antiserum cross-reacted with M synoviae polypeptides in ELISA and TA immunoblots. Two major M synoviae polypeptides (88 and 53 kilodaltons [kD]) cross-reacted with M gallisepticum antisera in TA immunoblots. An M gallisepticum polypeptide of 70 kD cross-reacted with M synoviae in TA immunoblots. In contrast, M gallisepticum and M synoviae reference antisera cross-reacted when immunoblotted with heterologous antigens. A monoclonal antibody specific for M gallisepticum bound to a 69-kD polypeptide in lectin-purified and whole-cell M gallisepticum protein fractions in immunoblot assays. The lectin-purified fraction hemagglutinated chicken RBC. Seemingly, the 69-kD polypeptide may constitute all or part of the M gallisepticum hemagglutinin.