Antibody responses in sera and respiratory secretions from chickens infected with Mycoplasma gallisepticum

Antibodies in sera and respiratory secretions from chickens infected with Mycoplasma gallisepticum (MG) were measured by an enzyme-linked immunosorbent assay (ELISA). Chickens intratracheally inoculated with 10(5) cells of MG showed a correlation between severity of tracheal lesions and extent of MG colonization in the tracheas in the first 3 weeks postinoculation. Antibody titers in tracheal washings (TWs) of the infected chickens increased during this phase. Thereafter, isolation of MG from the trachea decreased sharply, and there was a concomitant decrease in tracheal lesion scores. At 5 weeks postinfection, the chickens that recovered from the infection exhibited a consistent presence of antibodies in TWs. Chickens reexposed had a faster rate of MG elimination and substantially less severe inflammatory lesions in the tracheas than chickens observed after the first exposure. These findings suggest a possible role of antibodies of the respiratory secretions in resistance to MG. The ELISA was a sensitive and reliable test to detect a minute amount of antibodies in the secretions.     

Tracheal populations of Mycoplasma gallisepticum after challenge of bacterin-vaccinated chickens

Chickens vaccinated once or twice with inactivated oil-emulsion vaccine against Mycoplasma gallisepticum (MG) or left unvaccinated were challenged intratracheally with the R strain of MG. The population of MG organisms was determined by enumerating tracheal cultures periodically up to 28 weeks postchallenge (PC). The number of organisms in the respiratory tract increased rapidly after 4 days PC, and the number tended to decrease after 4 weeks PC. Tracheal populations of MG varied considerably among individual chickens. Bacterin-vaccinated chickens had numerically lower populations of MG in their tracheas up to 8 weeks PC, but the differences were small and considered of little practical significance.     

Adenovirus infection associated with respiratory disease in commercial chickens

The lesions and etiologic agents associated with 13 outbreaks of respiratory disease in commercial chickens were investigated. Adenoviruses were isolated from tracheal and lung tissues of affected chickens in all 13 outbreaks. Escherichia coli was isolated from the lung of an occasional bird. The tracheal specimens were consistently negative for Bordetella avium, but E. coli and occasionally Staphylococcus aureus were isolated. There was also serological evidence in one outbreak, and pathological evidence in another, of a concurrent infectious bursal disease virus (IBDV) infection of chickens affected with the disease. Gross and microscopic alterations in the tracheas and lungs of affected chickens were similar in all outbreaks and consisted of catarrhal tracheitis and occasionally multifocal pneumonia with mononuclear cell infiltrates. Hepatitis and splenitis with heterophil infiltrates occasionally were seen in birds with coliform septicemia. The tracheal and lung lesions in the present investigation were considered primarily of adenovirus etiology, complicated by secondary bacterial infection.     

Comparative use of direct organ cultures of infected chicken tracheas in isolating avian infectious bronchitis virus

Five trials were conducted to compare four in vitro methods of isolating avian infectious bronchitis virus (IBV)-direct organ culture of infected tracheal rings (DOC), inoculation of tracheal organ culture (OC), inoculation of chicken embryo, and inoculation of cultured cells. DOC was prepared from tracheas of chickens experimentally inoculated with field samples. In the other methods, pooled tracheal and kidney suspensions were used to inoculate OC, chicken embryos, and cultured cells. IBV was consistently isolated at the initial passage by the DOC and OC inoculation systems, but it was not always isolated by embryo inoculation and never isolated by cultured-cell inoculation. When combined with immunofluorescent staining, DOC was much more efficient than the OC inoculation system for isolation and identification of the five strains of IBV tested because of its simplicity and speed.     

Safety and efficacy of the avirulent Mycoplasma gallisepticum strain K5054 as a live vaccine in poultry

A Mycoplasma gallisepticum (MG) isolate from an atypically mild outbreak in turkey breeders was found to be similar to house finch isolates by DNA analyses. A preliminary study in turkeys showed that this isolate (K5054) caused very mild lesions and protected turkeys against subsequent challenge with a virulent MG strain. In this study, K5054 was further evaluated as a potential vaccine strain in commercial layer-type chickens and turkeys. The safety of K5054 was evaluated by aerosol challenge followed by evaluation of gross and histopathologic lesions as well as serologic reactions and isolation of MG from the trachea and air sacs. Infection of chickens (trial 1) and turkeys (trial 2) with K5054 resulted in little evidence of MG lesions. There was weak seroconversion, and K5054 was consistently reisolated from the tracheas of chickens and turkeys. The efficacy of K5054 as a vaccine was evaluated by aerosol challenge of vaccinated chickens (trial 3) and turkeys (trial 4) with virulent R strain. There was evidence of protection from lesions associated with MG.     

Comparison of a modified Edward-type medium and a modified SP4-type medium for primary isolation of Mycoplasma gallisepticum (MG) from chickens vaccinated with the F strain of MG

The efficacy of two media, an Edward-type medium (EPJ) and a modified SP4-type medium (SP4-PS), were compared for primary isolation of Mycoplasma gallisepticum (MG) from commercial layer chickens (n = 58) vaccinated with the live F strain of MG. Three groups of chickens that differed in the interval after vaccinal exposure to the F strain (32, 41, and 102 weeks) were studied at necropsy. Mycoplasma isolation was attempted from the trachea, sinus, and cloaca using lavage and swab techniques but was successful only from the trachea and sinus. MG was isolated from 39 (8.4%) of 463 culture attempts from 58 tracheal inocula and 58 sinus inocula. Isolation of MG was successful more frequently using EPJ medium than SP4-PS medium, and isolation occurred more often from the sinus than from the trachea. Of the 58 chickens studied, 19 (33%) were shown by culture to be infected with MG. Isolation was successful only from 32- and 41-week post-vaccination exposure groups. However, all chickens studied were serologically positive for MG antibody by rapid-plate agglutination and hemagglutination-inhibition assays.     

Escherichia coli colonization of the trachea in poultry: comparison of virulent and avirulent strains in gnotoxenic chickens

In chickens, virulent Escherichia coli strains express their pathogenicity in the respiratory tract. A quantitative comparison of tracheal colonization by virulent and avirulent E. coli was carried out in gnotoxenic chickens after intestinal implantation. Two-week-old axenic chicks reared in isolators were inoculated per os with various associations of identified E. coli strains. No clinical sign of disease was observed in any of the chicks, despite the presence of virulent strains in all the intestines and most of the tracheas. The virulent organism reached greater population sizes in the trachea and feces of monocontaminated chicks and of chicks contaminated simultaneously with a virulent and an avirulent strain. In holoxenic chicks, identified virulent and avirulent strains were outnumbered by the E. coli population of the intestinal flora previously established and could not be recovered from the tracheas of most chicks.     

An Alcaligenes faecalis isolate from turkeys: pathogenicity in selected avian and mammalian species

An Alcaligenes faecalis isolate of known pathogenicity for turkeys was examined for adherence and cytotoxicity in tracheal organ cultures of turkeys, chickens, Japanese quail, guinea pigs, hamsters, and mice, and for colonization and pathogenicity in these 6 species. Adherence and colonization were detected by fluorescent antibody staining. Infected and noninfected tracheal rings were examined by phase-contrast microscopy for cytotoxicity (ciliostasis, blebing of the cell membrane, and sloughing of the ciliated epithelium). Alcaligenes faecalis adhered to the tracheal rings of all species examined. Cytotoxicity was apparent in the tracheal rings of turkeys, quail, and chickens. Cytotoxicity was not detected in tracheal rings from the mammalian species. Alcaligenes faecalis colonization of turbinates and tracheas of intact turkeys and quail was detected. Clinical signs of alcaligenes rhinotracheitis were observed and histopathologic characteristics of the disease were detected. Chickens, guinea pigs, hamsters, and mice were refractory to infection with this isolate of A faecalis.     

Escherichia coli multiplication and lesions in the respiratory tract of chickens inoculated with infectious bronchitis virus and/or E. coli.

Escherichia coli numbers and histopathological changes were studied in the respiratory tract of line 151 chickens intranasally inoculated with infectious bronchitis virus (IBV) and/or virulent E. coli; this line is highly susceptible to IBV. Chickens inoculated with IBV alone showed increased numbers of E. coli in the trachea and had tracheitis, airsacculitis, and bronchiolitis. One of 17 chickens inoculated with IBV alone died with fibrinopurulent serositis. Chickens inoculated with IBV and E. coli had more severe and persistent respiratory lesions than those inoculated with IBV alone. E. coli was isolated from tracheas of chickens inoculated with IBV and E. coli more frequently than from chickens inoculated with IBV alone. In this group, 14 of 27 chickens died with tracheal plugs or with fibrinopurulent serositis. There was neither increased numbers of E. coli nor significant lesions in the respiratory tract of the group inoculated with E. coli alone. These results suggest that IBV may facilitate E. coli invasion into the lower respiratory tract of the chicken.     

Detection of infectious bronchitis virus antigen from experimentally infected chickens by indirect immunofluorescent assay with monoclonal antibody

Infectious bronchitis virus (IBV) was detected by indirect immunofluorescent assay with a monoclonal antibody (MAb-IFA). The monoclonal antibody was specific for the nucleocapsid protein of IBV strain M41. The MAb-IFA clearly detected IBV with high specificity in infected chicken kidney cells. The assay furthermore detected IBV in tracheal smears and sliced tracheas from experimentally infected chickens. The positive reaction was found to be longer than that in the virus recovery test. These results indicate that MAb-IFA is a useful method for the detection of IBV from chickens suspected to have infectious bronchitis.     

Evaluation of protection against colonization of the chicken trachea following administration of Mycoplasma gallisepticum bacterin

Twelve-week-old commercial white leghorn pullets were given one or two doses of an inactivated oil-emulsion Mycoplasma gallisepticum (MG) vaccine or kept as unvaccinated controls. At 24 weeks of age, all groups were challenged intratracheally with one of six dilutions of a low-passage R strain of MG. Three days postchallenge, the tracheas from all chickens were cultured for MG to determine the number of challenge organisms required to initiate infection. The log10 ID50 of chickens vaccinated 0, one, or two times was 2.9, 3.4, and 3.7, respectively, and the minimum infectious dose (the lowest challenge dose to infect a single bird) was 15, 150, and 1500 colony-forming units, respectively. It was concluded that the vaccine provided measurable, though limited, protection against infection under these experimental conditions.     

Effect of temperature-sensitive Mycoplasma gallisepticum vaccine preparations and routes of inoculation on resistance of white leghorns to challenge

One-week-old chickens were vaccinated with live or formalin-killed temperature-sensitive (TS) Mycoplasma gallisepticum (MG) either intranasally (IN) or subcutaneously (SQ). Live TS MG protected chickens against S6 strain challenge directly into the air sacs, regardless of route of vaccination. Killed MG, however, protected chickens only when administered SQ. Antibody to MG was detected in sera and in the tracheal and air-sac washings of only the chickens given live vaccine IN. The antibody present in tracheal and air-sac washings may be one of the mechanisms that play a role in resistance to MG challenge.     

A quantitative study of single and mixed infection of the chicken trachea by Mycoplasma gallisepticum

The interaction between Mycoplasma gallisepticum (MG) and the tracheal mucosa of the young chicken was studied. The use of a selective plating method permitted differentiation between a pathogenic tylosin-resistant strain (227) and a less pathogenic tylosin-sensitive vaccine strain (F). Both MG strains adhered to the tracheal mucosa and colonized equally well. In mixed infection, the presence or absence of the second strain did not change the efficiency of colonization by either strain. When chickens were exposed to the vaccine strain 24 hr or 2 weeks before superinfection by the pathogen, there was no significant reduction in the efficiency of superinfection, despite the presence of 10(6) colony-forming units of MG strain F in the trachea. However, chickens had an increased ability to resist superinfection 5 weeks after exposure via the air sac. These results suggest that the biological mechanism underlying protection of F-strain-vaccinated chickens against adventitious infection by the homologous species does not involve competition for adherence sites or blockage by prior colonization.     

Evaluation of Mycoplasma gallisepticum K-strain as a live vaccine in chickens

We evaluated the pathogenicity of three live Mycoplasma gallisepticum (MG) vaccine candidates by infection via aerosol of 3-wk-old chickens with log phase broth cultures (trial 1). Two of the candidates (K3020 and K4649A) colonized only 10% and 20% of the chickens, respectively, unlike K2101 (K-strain), which was reisolated from all of the vaccinated chickens tested. K-strain inoculation did not result in significant air sac or tracheal lesions in chickens at 10 and 39 days postinfection (P < or = 0.05). The efficacy of K-strain as a live vaccine was evaluated in trial 2, by challenge of vaccinated chickens with virulent R-strain via aerosol at 6 wk postvaccination. K-strain vaccination resulted in significant protection from air sac and tracheal lesions (P < or = 0.05). The K-strain was further investigated to evaluate transmissibility (trial 3), colonization and persistence of infection following aerosol administration (trial 4), genetic and phenotypic stability following back passage through chickens (trial 5), and vertical transmission (trial 6). The K-strain had a low rate of horizontal transmission; it remained primarily in the respiratory system of inoculated birds and persisted in the upper respiratory tract for the duration of the trial 4 (5 mo). There was no increase in virulence of K-strain when it was back passaged five times through chickens, and no vertical transmission of K-strain was detected. K-strain showed great potential as a safe and effective live MG vaccine.     

Histochemical study of normal and collapsed tracheas in dogs

Tracheas from 15 toy breed dogs with normal tracheas and 6 dogs with collapsed tracheas were examined histologically and histochemically. Tracheal cartilage was analyzed for chondroitin sulfate by means of alcian blue (CEC method) and for calcium with alizarin red S. Cartilage arcs from dogs with collapsed tracheas had areas that were apparently hypocellular, and some had other areas that appeared like fibrocartilage or fibrous tissue. Histochemically, collapsed tracheal cartilage had less chondroitin sulfate and calcium than did normal tracheal cartilage. Cartilage arcs from the collapsed tracheas were not uniformly affected to the same degree, and parts of a given tracheal arc appeared normal, whereas other parts had an abnormal histologic appearance.     

Immunity induced with an aluminum hydroxide-adsorbed Mycoplasma gallisepticum bacterin in chickens

The protective effect of an inactivated Mycoplasma gallisepticum (MG) bacterin was evaluated in chickens subsequently challenged intratracheally (IT) with the homologous strain. Antibody responses in sera and tracheal washings (TWs) from these chickens were determined by an enzyme-linked immunosorbent assay. A group of chickens was vaccinated intramuscularly (IM) with two doses of the bacterin containing aluminum hydroxide gel (IM + IM). Another group was vaccinated IM with the same bacterin followed by IT with bacterin lacking the adjuvant (IM + IT). Chickens of both vaccinated groups had similar levels of antibody in TWs at the time of challenge. MG was eliminated from the trachea at higher rates and inflammatory lesions in the trachea were less severe in vaccinated chickens than in unvaccinated controls. The protective effect in chickens vaccinated IM + IT was greater than that in chickens vaccinated IM + IM. Perhaps vaccinal immunity is mediated by local rather than systemic antibody responses, or perhaps resistance provided by vaccination IM + IT is conferred partly by another immune mechanism such as cell-mediated immunity.     

Influence of swab material on the detection of Mycoplasma gallisepticum and Mycoplasma synoviae by real-time PCR

Recent reports have shown an increased recovery of cells from flocked nylon swabs which may improve the specimen quality and the real sensitivity of diagnostic tests in a clinical setting. In this study, the detection of Mycoplasma gallisepticum (MG) and M. synoviae (MS), using dry swabs of different materials (nylon flocked, cotton, and polyester), was investigated using real-time TaqMan PCR protocols. Different types of samples, including dilutions of pure broth cultures of MG and MS as well as swabs from tracheas of experimentally infected chickens and field cases of infection, were analyzed. There were no statistical differences in real-time PCR results among the different swab types (P < 0.05), indicating that this is not likely to be a significant factor in MG and MS detection by this method.     

Response of chickens to inoculation with a temperature-sensitive mutant of Mycoplasma gallisepticum

Newly hatched chickens were inoculated intranasally with either the S6 or TS 100 strain of Mycoplasma gallisepticum (MG) or they were left uninoculated. The three groups of chickens did not differ discernibly in body, spleen, or bursa weight during the 27-day sampling period. However, the S6-inoculated chickens showed a more pronounced cellular response in the nasal passages and had nearly complete lymphoid depletion in the spleens. The TS 100-inoculated birds expressed only a mild cellular reaction, which was localized in the nasal passages. Uninoculated chickens appeared normal histologically. Serologic tests such as rapid serum plate agglutination, hemagglutination-inhibition, and radioimmunoassay were able to detect antibody responses of chickens to MG inoculations yet could not differentiate the response to TS 100 from the response to S6. Tracheal secretions in intact TS 100-inoculated chickens contained antibodies to MG, yet only one-half of the bursectomized inoculated chickens contained detectable antibody, which appeared to be IgG. This led to the conclusion that bursectomy suppresses the appearance of locally synthesized IgG antibodies to MG in tracheal washings. The locally produced antibody was considered important in the development of resistance induced by intranasal inoculation of TS mutants.     

一株类4/91病毒的初步研究

从发病鸡群中分离到一株病毒，通过鸡胚接种、电镜观察、动物回归试验、气管环交叉中和试验等，初步确定为传染性支气管炎病毒，暂命名为A2。外源病原检查结果为阴性。攻毒试验：在气管、肾以及肌肉等组织均有明显的病变；病理组织观察；在气管、肾脏和肌肉均发生了一定程度的病理变化。利用电泳对传统的病毒株和分离株进行了病毒结构蛋白分析，A2株出现了31kD的特殊蛋白条带。气管环交叉中和试验表明A2株与4／91病毒为同一血清型；临床的动物免疫试验结果表明，4／91疫苗有较好的保护作用。