Delineation of the lateral spread of Mycoplasma gallisepticum infection in chickens

Lateral spread of S6 strain Mycoplasma gallisepticum (MG) was studied in small populations of chickens. One experimentally exposed bird served as the source of infection, and the presence of MG-agglutinating antibody was evidence of infection in individuals. The results were subjected to survival data analysis. In the seven experiments, four similar but not identical phases of lateral spread were observed: phase 1, a generally long latent phase (median 15, range 12-21 days) before antibody was first detected in the MG-inoculated bird; phase 2, a generally short period (median 1, range 1-21 days) in which infection gradually appeared in 5-10% of the population; phase 3, a fairly constant characteristic phase (median 24, range 7-32 days) in which 90-95% of the remaining population developed MG antibody; phase 4, a generally short terminal phase (median 4, range 3-19 days) in which the remainder of the population became positive. Increasing the population density increased the rate at which lateral spread occurred.     

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.     

Monitoring Mycoplasma gallisepticum and Mycoplasma synoviae infection in breeder chickens after treatment with enrofloxacin.

Three experimental strains of breeder chickens were accidentally exposed to Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS), presumably from a newly introduced group of leghorn-type pullets. The experimental strains subsequently became infected and were diagnosed positive for MG and MS by the serum plate agglutination (SPA) test and confirmed by the hemagglutination inhibition (HI) test and the polymerase chain reaction (PCR) of tracheal swabs. Treatment with 10 mg/kg enrofloxacin via drinking water for 14 days was elected. Before and after initiation of treatment, MG and MS were monitored for changes by SPA, HI, PCR, and culture, with sampling intervals ranging from 1 wk to 7 wk. MG and MS SPA, HI, PCR, and culture were performed at each sampling period, with the exception of weeks 1.0 and 6.5. Week 1.0 included SPA and His for MG and MS. Week 6.5 included PCR and culture for MG and MS. The MG and MS SPA results were positive throughout the 29-wk trial period. MG HI titers declined until the last sampling, whereas the MS HI titers did not decline significantly. PCR for MG yielded only one positive result, which occurred before treatment. MS PCR remained positive throughout the trial period. MG was never isolated from any sample; however, one MS organism was isolated during treatment. The treatment regimen was effective for MG on the basis of PCR results. Treatment with enrofloxacin did not eliminate SPA reactions during the 29-wk trial period. MG HI titers remained in the suspicious range throughout the remainder of the trial period. Four weeks after the treatment ended, MG HIs were reduced by approximately 40%, with MS HIs remaining high throughout the 29-wk period. PCR appeared to be a sensitive and specific test on the basis of correlation with HIs. On the basis of the isolation of MS during treatment and continued subsequent PCR positive reactions, the treatment for MS with enrofloxacin was not as efficacious as for MG.     

Evaluation of protection against Mycoplasma gallisepticum infection in chickens vaccinated with the F strain of M. gallisepticum

The effect of vaccination with the F strain of Mycoplasma gallisepticum (MG) on protection against challenge with a tylosin-resistant strain of MG was evaluated. White leghorn chickens vaccinated via eyedrop at 6 weeks of age were subsequently challenged with various dilutions of the tylosin-resistant MG strain, as were unvaccinated controls. Three days later, tracheal swabs were collected and cultured in medium with and without tylosin to distinguish between the vaccine and challenge strains. The mean infectious dose of the challenge strains was 3.8 log10 higher in the vaccinated group than in the controls, and the vaccinated group harbored fewer challenge organisms in the trachea. These findings suggest that the F strain of MG induces protection against infection with field strains of MG and that long-term vaccination with the F strain in multiple-age layer farms may result in replacement of field MG strains by the F strain.     

罗红霉素对鸡霉形体病的药效观察

罗红霉素 (Roxithromycin)为十四元环的半合成新大环内酯类抗生素 ,抗菌谱较广 ,对多数革兰氏阳性菌 (如金葡菌、链球菌等 )、部分革兰氏阴性菌敏感 ,对某些厌氧菌及支原体、衣原体亦有较强的抗微生物作用[1] 。罗红霉素与红霉素有相似的抗菌谱和不完全的交叉耐药性 ,其优点是提高了对酸的稳定性 ,同时显著改善了药动学性质 ,如半衰期延长 ,能迅速经胃肠道吸收 ,具有较高的血药浓度 ,能广泛分布于全身各组织及体液中 ,其血药峰浓度及生物利用度是大环内酯类中相对最高者 [2～ 4 ] 。医学上主要用于呼吸道、泌尿道及皮肤软组织感染 [2 ]。国…     

Effect of medicated feed on tracheal infection and population of Mycoplasma gallisepticum in chickens

Six-week-old broilers were fed 50 g tylosin/ton, 400 g chlortetracycline (CTC)/ton, or unmedicated feed and then challenged intratracheally with R strain Mycoplasma gallisepticum (MG). Feed-grade antibiotic medication did not prevent infection, but medication did lower the number of isolations from treated birds compared with controls. Only tylosin significantly lowered MG counts in the trachea. The log10 ID50 of birds receiving tylosin, CTC, or unmedicated feed were 5.8, 4.4, and 2.9, respectively. Six-week-old leghorns were placed on the three diets described previously and challenged with the R strain of MG. Birds were sacrificed at various times up to 10 weeks postchallenge (PC). Compared with the control diet, the tylosin-medicated diet significantly reduced the tracheal MG count from day 1 to 4 weeks PC, whereas the CTC-medicated diet significantly lowered the tracheal MG count only at 3 days PC. In all groups, the MG count gradually declined after 1 week PC; by 8 weeks PC it was essentially the same in all groups. It was concluded that continuous medication has the potential for reducing MG tracheal populations only during the initial phase of an outbreak.     

Cloning and expression in Escherichia coli of Mycoplasma gallisepticum antigens recognized by sera from infected chickens

A clone bank of Mycoplasma gallisepticum (MG) strain A5969 DNA was prepared in the expression vector phage lambda gt11. Approximately 75% of the resulting phages were recombinants, based upon the insertional inactivation of the lacZ gene of the vector. Clones were screened immunologically with serum prepared from specific-pathogen-free white leghorn chickens that had been infected with aerosolized MG. Approximately 250 clones, or less than 1% of the recombinant phage, reacted positively to various degrees with the test serum and failed to react with serum from uninfected specific-pathogen-free control chickens. A single clone was chosen at random for comparison with a vector control by western immunoblot, revealing a polypeptide of 140,000 molecular weight in the clone profile but not the control profile that reacted with immune serum. Clones expressing MG antigens recognized during infection may provide an improved means for antigen preparation for serologic diagnosis of mycoplasmosis.     

藏鸡鸡毒霉形体的血清学调查

霉形体又称支原体 ,是一类缺乏细胞壁的原核微生物 ,分类学上属于软皮纲成员。在兽医学上是一类重要的病原微生物 ,对禽类等动物具有广泛的致病性。 2 0 0 2年 4月 ,西藏某些地区鸡发生一种以咳嗽 ,鼻炎 ,打喷嚏 ,呼吸有锣音等呼吸症状为主要特征的疾病。另外 ,还可见到鼻分泌物较多 ,部分鸡出现结膜炎 ,眼中伴有泡沫分泌物 ,产蛋量下降并有死亡现象。经血清学快速诊断为鸡毒霉形体。为了摸清鸡毒霉形体在藏鸡中的感染情况 ,我们应用血清学方法对西藏各地区的藏鸡进行霉形体检测。结果显示我区藏鸡的鸡毒霉形体感染率较高 ,对藏鸡的年龄、性…     

Evaluation and comparison of various PCR methods for detection of Mycoplasma gallisepticum infection in chickens

Four genetic Mycoplasma gallisepticum (MG) polymerase chain reactions (PCRs) (16s rRNA PCR, three newly developed PCR methods that target surface protein genes [mgc2, LP (nested) and gapA (nested)]) were compared for analytical specificity and sensitivity and for diagnostic sensitivity (Se) and specificity of detection from tracheal swabs. The licensed MG DNA Test Kit Flock Chek test (IDEXX, Laboratories, Inc., Westbrook, ME) was as well evaluated for the diagnostic specificity and sensitivity of detection from tracheal swabs. Analytical specificity was evaluated for the four generic PCR methods using a panel of DNA samples from microorganisms that may be isolated from the trachea of commercial poultry and other fowl. PCR methods mgc2, nLP, and ngapA only amplified DNA from MG, whereas 16S rRNA PCR amplified DNA from MG and Mycoplasma imitans. The analytical sensitivity of the four generic PCR methods expressed in color-changing units (CCU)/amplification reaction was estimated for each PCR method and ranged from 4 to 400 CCU/reaction; the sensitivities of single PCR methods 16S rRNA and mgc2 were estimated at 40 CCU/reaction, the nLP at 400 CCU/reaction, and the ngapA at 4 CCU/reaction. The diagnostic sensitivity and specificity of MG detection from tracheal swab pools, as compared to isolation from choanal cleft swabs, was evaluated for the five PCR methods using three groups of birds exposed to vaccine strains ts-11 and 6/85 and to challenge strain R. All PCR methods were able to detect the vaccine strains and the challenge strain R directly from tracheal swabs, indicating that PCR primers from the different methods amplified divergent MG strains. Isolation and PCR results correlated satisfactorily among the three experimentally infected groups, with agreement values (k) ranging from 0.52 to 1.00. The ngapA, IDEXX, and mgc2 PCRs showed the best sensitivity (Se) ratios for detection of M. gallisepticum strains as compared to isolation. Compared to the ngapA and IDEXX PCR methods, the mgc2 PCR has a faster turnaround time, since this test consists of a single amplification reaction and the amplification product is detected by gel electrophoresis. Therefore, among the PCR methods evaluated in this study, the mgc2 PCR is the method of choice to further validate in the field.     

Mycoplasma gallisepticum and Escherichia coli mixed infection model in broiler chickens for studying valnemulin pharmacokinetics

A Mycoplasma gallisepticum–Escherichia coli mixed infection model was developed in broiler chickens, which was applied to pharmacokinetics of valnemulin in the present experiment. The velogenic M. gallisepticum standard strain S6 was rejuvenated to establish the animal model, and the wild E. coli strain O78 was injected as supplementary inoculum to induce chronic respiratory disease in chickens. The disease model was evaluated based on its clinical signs, histopathological examination, bacteriological assay, and serum plate agglutination test. The pharmacokinetics of valnemulin in infected chickens was determined by intramuscular (i.m.) injection and oral administration (per os, p.o.) of a single dose of 10 mg/kg body weight (BW). Plasma samples were analyzed by liquid chromatography–tandem mass spectrometry. The plasma concentration–time curve of valnemulin was analyzed using the noncompartmental method. After the i.m. administration, the mean values of C_max, T_max, AUC_last, MRT, CL_β/F, V_z/F, and t_1⁄2β, were 27.94 μg/mL, 1.57 h, 171.63 μg·h/mL, 4.51 h, 0.06 L/h/kg, 0.56 L/kg, and 6.50 h, respectively. By contrast, the corresponding values after p.o. administration were 5.93 μg/mL, 7.14 h, 47.60 μg·h/mL, 9.80 h, 0.22 L/h/kg, 3.35 L/kg, and 10.60 h. The disposition of valnemulin was retarded in infected chickens after both modes of extravascular administration as compared to the healthy controls. More attention should be given to monitoring the therapeutic efficacy and adverse effects of mixed infection because of higher required plasma drug concentration and enlarged AUC with valnemulin treatment.     

Immunization of chickens with temperature-sensitive mutants of Mycoplasma gallisepticum

Temperature-sensitive (TS) mutants of the S6 strain of Mycoplasma gallisepticum (MG) were used to immunize newly hatched chickens. Immunized chickens developed antibodies to the wild-type (WT) S6 strain as demonstrated by serologic tests. MG was recovered from nasal cavities but not from the lower respiratory tract of the immunized chicks. Three weeks after intranasal immunization, chickens were challenged via the air sacs with the virulent S6 strain. Immunized chickens were significantly better protected from development of air-sac lesions than were controls.     

Immunohistochemical detection of Mycoplasma gallisepticum antigens in turkey respiratory tissues

An avidin-biotin-immunoperoxidase diagnostic test was developed to facilitate rapid identification of Mycoplasma gallisepticum in respiratory tissues of turkeys. This procedure used polyclonal primary antibodies produced in rabbits. Turkeys were inoculated into the infraorbital sinus and trachea with the R strain of M. gallisepticum, Mycoplasma synoviae, Mycoplasma meleagridis, or Frey's media. The outer walls of the infraorbital sinuses, lungs, and tracheas were collected and fixed in either 10% neutral formalin or pentanedial methyl glycol at 1, 2, 3, and 4 wk postinoculation. Tissues were subdivided and remained in each fixative for 6 or 24 hr. The avidin-biotin-immunoperoxidase diagnostic test was sufficiently sensitive to detect M. gallisepticum antigen at 1, 2, 3, and 4 wk postinoculation. Staining of M. gallisepticum was significantly more intense on infraorbital sinus epithelium than on respiratory epithelium from the trachea or lung. Statistical analysis indicated that the 6-hr fixation time offered better antigen preservation than 24 hr in a fixative. There was no difference in intensity of M. gallisepticum antigen staining in tissues fixed in methyl pentanedial glycol when compared with tissues fixed in 10% neutral buffered formalin. Significant differences in staining intensity were observed between weeks. Specificity of the avidin-biotin-immunoperoxidase test was not complete. None of the tissues from the M. meleagridis and control groups showed staining. No staining was observed in the ciliated brush border of infraorbital sinus epithelial cells from turkeys infected with M. synoviae. However, weak to moderate staining was observed in several tracheas of turkeys inoculated with M. synoviae. Improved specificity of an avidin-biotin-immunoperoxidase diagnostic test to detect M. gallisepticum in respiratory tissues of turkeys probably will require the use of multiple monoclonal antibodies directed against several different epitopes specific to the cell membrane of M. gallisepticum.     

Disposition kinetics of doxycycline in chickens naturally infected with Mycoplasma gallisepticum

1. The pharmacokinetic properties of doxycycline were determined in healthy chickens and chickens naturally infected with Mycoplasma gallisepticum after a single intravenous (i.v.) and oral administration of the drug at 20 mg/kg body weight. Tissue residues of the tested drug after an oral dose of 20 mg/kg given twice daily for 5 consecutive days were also estimated in diseased chickens. 2. The plasma concentrations of doxycycline following single i.v. and oral administration were higher in healthy chickens than in diseased ones. Following i.v. injection, the elimination half-life (t1/2beta), distribution half-life and mean residence time (MRT) were longer in healthy chickens than in diseased birds. The values of total body clearance (ClB) and volume of distribution (Vdss) were larger in healthy chickens than in diseased birds. 3. After single oral administration, the absorption half-life (tl/2ab) and the elimination half-life were longer in normal birds than in diseased ones. The maximum plasma concentration of the drug was higher in normal chickens than in diseased ones. 4. Following repeated oral administration, the concentration of doxycycline in all tissues except muscle was higher than the corresponding concentrations in plasma. Concentrations of doxycycline in different tissues were in the following order: kidney > liver > lung > muscle. The drug was detected in liver and kidney in substantial concentrations on d 5 post administration of the last dose whereas, on d 7, its concentration in all tissues was below the lower limit of the sensitivity of the assay method used. Because of the low sensitivity of the microbiological assay method used in this study, a safe withdrawal time for doxycycline in diseased birds could not be estimated for the meanwhile.     

DNA probes for Mycoplasma gallisepticum and Mycoplasma synoviae: application in experimentally infected chickens

DNA probes specific for Mycoplasma gallisepticum and M. synoviae were selected from genomic libraries prepared in the pUC13 vector. The probes hybridized with the DNA of a wide spectrum of strains within each homologous species, but did not react with the heterologous species or with DNA from any other avian mycoplasma or bacteria tested. Experimental infection and contact exposure of chickens to M. gallisepticum served as models to test the effectiveness of the DNA probe in diagnosis as compared with serological and culture detection methods carried out in parallel. A correlation was generally found between the level of M. gallisepticum in tracheal swabs and the effectiveness of the probe, although a predictably reactive level of mycoplasmas was not always detected. Treatment of clinical specimens with acetylcysteine to disrupt mucus improved the detection rate. Dot-blot hybridization with probe pMG4 enabled positive identification of M. gallisepticum at an early stage of infection, prior to the development of a serological response in the infected chicken. Results are obtainable within 4 days of sampling, much more rapidly than culture, and also in clinical specimens from which mycoplasma isolation is impossible, such as carcasses. The results indicate that the use of DNA probes for the early and rapid detection of M. gallisepticum infection is feasible; a development which can replace laborious culture techniques and less effective serological methods, and thus reduce the time required for diagnosis.     

Experimental infection of ducks with Mycoplasma gallisepticum

Specific-pathogen-free ducks 24 and 180 days old were inoculated intranasally with the S6 strain of Mycoplasma gallisepticum (MG). No significant gross lesions were found in trachea, lung or air sacs at 7 or 28 days postinfection (PI). MG was recovered from the infraorbital sinus and trachea but not from the air sacs 7 and 28 days PI. A few ducks responded serologically by developing agglutinating antibody. MG multiplied in embryonated duck eggs but to lower titers than in embryonated chicken eggs.     

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.     

延胡索酸泰妙菌素对支原体和大肠杆菌混合感染的治疗试验

以人工诱发鸡毒支原体和大肠杆菌混合感染为模型,以酒石酸泰乐菌素为对照药物,评价了延胡索酸泰妙菌素的疗效.按每1 L水中分别加入312.5、468、625 mg延胡索酸泰妙菌素及500 mg酒石酸泰乐菌素的用量给病鸡饮水给药,连用5 d.试验表明,用药组的成活率、日增重、料肉比、气囊损伤度与感染对照组比较差异极显著(P＜0.01);延胡索酸泰妙菌素大剂量组日增重与小剂量组比较差异显著(P＜0.05),与其他各用药组比较差异极显著(P＜0.01),料肉比与其他各用药组比较差异极显著(P＜0.01);酒石酸泰乐菌素组的料肉比与延胡索酸泰妙菌素小剂量组比较差异不显著(P＞0.05);而与其他各组比较差异极显著(P＜0.01).数据分析表明,延胡索酸泰妙菌素大剂量组能有效地降低气囊损伤度,提高饲料转化率.