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.     

Preliminary data on efficacy of Mycoplasma gallisepticum vaccines containing different adjuvants in laying hens

Chickens were vaccinated subcutaneously twice, at 13 and 17 weeks of age. The vaccines used were the whole organisms of Mycoplasma gallisepticum (MG) adjuvanted with multilamellar positively charged (MPC) liposomes or oil-emulsion. Other chickens received the same bacterins but supplemented with Salmonella typhimurium cell wall protein mitogen (STP) (50 micrograms/dose). At 21 weeks of age, each bird was challenged in the right and left caudal thoracic air sacs. The challenge dose/chicken was 1.3 x 10(5) CFU of MG (R-strain). A significant immunoglobulin (Ig) response specific to MG was observed in sera of chickens collected 3 weeks after the first and second vaccination with MG adjuvanted with MPC liposomes or oil-emulsion. The same two treatments had highly significant MG-titers in eggs collected during the first and second month post challenge. Both groups had highly significant protection (P less than 0.05) against MG transmission in eggs layed during the first month post challenge. Vaccination with MG organisms adjuvanted to MPC liposomes or oil-emulsion resulted in higher egg production, during the first month following challenge, in comparison to the unvaccinated-challenged birds; the same two groups had higher egg production in the second month following challenge compared to unvaccinated-challenged birds, but not significantly different (P greater than 0.05). The addition of STP to bacterins containing MG organisms adjuvanted to MPC liposomes or oil-emulsion, resulted in a significant reduction (P less than 0.05) of the Ig-specific to MG in sera and in a significant drop in egg production (P less than 0.05) during the first month following challenge.     

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.     

Evaluation of a Mycoplasma gallisepticum strain exhibiting reduced virulence for prevention and control of poultry mycoplasmosis.

Two experiments were conducted to evaluate the virulence and vaccination efficacy of a Mycoplasma gallisepticum (MG) isolate designated MG Intervet 6/85. Virulence of the strain was determined by evaluation of airsacculitis scores following aerosol exposure to the isolate before and after 10 sequential passes in either commercial broiler chickens or commercial turkeys. Two-week-old specific-pathogen-free chickens were vaccinated by aerosol exposure. The birds were challenged with the R' strain of MG at either 4 or 8 weeks post-vaccination. Efficacy was evaluated by airsacculitis scores determined 21 days after challenge. Ten repetitive back-passes of the isolate in chickens and turkeys did not substantially increase the virulence. Virulence for both chickens and turkeys was minimal, while protection elicited by aerosol vaccination in young chickens against virulent R' strain was significant (P less than or equal to 0.05) compared with unvaccinated controls.     

Protection and immunity in commercial chicken layers administered Mycoplasma gallisepticum liposomal bacterins

Six liposomal Mycoplasma gallisepticum (MG) bacterins, differing in charge and size, and two oil-emulsion vaccines (sonicated and non-sonicated) were given to white leghorns in two doses, at 13 weeks and again 1 month later. At 21 weeks of age, all chickens were challenged with a viable 20-hour culture of MG cells (17,800 colony-forming units) intratracheally and with nonviable MG organisms (0.09 mg protein) injected subcutaneously in the wattle center. The three chicken groups that had the lowest tracheal MG-infection rates postchallenge were those given adjuvants of small multilamellar positively charged liposomes (16.67%), large multilamellar negatively charged liposomes (16.67%), and non-sonicated oil-emulsion bacterin (37.5%). These three groups also had significant levels of antibody in sera 4 weeks after the second dose of vaccine. The group given the small multilamellar positively charged liposome also showed significant delayed-type hypersensitivity (wattle swelling) (P less than or equal to 0.05). The group given the large multilamellar negatively charged liposomes had the highest local antibody response (P less than or equal to 0.01) and was the only group that had no microscopic lesions in the trachea.     

Effect of Mycoplasma gallisepticum bacterin on egg-transmission and egg production

Groups of white leghorn hens were vaccinated twice with a Mycoplasma gallisepticum (MG) bacterin, once with bacterin, or left unvaccinated. Four weeks after vaccination, they were challenged with virulent R strain MG. Egg production was significantly higher in challenged vaccinated groups than in the challenged control group. Four challenged control hens went out of production, whereas only one twice-vaccinated hen did. MG was first isolated directly from eggs 5 days postchallenge (PC) in twice-vaccinated hens, 4 days PC in once-vaccinated hens, and 2 days PC in controls, and the hens continued to lay positive eggs till the end of the experiment 7 weeks PC. MG was found in 17.65%, 38.55%, and 45.90% of eggs cultured in twice-vaccinated, once-vaccinated, and control groups, respectively. Nine of 16 twice-vaccinated hens were found to be shedding MG through their eggs, whereas 15 of 17 once-vaccinated hens and 14 of 16 controls were shedding MG through their eggs.     

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.     

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.     

Identification of the antigenic components of the virulent Mycoplasma gallisepticum (R) in chickens: their role in differentiation from the vaccine strain (F)

The antibody response to different proteins of Mycoplasma gallisepticum (MG) was studied in chickens experimentally infected with virulent MG R strain. The chickens were challenged at 8 weeks of age by the intranasal route. Each cockerel received 1.3 X 10(6) colony-forming units (CFU). MG strains (R and F) were banded by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). The banding pattern was distinctively different between the two strains in the range of 92.5 to 200 kilodaltons (kD). Chicken sera collected at different times following challenge were analyzed by Western blot to determine the patterns of antibodies raised to specific MG proteins (R versus F strains). Early in infection (2 weeks postchallenge), antibodies to 60-kD and 75-kD polypeptides of MG R strain were produced. Subsequently (greater than or equal to 4 weeks postchallenge), antibodies recognized a larger number of MG antigens in both strains. The immunoblot patterns remained the same in the period 8-11 weeks postinfection in each of the two strains; however, the patterns were different when the two strains were compared. The early response recognized the 75-kD protein in the R strain while it recognized the 80-kD protein in the F strain. The late response recognized the 130-kD protein and the protein slightly heavier than 200 kD in the R strain. These two bands did not appear in the immunoblot performed against the F strain of MG. Electroeluted protein of MG R strain, namely adhesin (75 kD), showed a hemagglutination activity (HA) on chicken red blood cells. With the appearance of antibodies specific to the 60-kD and 75-kD polypeptides, there was a significant rise in hemagglutination-inhibition geometric mean titer of chicken sera.     

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.     

Elimination of mycoplasmal plate agglutination cross-reactions in sera from chickens inoculated with infectious bursal disease viruses

Sera from chickens inoculated with various challenge infectious bursal disease viruses or infectious bursal disease vaccines were found to cross-react in the Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) serum plate agglutination (SPA) tests. Two-fold dilutions of these cross-reacting sera with phosphate-buffered saline before retesting eliminated all non-specific agglutination in the MG and MS SPA tests. Cross-reactions were observed in the SPA test using sera from chickens inoculated with either MG or MS. Dilutions of these sera 1:2 had little effect on the number of these cross-reactions. At 1:4 serum dilutions, however, the number of cross-reactions between MG and MS was reduced. At 1:8 dilution of test sera, cross-reactions between MG and MS were further reduced. Some reduction in specific MG and MS SPA reactions, however, also occurred at the 1:8 dilution of sera with some of the plate antigen.     

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.     

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.     

Comparison of Mycoplasma gallisepticum subunit and whole organism vaccines containing different adjuvants by western immunoblotting

Chickens were vaccinated with subunit (adhesin protein) or whole organisms of Mycoplasma gallisepticum (MG) adjuvanted with multilamellar positively charged liposomes or oil-emulsion. Sera were collected before and following the first (13 weeks of age) and second (17 weeks of age) vaccination. The chicken sera were used in western immunoblotting against whole MG polypeptides. Vaccination with the subunit (MG-adhesin) bacterin containing positively charged liposomes resulted in antibody response specific to adhesin band (75 kD) at 3 weeks post the first and second vaccination; however, crossreactions of the same antibodies occurred to MG proteins of 85 kD (3 weeks after the first vaccination) and 56 kD (3 weeks after the second vaccination). Vaccination with whole MG proteins containing positively charged liposomes resulted in significant immunopotentiation of antibodies against low molecular weight polypeptides of MG (less than 48.0 kD). The addition of Salmonella typhimurium cell wall proteins mitogens (STP) to the different bacterins suppressed the antibody responses to some MG polypeptides.     

Further studies on the immunization of chickens with temperature-sensitive Mycoplasma gallisepticum mutant

Newly hatched chickens were immunized with a temperature-sensitive (TS) Mycoplasma gallisepticum (MG) mutant (TS 100). Immunized chickens resisted challenge with the virulent S6 strain. The dose of TS MG needed for protection was less than 3.3 X 10(4) colony-forming units. After immunization with TS 100, chickens were subjected to a variety of virus infection and immunosuppressive treatments. Neonatal bursectomy or thymectomy, infectious bursal disease virus infection, and infectious bronchitis virus vaccination or a combination of infectious bronchitis virus and Newcastle disease virus vaccination did not contribute to the development of air-sac lesions in TS MG-immunized chickens. Infectious bronchitis virus vaccination enabled MG to migrate from the nasal cavity to the trachea but not to the air sacs. The TS MG vaccine appears to be a safe immunogen.     

Detection of serum antibodies against Mycoplasma gallisepticum and Mycoplasma synoviae by a dot-immunobinding technique

Both Mycoplasma gallisepticum (MG) and M. synoviae (MS) antigens prepared for the routine haemagglutination inhibition (HI) test were diluted and absorbed to the separate pieces of durapore membrane for the measurement of dot-immunobinding (DIB) titers of test sera. Besides, durapore strips bearing both antigens were employed for a DIB test with chicken sera definitely diluted 100-fold. Shortening of reaction time of chicken sera with antigens as well as with the secondary serum markedly eliminated non-specific DIB reactions exhibited at low dilutions although the same condition was not so effective on the elimination of non-specific reactions among rabbit hyperimmune sera. Rapid and specific development of DIB antibody which continued at high titer up to 1:640 for 10 weeks postinoculation was proved in the sera of SPF chickens inoculated with MG or MS, while DIB titers of sera from uninoculated chickens remained 1:20 or lower. Non-specific reactions, which occurred in the routine serum plate agglutination test with a part of sera from the inoculated chickens, were not exhibited in the DIB as well as in the HI test with the same sera. Results of the DIB test with serum samples from 287 conventionally reared chickens definitely diluted 100-fold coincided with the results of HI test at a level of 90% with MG and 89% with MS antigen. This technique seems to be useful for a rapid, simple and specific diagnosis of avian mycoplasmosis.     

Duration of immunity induced in chickens by an attenuated live Salmonella enteritidis vaccine and an inactivated Salmonella enteritidis/typhimurium vaccine

The aim of this study was to examine the duration of immunity of different vaccination schemes using the S. enteritidis live vaccine Gallivac Se and the S. enteritidis-S. typhimurium inactivated vaccine Gallimune Se+St. Three groups of Lohman Brown chickens were used. Group one was vaccinated three times orally with Gallivac Se at weeks one, seven and 13 of age. Group two was vaccinated twice orally with Gallivac Se in weeks one and seven and once i.m. with Gallimune Se+St in week 14 of age. A third group was not vaccinated and served as the control group. Eight randomly selected chickens from each of the three groups were challenged with a nalidixic acid resistant S. enteritidis PT4 strain in weeks 24, 51 and 71 of age and the same number of animals were challenged with a S. typhimurium DT 104 strain in weeks 26, 54 and 73 (75) of age.The chickens were euthanised seven days post challenge and the number of challenge strain organisms (log10 cfu) in the liver and on caecal mucosa was determined.The quantitative investigation of the challenge strain in the liver and caecal mucosa revealed a statistically significant (p < 0.05) lower challenge strain burden in the vaccinated groups compared with the non-vaccinated control group up to week 71 (73) of age. The protective effects were demonstrated for both challenge strains.     

Resistance to fecal shedding of salmonellae in pigs and chickens vaccinated with an aromatic-dependent mutant of Salmonella typhimurium.

The purpose of this study was to evaluate the effectiveness of an aromatic-dependent mutant of Salmonella typhimurium as a parenteral vaccine for prevention of fecal shedding of Salmonella spp. Pigs and chickens were vaccinated IM, with 1 x 10(9) and 1 x 10(8) organisms, respectively, followed by a second identical vaccination 2 weeks later. Salmonella organisms were not detected by analysis of fecal or cloacal swab specimens from any animal after vaccination. Deleterious side effects were not noticed after vaccination. Pigs were challenge-inoculated PO with 1 x 10(12) virulent S typhimurium 1 week after the second vaccination. Chickens were challenge-inoculated PO with 3 x 10(8) organisms of either S enteritidis or the virulent parent strain of S typhimurium 3 weeks after the second vaccination. Vaccinated pigs shed Salmonella spp significantly less frequently than did nonvaccinated pigs. Vaccinated chickens challenge-inoculated with either S enteritidis or S typhimurium also shed Salmonella less frequently than the corresponding nonvaccinated control birds; however, the difference was not significant.     

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.     

Changes of some metabolic parameters of the liver in chickens under the influence of Entomoxan]

From the 3rd week of life Studler-Cornish-breed chickens received a food mixed with Entomoxan (15 ppm HCH-gamma-Isomer in the food) for an 8 weeks period. 2,4,6, and 8 weeks after the beginning of the experiment livers of the chickens were examined for RNA-, DNA-, total protein- and total aminonitrogen contents. The RNA-values decrease constantly where as those of DNA increase up to the 4th week and after that decline. Total protein contents rise up to the 4th week. After 2 and 8 weeks the contents of amino-nitrogen are lower in experimental birds than in controls.