ELIMINATION OF MYCOPLASMA FROM THE TURKEY SEMEN

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1 Bull. Vet. Inst. Pulawy 46, 11-15, 2002 ELIMINATION OF MYCOPLASMA FROM THE TURKEY SEMEN GRZEGORZ TOMCZYK AND ZENON MINTA Department of Poultry Diseases, National Veterinary Research Institute, Pulawy, Poland The studies were carried out in a breeding turkey flock in which mycoplasma infections were detected (serologically and by isolation from semen). Trials using tylosin and enrofloxacin were carried out to limit the infection. The antibiotics were given to the birds for 5 days in therapeutic doses and to semen at concentration of 1 mg/ ml of semen. The effectiveness of such treatments was confirmed by negative results of mycoplasma isolation from semen of turkey males and from the cloaca of turkey hens. As a result, an improvement in reproductive performances was observed. Hatchability was higher by 15% and 27% after the addition of enrofloxacin and tylosin, respectively.the progeny were free of mycoplasmas. The effectiveness of the method used was confirmed during two consecutive months. Key words: turkeys, mycoplasma, eradication, semen, antibiotics. Mycoplasma infections are one of the most important problems in the turkey production. They affect respiratory (8, 13), locomotory (1, 2) and reproductive systems (10). Direct contact of infected and non-infected birds (9, 13) and also genital infections (5, 7, 13) are the main causes of the spread of this disease. Moreover, mycoplasmas are transmitted through the egg to offspring (6, 12, 13). The semen from infected males contaminates clean semen when pooled for insemination. Mycoplasma eradication programs involve the removal of infected birds from the flocks after serological monitoring or isolation of mycoplasma. But the infected flocks have sometimes to be kept for other breading reasons. The treatment of eggs and turkey semen with an effective antibiotics are useful methods to reduce genical infection (3, 5). The aim of presented study was eradication of mycoplasmas from breeder turkeys by adding antibiotics to the infected semen of stags and treatment of the femals. Material and Methods Experimental animals. Thirty-six-week-old BUT BIG 6 turkeys obtained from breeding flocks (2880 females and 400 stags) were used. The health status of the birds was known. Mortality in birds ageing from 1 to 28 weeks and from 29 to 36 weeks was 8.5% and 2.5%, respectively. Before experiments the flock was examined serologically by the plate agglutination test and ELISA and bacteriologically using PPLO selective media. This flock manifested a drop in egg production and fertility.

2 12 Media for mycoplasma culture. B medium supplemented with 0.2% L arginine and PPLO agar were used for the propagation of mycoplasma in the in vitro culture and the primary isolation of mycoplasma (4). Drugs. Tylosin tartrate (Tylan) Elanco and enrofloxacin (Baytril) Bayer were used in the experiments. Our early studies on mycoplasma isolated from turkeys revealed their sensitivity to these antibiotics. Culture of mycoplasma sampled from birds and embryos. Cloaca swabs from females, semen of stags and the yolk sacs of embryos were cultured. Administration of the drugs. The eradication program was performed for 8 weeks in birds assigned to three individual groups. This program contained treatment with tylosin 0.5g/l of drinking water for 5 days and enrofloxacin at the same dose. The females were treated with tylosin and enrofloxacin twice: at the beginning of the eradication program (5th week of laying season) and in four weeks after the start of the program (9th week of laying season). The stags were treated once with tylosin at the beginning of the eradication and each insemination dose of semen was supplemented with tylosin (Group 1) or enrofloxacin (Group 2) at a dose of 1 mg/ml of semen for 8 weeks. Birds in group 3 (controls) were not treated. Results The results of mycoplasma isolation from turkeys are presented in Table 1. The mycoplasma strains isolated from this flock were identified as Mycoplasma meleagridis (MM). Before treatment, 28% 40%, and 28% of semen samples, cloaca swabs and yolk sacs of embryos, respectively, were infected, whereas no positive mycoplasma isolations were found after a 4 and 8-week eradication program realisation. The mycoplasmas were isolated from all the controls. Sample Semen 7/25 (28) Cloaca swabs Table 1 Isolation of mycoplasma from turkeys Mycoplasma isolation before treatment 4 weeks after 8 weeks after antibiotic treatment antibiotic treatment (%) (%) (%) 6/25 3 (24) 6/25 3 (24) 7/25 4 (28) 7/25 4 (28) 10/25 (40) 8/25 3 (32) 9/25 4 (36) 0/10 1 0/10 2 3/10 3 (30) 4/10 4 (40) 8/25 3 (32) 11/25 4 (44) 0/10 1 Embryos (yolk sac) 7/25 (28) 0/10 2 4/10 3 (40) 4/10 4 (40) 1 treated with tylosin; 2 treated with enrofloxacin; 3 control for tylosin; 4 control for enrofloxacin

3 13 Further examinations revealed that the effectiveness of the enrofloxacin and tylosin treatment was markedly higher in the case of the fertility of eggs in comparison to that of the controls. Table 2 Effect of mycoplasma eradication program on the turkey semen (group treated with enrofloxacin) Weeks of Fertility of eggs (%), group of birds eradication Enrofloxacin* Control (not treated) Difference I II III IV V VI *semen for insemination treated with enrofloxacin Table 3 Effect of mycoplasma eradication program on the turkey semen (group treated with tylosin) Weeks of Fertility of eggs (%), group of birds eradication Tylosin* Control (not treated) Difference I II III IV V VI *semen for insemination treated with tylosin In further studies, the reproductive performance traits (infertile eggs, dead embryos and the percentage of hatchability) were compared between treated and control groups after 8 weeks. The results are presented in Table 4.

4 14 Table 4 Reproductive performance (after 8 weeks of eradication program) Reproductive traits No of set eggs Group of reproductors (treated) Tylosin Enrofloxacin Control 1 (not treated) 735 Control 2 (not treated) 651 Infertile eggs 249 (46.1) (53.4) 461 (62.7) 452 (69.4) Dead embryos 0 (0) 7 (1.8) 2 (0.4) 3 (0.6) Percent of hatchability control group for tylosin; 2 control group for enroflaxacin; 3 percentage 30.6 Discussion It was found that the eradication program applied in breeding turkeys infected with mycoplasma was effective. Results of the examination of Mycoplasma meleagridis (MM) occurrence by the microbe isolation from the reproductive system and by serological examinations revealed, before the treatment, a 40% and 28% incidence of the infection in examined females (cloaca swabs) and in the semen of stage donors, respectively. Both tylosin and enrofloxacin used in a regular treatment of birds and as an additive to the semen for insemination completely eliminated mycoplasma occurrence in semen samples, cloaca swabs from laying turkey hens and in their offspring after a 4-week realisation of the program whereas the control birds revealed a 24% to 44% incidence of MM infection during the whole experimental period (8 weeks). In breeding turkeys, attempts to reduce the egg-transmission rate are made in numerous countries (13, 14): MM eradication program in breeding flocks has markedly reduced the MM prevalence. Currently, the economic losses caused by MM infection in turkeys have been reduced significantly. However, the vertical transmission of MM infection plays a significant role (3, 11); laying turkey hens may be infected while mated or by infected semen (12, 14). Tylosin and other therapeutics (5, 13) effectively disrupt MM infection and can reduce the microbe transmission. In our experiment tylosin and enrofloxacin used in regular treatments and added to the turkey semen increased the fertility rate of eggs by 3.8% and 16.5% after 2-week and 6-week treatments, respectively (Tables 2 and 3). In the case of tylosin a 23.1% increase resulting from 4-week eradication program was found. This procedure similarly to dipping of eggs in antibiotics solution (5) may be used to improve hatchability and to obtain the MM-free progeny. MM and other mycoplasma infections markedly affect fertility and hatchability of eggs especially in the late stage of hatching. In the experiment carried out in our laboratory, the use of the eradication program significantly increased the egg fertilisation rate by 16.6% and 6.7% in the groups treated with tylosin and enrofloxacin, respectively (Tab.4). Moreover, tylosin (Tab.4) effectively increased hatchability and reduced embryo mortality.

5 15 References 1. Chin R.P.,. Daft B.M, Meteyer C.U., Yamamoto R.: Meningoencephalitis in commercial meat turkeys associated with Mycoplasma gallisepticum. Avian Dis., 1991, 35, Dingfelder R.S., Ley D.H., Mc Laren J.M., Borwnie C.: Experimental infection of turkeys with Mycoplasma gallisepticum of low virulence, transmissibility, and immunogenicity. Avian Dis., 1991, 35, Edson R.K., Yammamoto R., Farver T.B.: Mycoplasma meleagridis of turkeys: Probability of eliminating egg-borne infection. Avian Dis., 1987, 31, Frey M.L.,Hawk S.T, Anderson D. P.: A medium for the isolation of avian Mycoplasmas. Am. J. Vet. Res., 1968, 29, Ghazikhanian G., Yamamoto R., McCapes R.H., Dungan W.M., Ortmayer H.B.: Combination dip and injection of turkey eggs with antibiotics to eliminate Mycoplasma meleagridis infection from a primary breeding stock. Avian Dis., 1980, 24, Kleven S.H., Pomeroy B.S.: Role of the female in egg transmission of Mycoplasma meleagridis in turkeys. Avian Dis., 1971, 15, Kumar M.C., Pomeroy B.S.: Transmission of Mycoplasma meleagridis in turkeys. Am. J. Vet. Res., 1969, 30, Minta Z., Tomczyk G.: Mykoplazmozy indyków. VIII Sympozjum Drobiarskie.Polanica Zdrój, ,1997, Mohamed Y.S., E.H. Bohl.: Studies on the transmission of Mycoplasma meleagridis. Avian Dis., 1967, 11, Rhoades K.R.: Experimentaly induced Mycoplasma meleagridis infection of turkey reproductive tracts. Avian Dis., 1969, 13, Yamamoto R., Ortmayer H.B.: Hatcher and intraflock transmision of Mycoplasma meleagridis. Avian Dis., 1967, 11, Yammamoto R., Ortmayer H.B.: Egg transmission of Mycoplasma meleagridis in naturaly infected turkeys under different mating systems. Poult. Sci., 1969, 48, Yammamoto R.: Mycoplasma meleagridis infection. In B.W. Calnek, H.J. Beard, W.M. Reid, and H.W. Yoder, Jr. Diseases of Poultry, (9-th ed). edited by Iowa State University Press, Ames, IA, 1991, pp Yoder H.W.Jr.: Preincubation neat treatment of chicken hatching eggs to inactivate Mycoplasma. Avian Dis., 1970, 14,

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