Influence of bovine blood serum on growth properties of nutrient media for Mycoplasma gallisepticum and Mycoplasma synoviae cultivation

The growth properties of the nutrient medium for the cultivation of pathogenic mycoplasmas depend on the type of blood serum it is supplemented with. Comparative tests of two cell-free nutrient media supplemented with bovine and porcine blood sera for the cultivation of strains “S6” Mycoplasma gallisepticum and “WVU 1853” Mycoplasma synoviae were performed. Growth properties of the tested nutrient media were assessed by determining the activity of the resulting biomass in the hemagglutination and agglutination assays, as well as by determining  the concentration of viable cells after the 9th passage. It has been shown that a cell-free nutrient medium supplemented with the porcine blood serum is optimal for the cultivation of pathogenic mycoplasma species causing infectious diseases in birds. The hemagglutinating activity of the Mycoplasma gallisepticum culture reached 5 HAU log₂ after 72 hours of cultivation, the agglutinating activity of Mycoplasma synoviae reached 5 AU log₂  during the 88-hour incubation period, the concentration  of viable cells of both strains was 10⁶ CFU/cm³. The low growth properties of the medium  prepared  with the addition  of bovine blood serum are most likely associated with its biochemical composition, which contains 5–20 times more provitamin A than the porcine blood serum, and high density lipoprotein cholesterol. On the contrary, in the porcine blood serum, most of the lipoproteins have a low density, containing a large amount of fatty acids and cholesterol, which are the main structural elements of mycoplasma cells. The obtained test results are of practical value and can be used in the technology of cultivation of pathogenic  species of avian mycoplasmas in the production of diagnostic and preventive tools.

1. Borchsenius S. N., Chernova O. A. Mycoplasmas: molecular and cellular biology, pathogenicity, diagnostics. Leningrad: Nauka; 1989. 156 р. (in Russ.)

2. Kagan G. Ya., Rakovskaya I. V. Mycoplasma infection in tissue cultures. Leningrad: Meditsina; 1968; 161. (in Russ.)

3. Prozorovskiĭ S. V., Rakovskaya I. V., Vulfovich Yu. V. Medical mycoplasmology. Moscow: Meditsina; 1995. 286 p. (in Russ.)

4. Prozorovskiĭ S. V., Pronin A. V., Sanin A. V. Immunologicheskie mekhanizmy persistentsii mikoplazm = Immunologic mechanisms of the persistence of Mycoplasma. Vestnik Akademii Meditsinskikh Nauk SSSR. 1985; 10: 43–51. PMID: 4082756. (in Russ.)

5. Yadav J. P., Tomar P., Singh Y., Khurana S. K. Insights on Mycoplasma gallisepticum and Mycoplasma synoviae infection in poultry: a systematic review. Anim. Biotechnol. 2021; DOI: 10.1080/10495398.2021.1908316.

6. Gautier-Bouchardon A. V. Antimicrobial resistance in Mycoplasma spp. Microbiol. Spectr. 2018; 6 (4). DOI: 10.1128/microbiolspec.ARBA-0030-2018.

7. Bessarabov B. F. Poultry diseases. Мoscow: Kolos; 1970; 80–82. (in Russ.)

8. Malushko V. V. Bor’ba s infektsionnymi boleznyami v promyshlennom ptitsevodstve = Infectious disease control in commercial poultry production. Veterinariya. 1982; 5: 36–37. (in Russ.)

9. Bessarabov B. F., Vasilevich F. I., Mel’nikova I. I., et al. Guide on avian diseases. Moscow: KolosS; 2005. 200 p. (in Russ.)

10. Animal mycoplasmoses. Ed. by Ya. R. Kovalenko. Мoscow: Kolos; 1976. 304 p. (in Russ.)

11. Kleven S. H. Mycoplasma synoviae infection. In: Deseases of Poultry. Ed. by Y. M. Saif, H. J. Barnes, J. R. Glisson, A. M. Fadly, L. R. McDougald, D. E. Swayne. 11th ed. Ames: Iowa State University Press; 2003; 756–766.

12. Irza V. N. Infectious synovitis birds – epizootiology and prevention. Ptitsevodstvo. 2009; 11: 39–40. eLIBRARY ID: 17089360. (in Russ.)

13. Korovin R. N., ZelenskyV. P., GroshevaG. A. Laboratory diagnosis of avian diseases: Reference guide. Мoscow: Agropromizdat; 1989; 172–174. (in Russ.)

14. Kulappu Arachchige S. N., Kanci Condello A., Zhu L., Shil P. K., Tivendale K. A., Underwood G. J., et al. Effects of immunosuppression on the efficacy of vaccination against Mycoplasma gallisepticum infection in chickens. Vet. Microbiol. 2021; 260:109182. DOI: 10.1016/j.vetmic.2021.109182.

15. Irza V. N., Sorokina M. I., Chernyaeva T. Yu., Volkov M. S., Zhalin M. V., Kolotilov A. N. Methods for refreshing and maintaining the Mycoplasma gallisepticum “S6”production strain. Vladimir: FGI“ARRIAH”; 2006. 7 p. (in Russ.)

16. Irza V. N., Volkov M. S., Chernyaeva T. Yu., Sorokina M. I., Kolotilov A. N. Methods for refreshing and maintaining Mycoplasma synoviae “WVU 1853” production strain. Vladimir: FGI “ARRIAH”; 2006. 7 p. (in Russ.)

17. Chernyaeva T. Yu., Sorokina M. I., Kozlov D. A., Volkov M. S. Guidelines for the determination of hemagglutinating activity of the causative agent of avian mycoplasmosis (Mycoplasma gallisepticum): approved by FGBI “ARRIAH”. December 23, 2020. No. 95-20. Vladimir: FGBI “ARRIAH”; 2020. 29 p. (in Russ.)

18. Girin M. V., Borisov A. V., Kulakov V. Yu., Sorokina M. I., Irza V. N. Izmerenie rosta Mycoplasma gallisepticum = Mycoplasma gallisepticum growth measurement. Dostizheniya molodykh uchenykh – v veterinarnuyu praktiku: materialy konferentsii molodykh uchenykh (16–18 noyabrya 2000 g.) = Achievements of early carrier researchers – to veterinary practice: proceedings of the conference of early carrier researchers (November 16–18, 2000). Vladimir: ARRIAH; 2000; 38–42. (in Russ.)

19. Kondrakhin I. P., Talanov G. A., Pak V. V. Internal non-communicable diseases of animals: a textbook. Moscow: KolosS; 2003. 461 p. (in Russ.)

20. Motuzko I. S., Khvostova O. V. Lipoprotein age dynamics of bovine blood serum. Vestnik of Vitebsk State Medical University. 2003; 2 (4): 116–118. Available at: https://www.elib.vsmu.by/bitstream/123/8975/1/vVG-MU_2003_4_116-118.pdf. (in Russ.)

21. Lipoproteiny. Vidy i fiziologiya lipoproteinov = Lipoproteins. Types and physiology of lipoproteins. MedUniver. Available at: https://meduniver.com/Medical/Physiology/1184.html (date of access: 27.08.2021). (in Russ.)