STUDY OF ANTIMICROBIAL ACTIVITY OF PROBIOTIC STRAINS OF BACILLUS
Keywords:
calves, bacterial antagonism, antibiotic resistance, B. amyloliquefaciense, B. mucilaginosus, B. coagulans, B. megaterium, B. pumilus
Abstract
Improper and uncontrolled use of antimicrobials in livestock can lead to increased antibiotic resistance and affect animal and human health. The research were conducted in the farm of Holstein cattle LLC AF “Lan” Ukraine in March-April 2021. We monitored microorganisms on the farm, determined their number and species. Endo agar was used as the elective medium for Escherichia; Staphylococcus aureus was determined on Chistovich's agar, and fungi and yeast were determined on Saburo's agar. Polymerase chain reaction was used to determine Mycoplasma spp.. Antagonistic properties of probiotic strains of Bacillus spp. by diffusion into agar wells were also determined. The size of growth inhibition zone around different strains was determined in mm: Bacillus amyloliquefaciense NR 59, Bacillus mucilaginosus ACH 82, Bacillus coagulans ALM86, Bacillus megaterium NCH 55, Bacillus pumilus LA 56 in a dilution of 1 × 109, CFU/g. Disks with the antibiotic cephalexin were used as control. An appropriate strain of probiotic microorganism was poured into each well of meatpeptone agar with the appropriate isolate. Then incubated for 24 hours at 37 ºC and determined the demarcation zone around each well. The main pathogens of dairy calves on the farm are identified: S. agalactiae (23 %), S. aureus (11 %), S. epidermidis (18 %), E. fecalis (10 %), E. coli (12 %), Mycoplasma spp. (7 %), fungi Candida (9 %) and associated microflora (10 %). Three probiotic strains of microorganisms were identified, to which microorganisms which were isolated in the indoor of calf showed the greatest sensitivity. It was found that Bacillus coagulans ALM 86 showed more antagonistic properties compared to the antibiotic against S. agalactiae - by 18.93%; Candida - by 29.16%; S. aureus - by 15.56%. Bacillus pumilus LA 56 strain inhibited the colony's growth of S. epidermidis by 20.49%; E. coli 28.78%; Candida - by 7.33% more than cephalexin. B. megaterium NCH 55 showed antimicrobial properties against S. aureus and E. fecalis identical to the antibiotic cephalexin. As a result of the conducted research probiotics which can become an alternative of antibiotics are defined. The prospect of further research in this direction is to determine the mechanism of action of probiotics Bacillus megaterium NCH 55, Bacillus coagulans ALM 86 and Bacillus pumilus LA 56 on pathogenic microorganisms and determine the therapeutic effect on animals.
References
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15. Izuddin, W. I., Humam, A. M., Loh, T. C., Foo, H. L., & Samsudin, A. A. (2020). Dietary Postbiotic Lactobacillus plantarum Improves Serum and Ruminal Antioxidant Activity and Upregulates Hepatic Antioxidant Enzymes and Ruminal Barrier Function in Post-Weaning Lambs. Antioxidants (Basel, Switzerland), 9(3), 250. https://doi.org/doi.org/10.3390/antiox9030250
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17. Kong, L., Yang, C., Dong, L., Diao, Q., Si, B., Ma, J., & Tu, Y. (2019). Rumen Fermentation Characteristics in Preand Post-Weaning Calves upon Feeding with Mulberry Leaf Flavonoids and Candida tropicalis Individually or in Combination as a Supplement. Animals : an open access journal from MDPI, 9(11), 990. https://doi.org/10.3390/ani9110990
18. Maier, G. U., Love, W. J., Karle, B. M., Dubrovsky, S. A., Williams, D. R., Champagne, J. D., Anderson, R. J., Rowe, J. D., Lehenbauer, T. W., Van Eenennaam, A. L., & Aly, S. S. (2019). Management factors associated with bovine respiratory disease in preweaned calves on California dairies: The BRD 100 study. Journal of dairy science, 102(8), 7288–7305. https://doi.org/10.3168/jds.2018-14773
19. Manyi-Loh, C., Mamphweli, S., Meyer, E., & Okoh, A. (2018). Antibiotic Use in Agriculture and Its Consequential Resistance in Environmental Sources: Potential Public Health Implications. Molecules (Basel, Switzerland), 23(4), 795. https://doi.org/10.3390/molecules23040795
20. Mao, S., Zhang, R., Wang, D., & Zhu, W. (2012). The diversity of the fecal bacterial community and its relationship with the concentration of volatile fatty acids in the feces during subacute rumen acidosis in dairy cows. BMC veterinary research, 8, 237. https://doi.org/10.1186/1746-6148-8-237
21. Mingmongkolchai, S., & Panbangred, W. (2018). Bacillus probiotics: an alternative to antibiotics for livestock production. Journal of applied microbiology, 124(6), 1334–1346. https://doi.org/10.1111/jam.13690
a. Nguyen, Tu HK., Thu, Le B. (2015). Evaluation of antimicrobial activities of Bacillus megaterium with a thirdgeneration cephalosporin (ceftriaxone). 5 (09):016-020.10.7324/JAPS.2015.50903https://www.japsonline.com/admin/php/uploads/1616_pdf.pdf
22. Ozutsumi, Y., Hayashi, H., Sakamoto, M., Itabashi, H., & Benno, Y. (2005). Culture-independent analysis of fecal microbiota in cattle. Bioscience, biotechnology, and biochemistry, 69(9), 1793–1797. https://doi.org/10.1271/bbb.69.1793
23. Palma-Hidalgo, J. M., Jiménez, E., Popova, M., Morgavi, D. P., Martín-García, A. I., Yáñez-Ruiz, D. R., & Belanche, A. (2021). Inoculation with rumen fluid in early life accelerates the rumen microbial development and favours the weaning process in goats. Animal microbiome, 3(1), 11. https://doi.org/10.1186/s42523-021-00073-9
24. Ruiz, L., Ruas-Madiedo, P., Gueimonde, M., de Los Reyes-Gavilán, C. G., Margolles, A., & Sánchez, B. (2011). How do bifidobacteria counteract environmental challenges? Mechanisms involved and physiological consequences. Genes & nutrition, 6(3), 307–318. https://doi.org/10.1007/s12263-010-0207-5
25. Rybachuk, Z., Shkromada, O., Predko, A., & Dudchenko, Y. (2020). Influence of probiotics “Immunobacterin-D” on biocenoses and development of the gastrointestinal tract of calves. Scientific Messenger of LNU of Veterinary Medicine and Biotechnologies. Series: Veterinary Sciences, 22(98), 22-27. https://doi.org/10.32718/nvlvet9804
26. Shkromada O., Skliar O., Paliy A., Ulko L., Gerun I., Naumenko О., Ishchenko K., Kysterna O., Musiienko O., Paliy A., 2019. Development of measures to improve milk quality and safety during production. Eastern-European Journal of Enterprise Technologies, 3/11(99), 30-39. https://doi.org/10.15587/1729-4061.2019.168762
27. Shkromada, O., Skliar, O., Pikhtirova, A. & Inessa, G. (2019). Pathogens Transmission and Cytological Composition of Cow’s Milk . Acta Veterinaria Eurasia, 45 (3) , 73-79 . https://dergipark.org.tr/tr/pub/actavet/issue/50595/608279
28. Tanih, N. F., Sekwadi, E., Ndip, R. N., & Bessong, P. O. (2015). Detection of pathogenic Escherichia coli and Staphylococcus aureus from cattle and pigs slaughtered in abattoirs in Vhembe District, South Africa. TheScientificWorldJournal, 2015, 195972. https://doi.org/10.1155/2015/195972
29. Wu, H.J., Sun, L.B., Li, C.B., Li, Z.Z., Zhang, Z., Wen, X.B., Hu, Z., Zhang, Y.L. (2014) Enhancement of the immune response and protection against Vibrio parahaemolyticus by indigenous probiotic Bacillus strains in mud crab (Scylla paramamosain). Fish Shellfish Immunol 41, 156– 162. https://doi.org/10.1016/j.fsi.2014.08.027
30. Yáñez-Ruiz, D. R., Abecia, L., & Newbold, C. J. (2015). Manipulating rumen microbiome and fermentation through interventions during early life: a review. Frontiers in microbiology, 6, 1133. https://doi.org/10.3389/fmicb.2015.01133
2. Benedictus, L., Ravesloot, L., Poppe, K., Daemen, I., Boerhout, E., van Strijp, J., Broere, F., Rutten, V., Koets, A., & Eisenberg, S. (2019). Immunization of young heifers with staphylococcal immune evasion proteins before natural exposure to Staphylococcus aureus induces a humoral immune response in serum and milk. BMC veterinary research, 15(1), 15. https://doi.org/10.1186/s12917-018-1765-9
3. Calvo-Lorenzo, M. S., Hulbert, L. E., Fowler, A. L., Louie, A., Gershwin, L. J., Pinkerton, K. E., Ballou, M. A., Klasing, K. C., & Mitloehner, F. M. (2016). Wooden hutch space allowance influences male Holstein calf health, performance, daily lying time, and respiratory immunity. Journal of dairy science, 99(6), 4678–4692. https://doi.org/10.3168/jds.2016-10888
4. Cheng, G., Hao, H., Xie, S., Wang, X., Dai, M., Huang, L. and Yuan, Z. (2014) Antibiotic alternatives: the substitution of antibiotics in animal husbandry? Front Microbiol 5, 69– 83. https://doi.org/10.3389/fmicb.2014.00217
5. da Silva Duarte, V., Treu, L., Sartori, C., Dias, R. S., da Silva Paes, I., Vieira, M. S., Santana, G. R., Marcondes, M. I., Giacomini, A., Corich, V., Campanaro, S., da Silva, C. C., & de Paula, S. O. (2020). Milk microbial composition of Brazilian dairy cows entering the dry period and genomic comparison between Staphylococcus aureus strains susceptible to the bacteriophage vB_SauM-UFV_DC4. Scientific reports, 10(1), 5520. https://doi.org/10.1038/s41598-020-62499-6
6. EFSA Panel on Biological Hazards (BIOHAZ), Ricci, A., Allende, A., Bolton, D., Chemaly, M., Davies, R., Fernández Escámez, P. S., Girones, R., Koutsoumanis, K., Lindqvist, R., Nørrung, B., Robertson, L., Ru, G., Sanaa, M., Simmons, M., Skandamis, P., Snary, E., Speybroeck, N., Kuile, B. T., Threlfall, J., … Herman, L. (2017). Risk for the development of Antimicrobial Resistance (AMR) due to feeding of calves with milk containing residues of antibiotics. EFSA journal. European Food Safety Authority, 15(1), e04665. https://doi.org/10.2903/j.efsa.2017.4665
7. Evans, S. E., Scott, B. L., Clement, C. G., Larson, D. T., Kontoyiannis, D., Lewis, R. E., Lasala, P. R., Pawlik, J., Peterson, J. W., Chopra, A. K., Klimpel, G., Bowden, G., Höök, M., Xu, Y., Tuvim, M. J., & Dickey, B. F. (2010). Stimulated innate resistance of lung epithelium protects mice broadly against bacteria and fungi. American journal of respiratory cell and molecular biology, 42(1), 40–50. https://doi.org/10.1165/rcmb.2008-0260OC
8. Garkavenko, TO, Gorbatyuk, OI, Kozytska, TG, Anriashchuk, VO, Garkavenko, VM, Dybkova, SM, Azirkina IM (2021) Methodical recommendations for determining the sensitivity of microorganisms to antibacterial drugs, K .: DNDILVSE, 101.
9. Govender, M., Choonara, Y. E., Kumar, P., du Toit, L. C., van Vuuren, S., & Pillay, V. (2014). A review of the advancements in probiotic delivery: Conventional vs. non-conventional formulations for intestinal flora supplementation. AAPS PharmSciTech, 15(1), 29–43. https://doi.org/10.1208/s12249-013-0027-1
10. Guterbock W. M. (2014). The impact of BRD: the current dairy experience. Animal health research reviews, 15(2), 130–134. https://doi.org/10.1017/S1466252314000140
11. Haldar, L., & Gandhi, D. N. (2016). Effect of oral administration of Bacillus coagulans B37 and Bacillus pumilus B9 strains on fecal coliforms, Lactobacillus and Bacillus spp. in rat animal model. Veterinary world, 9(7), 766–772. https://doi.org/10.14202/vetworld.2016.766-772
12. Hartung, T. (2010). Comparative analysis of the revised Directive 2010/63/EU for the protection of laboratory animals with its predecessor 86/609/EEC – a t4 report. ALTEX, 27(4), 285-303. doi: 10.14573/altex.2010.4.285
13. http://www.vetlabresearch.gov.ua/derzhavni-zakupivli/docs/%D0%90%D0%BD%D1%82%D0%B8%D0%B1%D1%96%D0%BE%D1%82%D0%B8%D0%BA%D0%BE%D1%80%D0%B5%D0%B7%D0%B8%D1%81%D1%82%D0%B5%D0%BD%D1%82%D0%BD%D1%96%D1%81%D1%82%D1%8C.pdf
14. Islam, K. S., Shiraj-Um-Mahmuda, S., & Hazzaz-Bin-Kabir, M. (2016). Antibiotic usage patterns in selected broiler farms of Bangladesh and their public health implications. Journal of Public Health in Developing Countries, 2(3), 276-284 https://www.jphdc.org/index.php/jphdc/article/view/84
15. Izuddin, W. I., Humam, A. M., Loh, T. C., Foo, H. L., & Samsudin, A. A. (2020). Dietary Postbiotic Lactobacillus plantarum Improves Serum and Ruminal Antioxidant Activity and Upregulates Hepatic Antioxidant Enzymes and Ruminal Barrier Function in Post-Weaning Lambs. Antioxidants (Basel, Switzerland), 9(3), 250. https://doi.org/doi.org/10.3390/antiox9030250
16. Köllmann, K., Wente, N., Zhang, Y., & Krömker, V. (2021). Investigations on Transfer of Pathogens between Foster Cows and Calves during the Suckling Period. Animals : an open access journal from MDPI, 11(9), 2738. https://doi.org/10.3390/ani11092738
17. Kong, L., Yang, C., Dong, L., Diao, Q., Si, B., Ma, J., & Tu, Y. (2019). Rumen Fermentation Characteristics in Preand Post-Weaning Calves upon Feeding with Mulberry Leaf Flavonoids and Candida tropicalis Individually or in Combination as a Supplement. Animals : an open access journal from MDPI, 9(11), 990. https://doi.org/10.3390/ani9110990
18. Maier, G. U., Love, W. J., Karle, B. M., Dubrovsky, S. A., Williams, D. R., Champagne, J. D., Anderson, R. J., Rowe, J. D., Lehenbauer, T. W., Van Eenennaam, A. L., & Aly, S. S. (2019). Management factors associated with bovine respiratory disease in preweaned calves on California dairies: The BRD 100 study. Journal of dairy science, 102(8), 7288–7305. https://doi.org/10.3168/jds.2018-14773
19. Manyi-Loh, C., Mamphweli, S., Meyer, E., & Okoh, A. (2018). Antibiotic Use in Agriculture and Its Consequential Resistance in Environmental Sources: Potential Public Health Implications. Molecules (Basel, Switzerland), 23(4), 795. https://doi.org/10.3390/molecules23040795
20. Mao, S., Zhang, R., Wang, D., & Zhu, W. (2012). The diversity of the fecal bacterial community and its relationship with the concentration of volatile fatty acids in the feces during subacute rumen acidosis in dairy cows. BMC veterinary research, 8, 237. https://doi.org/10.1186/1746-6148-8-237
21. Mingmongkolchai, S., & Panbangred, W. (2018). Bacillus probiotics: an alternative to antibiotics for livestock production. Journal of applied microbiology, 124(6), 1334–1346. https://doi.org/10.1111/jam.13690
a. Nguyen, Tu HK., Thu, Le B. (2015). Evaluation of antimicrobial activities of Bacillus megaterium with a thirdgeneration cephalosporin (ceftriaxone). 5 (09):016-020.10.7324/JAPS.2015.50903https://www.japsonline.com/admin/php/uploads/1616_pdf.pdf
22. Ozutsumi, Y., Hayashi, H., Sakamoto, M., Itabashi, H., & Benno, Y. (2005). Culture-independent analysis of fecal microbiota in cattle. Bioscience, biotechnology, and biochemistry, 69(9), 1793–1797. https://doi.org/10.1271/bbb.69.1793
23. Palma-Hidalgo, J. M., Jiménez, E., Popova, M., Morgavi, D. P., Martín-García, A. I., Yáñez-Ruiz, D. R., & Belanche, A. (2021). Inoculation with rumen fluid in early life accelerates the rumen microbial development and favours the weaning process in goats. Animal microbiome, 3(1), 11. https://doi.org/10.1186/s42523-021-00073-9
24. Ruiz, L., Ruas-Madiedo, P., Gueimonde, M., de Los Reyes-Gavilán, C. G., Margolles, A., & Sánchez, B. (2011). How do bifidobacteria counteract environmental challenges? Mechanisms involved and physiological consequences. Genes & nutrition, 6(3), 307–318. https://doi.org/10.1007/s12263-010-0207-5
25. Rybachuk, Z., Shkromada, O., Predko, A., & Dudchenko, Y. (2020). Influence of probiotics “Immunobacterin-D” on biocenoses and development of the gastrointestinal tract of calves. Scientific Messenger of LNU of Veterinary Medicine and Biotechnologies. Series: Veterinary Sciences, 22(98), 22-27. https://doi.org/10.32718/nvlvet9804
26. Shkromada O., Skliar O., Paliy A., Ulko L., Gerun I., Naumenko О., Ishchenko K., Kysterna O., Musiienko O., Paliy A., 2019. Development of measures to improve milk quality and safety during production. Eastern-European Journal of Enterprise Technologies, 3/11(99), 30-39. https://doi.org/10.15587/1729-4061.2019.168762
27. Shkromada, O., Skliar, O., Pikhtirova, A. & Inessa, G. (2019). Pathogens Transmission and Cytological Composition of Cow’s Milk . Acta Veterinaria Eurasia, 45 (3) , 73-79 . https://dergipark.org.tr/tr/pub/actavet/issue/50595/608279
28. Tanih, N. F., Sekwadi, E., Ndip, R. N., & Bessong, P. O. (2015). Detection of pathogenic Escherichia coli and Staphylococcus aureus from cattle and pigs slaughtered in abattoirs in Vhembe District, South Africa. TheScientificWorldJournal, 2015, 195972. https://doi.org/10.1155/2015/195972
29. Wu, H.J., Sun, L.B., Li, C.B., Li, Z.Z., Zhang, Z., Wen, X.B., Hu, Z., Zhang, Y.L. (2014) Enhancement of the immune response and protection against Vibrio parahaemolyticus by indigenous probiotic Bacillus strains in mud crab (Scylla paramamosain). Fish Shellfish Immunol 41, 156– 162. https://doi.org/10.1016/j.fsi.2014.08.027
30. Yáñez-Ruiz, D. R., Abecia, L., & Newbold, C. J. (2015). Manipulating rumen microbiome and fermentation through interventions during early life: a review. Frontiers in microbiology, 6, 1133. https://doi.org/10.3389/fmicb.2015.01133
Published
2021-12-27
How to Cite
Shkromada, O. I., & Dudchenko, Y. A. (2021). STUDY OF ANTIMICROBIAL ACTIVITY OF PROBIOTIC STRAINS OF BACILLUS. Bulletin of Sumy National Agrarian University. The Series: Veterinary Medicine, (4 (55), 38-43. https://doi.org/10.32845/bsnau.vet.2021.4.6
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