Harmasheva I. Lactic acid bacteria of traditional fermented products, their occurrence, biological activity and practical use

Українська версія

Thesis for the degree of Doctor of Science (DSc)

State registration number

0525U000444

Thesis Registration Form

0525U000444.pdf

Applicant for

Specialization

  • 03.00.07 - Мікробіологія

24-09-2025

Specialized Academic Board

Д 26. 233. 01

Institute of Microbiology and Virology named after D. K. Zabolotny of the National Academy of Sciences of Ukraine

Essay

The dissertation is devoted to a comprehensive study of lactic acid bacteria (LAB) in traditional Ukrainian fermented foods of animal and plant origin obtained by spontaneous fermentation, based on which a scientifically based concept has been formulated regarding the dependence of the qualitative and quantitative composition of LAB, their physiological, biochemical and chemotaxonomic properties, and biological activity on different sources of isolation. Based on new, scientifically based, comprehensive approaches, the functional properties and safety of LAB have been assessed. The role of some LAB metabolites in mediating functional activity was revealed, and active LAB were selected for practical use in the food industry, agriculture and medicine. From dairy products, fermented fruits, and vegetables, 1,226 strains of LAB were isolated, which represent the genera Enterococcus, Lactococcus, Lactobacillus, Leuconostoc, and Pediococcus. Coccal forms of LAB dominated in dairy products, while representatives of the genera Lactobacillus, Leuconostoc and Pediococcus dominated in fermented products of plant origin, depending on the product. Analysis of LAB resistance to stressful cultivation conditions revealed variability in the ability to grow at temperatures of 10 and 42-45 °C, as well as resistance to 6.5% NaCl, depending on the source of isolation. For the first time, a difference in the fatty acid composition of cell lipids of L. plantarum strains was shown depending on their ability to grow under stressful conditions. For the first time, a genus-specific manifestation of the antagonistic activity of LAB against opportunistic pathogens was established. Growth inhibition after 24 hours of cultivation was mediated by the production of a complex of specific metabolites, including bacteriocins. Methodological approaches for rapid screening of potentially bacteriocinogenic LAB strains have been substantiated. Knowledge of the manifestation of antimicrobial activity depending on cultivation conditions has been deepened, and the effectiveness of using media based on food production waste for screening bacteriocinogenic LAB strains has been demonstrated. Strains of enterococci that are promising producers of bacteriocins have been selected. Exopolysaccharides (EPS) produced by LAB strains have been characterised. The role of EPS in the interaction of LAB with plant pathogens has been established. For the first time, the inhibitory or stimulating effect of LAB EPSs on the growth and biofilm formation of phytopathogenic bacteria of the species Pseudomonas fluorescens, P. syringae, Pectobacterium carotovorum, Clavibacter michiganensis, Xanthomonas campestris and X. vesicatoria was revealed. The role of LAB EPS in the antiviral activity of LAB against human herpes simplex virus type 1 (HSV-1) and tobacco mosaic virus (TMV) was demonstrated for the first time. It has been shown that LAB strains isolated from fermented products of plant and animal origin exhibit antioxidant activity in vitro. EPS at a concentration of 5 mg/ml exhibits antioxidant activity by binding hydroxyl radicals, which reaches 50% of the activity of vitamin C. LAB and their EPS (including capsular) are promising for use as reducing agents and stabilisers in nanobiotechnology for the ‘green’ synthesis of silver nanoparticles with antimicrobial activity against opportunistic pathogens. The ability of LAB to reduce silver ions with the formation of nanoparticles was strain-specific. Enterococci in traditional dairy products of spontaneous fermentation are represented by the species E. faecalis (66% of strains), E. durans (19% of strains) and E. faecium (6.5% of strains). Analysis according to safety criteria for production cultures showed that a significant number of Enterococcus strains do not meet the established requirements for industrial cultures, although they exhibit satisfactory technological properties. In 57% of enterococcus strains, multiple resistance to antibacterial drugs with different mechanisms of action was detected, biogenic amines were produced, and 20% of enterococcus strains had β-haemolytic activity. Lactococci exhibited a number of strain-specific technological characteristics, such as aroma formation, autolytic activity, moisture retention capacity, growth in the temperature range of 10-42°C and resistance to 6.5% NaCl, which meet the requirements for LAB starter cultures for dairy products. The acid formation of L. plantarum strains when growing in nutrient media based on vegetable raw materials is influenced to a greater extent by the composition of the medium than by the concentration of NaCl. The bacterial preparation LactoCap has been developed for the fermentation of vegetable raw materials, which includes the strains L. plantarum IMV B-7565, IMV B-7566, IMV B-7597 and IMV B-7598. The use of LactoCap reduces fermentation time and improves the microbiological quality of the final product.

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Research papers

1. Гармашева ІЛ, Коваленко НК, Олещенко ЛТ, Василюк ОМ. Властивості штамів лактококів, ізольованих із традиційних кисломолочних продуктів. Мікробіол журн. 2017;79(6):3-12.

2. Гармашева ІЛ, Коваленко НК, Василюк ОМ, Олещенко ЛТ. Продукція езкополісахаридів штамами молочнокислих бактерій, ізольованих з ферментованих продуктів. Мікробіол біотехнол. 2017;4:76-84.

3. Гармашева ІЛ, Коваленко НК, Олещенко ЛТ. Стійкість до антибіотиків, декарбоксилазна та гемолітична активності ентерококів, ізольованих із традиційних кисломолочних продуктів. Мікробіол журн. 2018;80(1):3-14.

4. Garmasheva IL, Kovalenko NK, Oleschenko LT. Biological traits of enterococci and lactococci isolated from traditional dairy products of Ukraine. Mikrobiol Z. 2020;82(1):22-32.

5. Garmasheva IL, Vasyliuk OM and Oleschenko LT. Effect of cocultivation on Lactobacillus plantarum strains growth and antagonistic activity. Mikrobiol Z. 2021;83(1):12-20.

6. Garmasheva I, Vasyliuk O, Kovalenko N, Oleschenko L, Ostapchuk A. Intraspecies cellular fatty acids heterogeneity of Lactobacillus plantarum strains isolated from fermented foods in Ukraine. Lett Appl Microbiol. 2015;61:283-292.

7. Garmasheva I. Isolation and characterization of lactic acid bacteria from Ukrainian traditional dairy products. AIMS Microbiol. 2016;2(3):372-387.

8. Livinska O, Ivaschenko O, Garmasheva I, Kovalenko N. The screening of lactic acid bacteria with antioxidant properties. AIMS Microbiol. 2016;2(4): 447-459.

9. Garmasheva I, Kovalenko N, Voychuk S, Ostapchuk A, Livins’ka O, Oleschenko L. Lactobacillus species mediated synthesis of silver nanoparticles and their antibacterial activity against opportunistic pathogens in vitro. BioImpacts. 2016;6(4):219-223.

10. Garmasheva I, Vasyliuk O, Kovalenko N, Oleschenko L. New approach for fast screening of lactic acid bacteria for vegetable fermentation. J Microbiol Biotechnol Food Sci. 2019;8(4):1066-1071.

11. Naumenko K, Biliavska L, Pankivska Y, Povnitsa O, Vasyliuk O, Garmasheva I, Zagorodnya S. Anti-herpetic activity exopolysaccharides produced by different species of lactic acid bacteria. J Adv Biol. 2019;12:2307-2315.

12. Garmasheva IL, Oleschenko LT. A comparative study of antagonistic activity spectra of lactic acid bacteria isolated from fermented foods. Lett Appl Microbiol. 2022;75(4):991-999.

13. Garmasheva IL, Oleschenko LT. Screening of bacteriocin-producing dairy Enterococcus strains using low-cost culture media. Front Microbiol. 2023;14:1168835.

14. Garmasheva I, Tomila T, Kharkhota M, Oleschenko L. Exopolysaccharides of lactic acid bacteria as protective agents against bacterial and viral plant pathogen. Int J Biol Macromol. 2024;276(1):133851

15. Гармашева ІЛ, Коваленко НК, Олещенко ЛТ. Молочнокислі бактерії традиційних кисломолочних продуктів України та їх біологічна активність. В: Матеріали ХІІІ з’їзду Товариства мікробіологів України ім. С.М. Виноградського; 1-6 жовтня 2013: Ялта, Україна. Ялта, 2013. с. 76.

16. Livinska OP, Ivaschenko OY, Garmasheva IL, Kovalenko NK. Isolation and screening of strains of lactic acid bacteria with antioxidative properties. In: Abstract book of the II International Scientific Conference “Microbiology and immunology – the development outlook in the 21th century”; April 14-15, 2016: Kyiv, Ukraine. Kyiv, 2016a. p. 46.

17. Василюк ОМ, Гармашева ІЛ, Коваленко НК, Олещенко ЛТ. Біологічно-активні штами Lactobacillus plantarum IМВ В-7565 та В-7566 для ферментування овочевої сировини. В: Тези доповідей XV з'їзду Товариства мікробіологів України iм. С.М. Виноградського; 11-15 вересня 2017: Одеса, Україна. Одеса, 2017. с. 37.

18. Гармашева ІЛ, Коваленко НК, Василюк ОМ, Олещенко ЛТ. Ідентифікація та біологічна активність ентерококів традиційних ферментованих продуктів. В: Матеріали XV з'їзду Товариства мікробіологів України iм. С.М. Виноградського; 11-15 вересня 2017: Одеса, Україна. Одеса, 2017б. с. 43.

19. Garmasheva IL. Comparative characteristic of lactic acid bacteria properties isolated from traditional fermented foods. In: Abstracts book III International Scientific Conference Microbiology and Immunology – the development outlook in the 21st century; April 19-20 2018: Kyiv, Ukraine. Kyiv, 2018a. p. 6-7.

20. Biliavska L, Pankivska Y, Povnitsa O., Vasuliuk O, Garmasheva I, Zagorodnya S. Antiviral potential of exopolysaccharides produced by lactic acid bacteria. In: Abstract book International Scientific Conference on Microbial Biotechnology (4th edition); 11–12 october, 2018: Chisinau, Moldova. Chisinau, 2018. p. 62.

21. Biliavska L, Pankivska Y, Povnitsa O, Vasyliuk O, Naumenko K, Garmasheva I, Zagorodnya S. Antiviral activity of the exopolysaccharides produced by Leuconostoc sp. against HSV-1. 4th International electronic conference on medical chemistry 1-30 November 2018.

22. Василюк ОМ, Гармашева ІЛ, Коваленко НК, Олещенко ЛТ. Інгібуюча активність штамів молочнокислих бактерій щодо фітопатогенних бактерій. В: Abstract book International scientific and practical conference “Prospects for the development of natural sciences in EU countries and Ukraine”; December 21-22, 2018: Wloclawek, Poland. Wloclawek, 2018. p. 10-11.

23. Vasyliuk O, Garmasheva I. Investigation of antibacterial metabolites of lactic acid bacteria against phytopathogenic bacteria. In: Conference materials of the young scientists conference “Youth and modern problems of microbiology and virology”; 12-14 November 2019: Kyiv, Ukraine. Kyiv, 2019. p. 32.

24. Василюк ОМ, Гармашева ІЛ. Антагоністична активнісь молочнокислих бактерій щодо збудників бактеріальних хвороб рослин. В: Матеріали XVІ наукової Гармашева ІЛ, Савчук ЯІ. Роль молочнокислих бактерій у підвищенні безпеки харчових продуктів. В: Матеріали V

25. Василюк ОМ, Гармашева ІЛ. Антагоністична активнісь молочнокислих бактерій щодо збудників бактеріальних хвороб рослин. В: Матеріали XVІ наукової конференції молодих учених «Мікробіологія в сучасному сільськогосподарському виробництві; 25 жовтня 2023:Чернігів, Україна. Чернігів, 2023. с. 12-13.

26. Пат. на корисну модель UA 113580 U Штам Lactobacillus plantarum 1047к з пробіотичними властивостями для заквашування овочевої сировини / Гармашева ІЛ, Василюк ОМ, Коваленко НК, Підгорський ВС, Олещенко ЛТ. Опубл. 10.02.2017. Бюл. № 3/2017.

27. Пат. на корисну модель UA 113581 U Штам Lactobacillus plantarum 47 СМ з антиоксидантною активністю для отримання ферментованих овочевих продуктів функціонального харчування / Гармашева ІЛ, Василюк ОМ, Коваленко НК, Підгорський ВС, Олещенко ЛТ, Лівінська ОП. Опубл. 10.02.2017. Бюл. №3/2017.

28. Пат. на корисну модель UA 140653 U Спосіб ферментації капусти / Василюк ОМ, Гармашева ІЛ, Коваленко НК, Олещенко ЛТ, Підгорський ВС. Опубл. 10.03.2020. Бюл. №5/2020.

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Files

autoreferat-Реферат Гармашева І-Л-.pdf

Дисертація Гармашева І-Л- 21-08-2025.pdf

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