THE EXPERIMENTAL DETERMINATION OF THE NUMBER OF FREEZING STAGES OF CANDIDA TROPICALIS FUNGAL CELLS FOR THEIR DESTRUCTION AND OBTAINING OF PROTEINS AND POLYSACCHARIDES
DOI:
https://doi.org/10.31861/biosystems2023.01.009Keywords:
Candida tropicalis, freezing, candidiasis, proteins, polysaccharides, monosaccharides, antigen, vaccineAbstract
Candidiasis is a disease that occurs due to excessive growth of the Candida fungus. Several types of this pathogen are known, each of which can cause damage to various organs and systems. Recently, there have been reports of the loss of sensitivity of Candida fungi to most of antifungal drugs that have been used for more than 40 years. An alternative to antifungal drugs for candidiasis can be the use of a vaccine for the prevention and treatment of candidal infection. In our opinion, it is promising to use a candidal vaccine based on a subunit vaccine that contains fragments of a microorganism without ballast substances. Cell fragments of Candida fungi that have antigenic properties include proteins and polysaccharides. To isolate antigenic substances from Candida fungal cells, we chose freezing as the method of destruction. The aim of this work was to experimentally substantiate the number of freezing stages for the destruction of C. tropicalis АТСС 20336 fungi cells and the subsequent production of proteins and polysaccharides. To determine the optimal number of freezing stages in the temperature range from (25 ± 2)°С to (-25 ± 2)°С, in order to destroy the cells of C. tropicalis fungi, the studies were conducted with freezing stages 3, 4, 5, and 6. Centrifugation was used to separate the excess components. Then the preliminary and sterilizing filtration was performed on membrane filters with pore diameters of 0.45 μm and 0.22 μm. The content of protein, polysaccharides and monosaccharides were determined in each case. The protein content was determined according to the State Pharmacopoeia of Ukraine (SPhU). To determine polysaccharides, a reaction with phenol and sulfuric acid was performed. Chromatographic studies of monosaccharides were conducted using paper chromatography according to the SPhU. Based on the results of the research, it was found that solutions obtained at stages 5 and 6 of freezing the biomass of C. tropicalis fungi cells at a temperature from (25 ± 2) °C to (-25 ± 2) °C contained the highest amount of proteins and polysaccharides. It is likely that with these number of stages for freezing the biomass of C. tropicalis fungi cells, active substances are released from all layers of Candida fungi cells. Thus, it can be concluded that five stages of freezing are the optimal number that ensures the maximum extraction of polysaccharides and proteins quickly and economically.
References
Da Silva L.B.R., Taborda C.P., Nosanchuk J.D. Advances in Fungal Peptide Vaccines. J Fungi (Basel). 2020; 6 (3): 119. https://doi.org/10.3390/jof6030119.
Edwards J.E.Jr., Schwartz M.M., Schmidt C.S., et al.. A fungal immunotherapeutic vaccine (ndv-3a) for treatment of recurrent vulvovaginal candidiasis-a phase 2 randomized, double-blind, placebo-controlled trial. Clin Infect Dis. 2018; 66 (12): 1928-1936. https://doi.org/10.1093/cid/ciy185.
Howley M.M., Carter T.C., Browne M.L., at al. Fluconazole use and birth defects in the National Birth Defects Prevention Study. Am J Obstet Gynecol. 2016; 214 (5): 657. https://doi.org/10.1016/j.ajog.2015.11.022.
Mercer D.K., O'Neil D.A. Innate Inspiration: Antifungal Peptides and Other Immunotherapeutics From the Host Immune Response. Front Immunol. 2020; 17 (11): 2177. https://doi.org/10.3389/fimmu.2020.02177.
Nami S., Mohammadi R.., Vakili M. at al. Fungal vaccines, mechanism of actions and immunology: A comprehensive review. Biomedicine & Pharmacotherapy. 2019; 109: 333-344. https://doi.org/10.1016/j.biopha.2018.10.075.
Piccione D., Mirabelli S., Minto, N., Bouklas T. Difficult but not impossible: in search of an anti-Candida vaccine. Current Tropical Medicine Reports. 2019; 6: 42–49. https://doi.org/10.1007/s40475-019-00173-2.
Polesello V., Segat L.,Crovella S., Zupin L. Candida infections and human defensins. Protein Pept. Lett. 2017; 24 (8): 747-756. https://doi.org/10.2174/0929866524666170807125245.
Tarang S., Kesherwani V., LaTendresse B. at al. In silico design of a multivalent vaccine against Candida albicans. Scientific Reports. 2020; 1066 (2020): https://doi.org/10.1038/s41598-020-57906-x.
Tso G.H.W., Reales-Calderon J.A., Pavelka N. The elusive anti-Candida vaccine: lessons from the past and opportunities for the future. Front Immunol. 2018; 9: 897. https://doi.org/10.3389/fimmu.2018.00897. eCollection 2018.
Wang X., Sui X., Yan L. at al. Vaccines in the treatment of invasive candidiasis. Virulence. 2015; 6 (4): 309-315. https://doi.org/10.4161/21505594.2014.983015
