Design and safety of phytobiotic with beneficial vaginal lactobacilli for the prevention of urogenital tract infections

MARCHESI ANTONELLA ; SILVA JESSICA ALEJANDRA; DE GREGORIO PRISCILLA ROMINA; WIESE BIRGITT; VIGNOLO GRACIELA; NADER-MACÍAS MARÍA ELENA FÁTIMA

Conferencia; International Scientific Conference Probiotics, Prebiotics Gut Microbiota and Health. Prague,; 2019

The vegetal extracts are applied to decrease or counteract Urogenital Tract (UGT) Infections symptoms, most of them produced as consequence of vaginal microbioma disbiosis, and host genetic polymorphism. Lactobacilli are the dominant bacteria in human vagina, exerting different type of effects both in healthy, sexually active and pregnant women. The complementation of phytoderivatives and BVL can be used to restore the ecological equilibrium of vaginal microbioma and prevent UGTI, which represent very high costs in public health systems and effect, mainly in pregnant and newborn. The objective of this work is to evaluate the compatibility between phytoderivatives and BVL and their safety to advance in the design of phytobiotic formulas for the UGT. Materials and methods: a) Compatibility between phytocompounds-BVL: different BVL (24 strains identified as Lactobacillus reuteri, L.rhamnosus, L.delbrueckii, L.johnsonii, L.salivarius, L.gasseri, L.paracasei) with pure or combined phytocompounds (Hydrocotile-asiatica, Calendula-officinalis, Peumus-boldu, Hamamelis-virginiana, Atropa-belladona, Cyclolepis-genistoides, Equisetum-arvence, Aesculus-hippocastanum, Amaranthus-muricatus, Acrtostaphylos-uva-ursi, Matricaria-recutita, Urtica-dioica, Smilax-aspera, Echinacea-purpurea, Chelidonium-majus, Plantago-lanceolata, Thymus-vulgaris, Minthostachys-mollis, Aloysia-polystachya) were incubated in polystyrene microplates at 37°C during 24h. Optical density (OD560nm) at 3, 6, 9, 12 and 24h were used to calculate the growth parameters, behavior and degree of compatibility. b) Inhibition of pathogens by phytocompounds: inhibition was determined by plate diffusion technique against urogenital pathogens: Candida albicans, C.tropicalis, Streptococcus aureus, Stp.agalactiae, Staphylococcus saprophyticus, Stap. aureus, Escherichia coli and Enterococcus faecalis. c) Safety assays: antimicrobial resistance of BVL to Ampicillin, Tetracycline, Chloramphenicol, Gentamicin, Streptomycin, Kanamycin, Clyndamicin, Vancomycin and Erythromycin was determined by microdilution technique, and MIC (minimum inhibitory concentration) in polystyrene-microplates in LSM broth, in microaerophyllic conditions at 37ºC for 48h, results defined by EFSA directions. Virulence factors were evaluated in agarized media containing specific substrates for the expression of Lecithinase, Gelatinase and Hemolysin. d) Animal assays: estrogenized mice were intravaginally (i.v.) inoculated once/day with different phytocompounds-BVL during 7 days, defining the estral cycle through vaginal washing and absence of inflammatory response by Giemsa stain and histology. Results-Discussion. Most of the phytocompounds exerted different type of effect on BVL growth. Some of them (Smilax aspera) stimulated BVL: L.ga.1263, L.ga.1509, L.ga.1320, L.rh.1261, L.rh.1508, while some other showed no effect: L.rh.1511, L.re.1327, L.re.1324, L.ga.1264. Some phytocompounds had a stimulating effect against BVL: Betula, Urtica-dioica, Hydrocotile-asiatica and Carica-papaya, while Echinacea-purpurea, Lippia, Chelidonium-majus, Hamamelis-virginiana, Acrtostaphylos-uva-ursi exerted inhibition. The extract inhibited most of the pathogens assayed. High number of BVL were sensitive to CHLOR/STREP/GEN/CLYN, frequently applied for UGTI therapy, being most of them resistant to ERY. No virulence factors were detected or expressed in BVL evaluated. Mice assays did not show inflammatory response or adverse effects after i.v.-7-daily-doses-phytobiotics formulated with selected phytoderivatives-BVL. The results support the design of safe phytobiotics containing selected BVL-phytoderivatives for the prevention UGTI.