Essential oil and plant extract of oregano as agents influencing the virulence factors of Candida albicans.

Candida albicans, a polymorphic yeast, is a physiological component of the human and animal commensal microbiome. It is an etiological factor of candidiasis, which is treated by azole antifungals. Growing resistance to azoles is a reason to look for other alternative treatment options. The pharmacotherapeutic use of plant extracts and essential oils has become increasingly important. In our experiment, C. albicans showed susceptibility to four observed plant extracts and essential oils from peppermint ( Mentha piperita), thyme ( Thymus vulgaris), sage ( Salvia officinalis), and oregano ( Origanum vulgare). Oregano plant extract and essential oil showed the highest antifungal activity, at MIC values of 4.9 mg/mL and 0.4 mg/mL respectively. Therefore, it was subjected to further research on the influence of virulence factors - biofilm formation, extracellular phospholipase production and germ tube formation. Oregano plant extract and essential oil showed an inhibitory effect on the observed C. albicans virulence factors at relatively low concentrations. The extract inhibited the adherence of cells at MIC 12.5 mg/mL and essential oil at MIC 0.25 mg/mL. Degradation of the formed biofilm was detected at MIC 14.1 mg/mL for plant extract and at MIC 0.4 mg/mL for essential oil. Extracellular phospholipase production was most effectively inhibited by the essential oil. In particular, the number of isolates with intensive extracellular phospholipase production decreased significantly. Of the 12 isolates intensively producing extracellular phospholipase, only 1 isolate (4.5%) retained intense production. Essential oil caused up to a 100 % reduction in germ tubes formation and plant extract reduced their formation depending on the concentration as follows: 2.6% (0.8 mg/mL), 21.2 % (6.25 mg/mL), and 64.5 % (12.5 mg/mL) compared to the control.

Efficacy of manuka honey with conventional antifungals on Malassezia pachydermatis.

Yeast infections such as otitis externa and seborrheic dermatitis in dogs and cats are frequently associated with Malassezia pachydermatis secondary infection. It is part of the normal cutaneous microflora of most warm-blooded vertebrates, however, under certain conditions, it can become a causative agent of infection that needs to be treated pharmacologically. Azole derivatives are the drugs of the first choice. An interesting trend in developing resistance is the use of natural substances, which include manuka honey with confirmed antimicrobial properties. The main intention of this research was to evaluate the mutual effect of manuka honey in combination with four conventional azole antifungals - clotrimazole, fluconazole, itraconazole, and miconazole - on 14 Malassezia pachydermatis isolates obtained from dogs and 1 reference strain. A slightly modified M27-A3 method (CLSI 2008) and the checkerboard test (Nikolic et al. 2017) were used for this purpose. Our results show an additive effect of all 4 antifungals with manuka honey concurrent use. Based on the determined values of fractional inhibitory concentration index (FICI - 0.74±0.03 when manuka honey combined with clotrimazole, 0.96±0.08 with fluconazole, 1.0±0 with miconazole and 1.16±0.26 with itraconazole), it was found in all cases that the effect of substances used is more pronounced in mutual combination than when used separately.

Efficacy of Lamiaceae essential oils with selected azoles against Candida albicans clinical isolates.

A current problem in candidiasis treatment is increasing resistance to azoles, which are often prescribed to patients. The study underlines the high resistance of yeasts to fluconazole, which achieved high MIC (minimal inhibitory concentration) values both alone and in combination with essential oils (EOs). Antifungal activity of Hyssopus officinalis, Thymus vulgaris, Salvia officinalis and Rosmarinus officinalis EOs was determined against 13 clinical isolates of Candida albicans and reference strain Candida albicans ATCC 10231. The synergistic effect was investigated for the combination of itraconazole and fluconazole with Hyssopus officinalis and Thymus vulgaris EOs. Based on the fractional inhibitory concentration index, the synergistic effect was achieved in all of the samples exposed to itraconazole with Hyssopus officinalis (FICI 0.3±0.06). On the other side, the additive effect was proven in use of itraconazole with Thymus vulgaris (FICI 0.75±0.35) and fluconazole with both EOs tested (FICI 0.81±0.19; 0.88±0.57) This study shows the importance of monitoring the synergistic effect of antifungals combined with EOs, because it is a possible solution for reducing the resistance and improving the disease prognosis.

Interspecific differences in cryoresistance of lichen symbiotic algae of genus Trebouxia assessed by cell viability and chlorophyll fluorescence.

Unicellular algae of genus Trebouxia are the most frequent symbiotic photobionts found in lichen species adapted to extreme environments. When lichenised, they cope well with freezing temperature of polar regions, high-mountains environments and were successfully tested in open-space experiments. Trebouxia sp. is considered potential model species for exobiological experiments. The aim of this paper is to evaluate cryoresistence of Trebouxia sp. when isolated from lichen thalli and cultivated on media. In our study, six algal strains were exposed to repeated freezing/thawing cycles. The strains of Trebouxia sp. (freshly isolated from lichen Lasallia pustulata), Trebouxia erici, Trebouxia asymmetrica, Trebouxia glomerata, Trebouxia irregularis, and Trebouxia jamesii from culture collection were cooled from 25 to -40 °C at two different rates. The strains were also shock frozen in liquid nitrogen. After repeated treatment, the strains were inoculated and cultivated on a BBM agar for 7 days. Then, cell viability was assessed as relative share of living cells. Potential quantum yield of photochemical reactions in PS II (F(V)/F(M)), and effective quantum yield of photochemical reactions in PS II (Φ(PSII)) were measured. While the slow cooling rate (0.5 °C min(-1)) did not cause any change in viability, F(V)/F(M), and Φ(PSII), the fast cooling rate (6.0 °C min(-1)) caused species-specific decrease in all parameters. The most pronounced interspecific differences in cryoresistance were found after shock freezing and consequent cultivation. While T. asymmetrica and T. jamesii exhibited low viability of living cells (18.9% and 34.7%) and full suppression of photosynthetic processes, the other strains had viability over 60%, and unaffected values of F(V)/F(M), and Φ(PSII). This indicated a high degree of cryoresistance of T. glomerata, T. erici, T. irregularis and Trebouxia sp. strains. These strains could be used for detailed investigation of underlying physiological mechanisms and as models for astrobiological tests taken in the Earth facilities.

Induction of micronuclei in rat bone marrow after chronic exposure to lead acetate trihydrate.

Lead increasingly contributes to pollution of the environment and may play a role in the development of adverse effects in the human and animal body. Data concerning its mutagenic, clastogenic, and carcinogenic properties have been conflicting. In this study, we evaluated the frequency of micronuclei in bone marrow erythrocytes of rats treated with lead acetate trihydrate. Outbred Wistar rats were exposed to a daily dose of 100 mg/L drinking water for 125 days. The mean value of the total number of micronuclei observed in polychromatic erythrocytes of female rats was significantly higher than that found in the control group (13.375 +/- 2.722 against 9.625 +/- 3.204 micronuclei/1000 cells; P = 0.024 in ANOVA). In exposed female animals, no significant reduction of the ratio of polychromatic to normochromatic erythrocytes was observed (0.990 +/- 0.228 against 1.208 +/- 0.195; P = 0.060 in ANOVA). The effects of lead acetate trihydrate in male rats are both cytotoxic and genotoxic because of a decrease in ratio of polychromatic to normochromatic erythrocytes (0.715 +/- 0.431 against 1.343 +/- 0.306; P = 0.023, ANOVA followed by Tukey test) and an increase in frequency of micronucleated polychromatic erythrocytes (24.167 +/- 7.859 against 4.0 +/- 4.528 micronuclei/1000 cells; P < or = 0.001, ANOVA followed by Tukey test), respectively.