Helium Cold Atmospheric Plasma Causes Morphological and Biochemical Alterations in Candida albicans Cells.

(1) Background: Previous studies reported the promising inhibitory effect of cold atmospheric plasma (CAP) on Candida albicans. However, the exact mechanisms of CAP's action on the fungal cell are still poorly understood. This study aims to elucidate the CAP effect on C. albicans cell wall, by evaluating the alterations on its structure and biochemical composition; (2) Methods: C. albicans cells treated with Helium-CAP were analyzed by atomic force microscopy (AFM) and Fourier transform infrared spectroscopy (FTIR) in order to detect morphological, topographic and biochemical changes in the fungal cell wall. Cells treated with caspofungin were also analyzed for comparative purposes; (3) Results: Expressive morphological and topographic changes, such as increased roughness and shape modification, were observed in the cells after CAP exposure. The alterations detected were similar to those observed after the treatment with caspofungin. The main biochemical changes occurred in polysaccharides content, and an overall decrease in glucans and an increase in chitin synthesis were detected; (4) Conclusions: Helium-CAP caused morphological and topographic alterations in C. albicans cells and affected the cell wall polysaccharide content.

Effect of Cold Atmospheric Plasma Jet Associated to Polyene Antifungals on Candida albicans Biofilms.

The increasing incidence of antifungal resistance represents a great challenge in the medical area and, for this reason, new therapeutic alternatives for the treatment of fungal infections are urgently required. Cold atmospheric plasma (CAP) has been proposed as a promising alternative technique for the treatment of superficial candidiasis, with inhibitory effect both in vitro and in vivo. However, little is known on the association of CAP with conventional antifungals. The aim of this study was to evaluate the effects of the association between CAP and conventional polyene antifungals on Candida albicans biofilms. C. albicans SC 5314 and a clinical isolate were used to grow 24 or 48 h biofilms, under standardized conditions. After that, the biofilms were exposed to nystatin, amphotericin B and CAP, separately or in combination. Different concentrations of the antifungals and sequences of treatment were evaluated to establish the most effective protocol. Biofilms viability after the treatments was compared to negative control. Data were compared by One-way ANOVA and post hoc Tukey (5%). The results demonstrate that 5 min exposure to CAP showed more effective antifungal effect on biofilms when compared to nystatin and amphotericin B. Additionally, it was detected that CAP showed similar (but smaller in magnitude) effects when applied in association with nystatin and amphotericin B at 40 µg/mL and 60 µg/mL. Therefore, it can be concluded that the application of CAP alone was more effective against C. albicans biofilms than in combination with conventional polyene antifungal agents.

Cold Atmospheric Plasma Jet as a Possible Adjuvant Therapy for Periodontal Disease.

Due to the limitations of traditional periodontal therapies, and reported cold atmospheric plasma anti-inflammatory/antimicrobial activities, plasma could be an adjuvant therapy to periodontitis. Porphyromonas gingivalis was grown in blood agar. Standardized suspensions were plated on blood agar and plasma-treated for planktonic growth. For biofilm, dual-species Streptococcus gordonii + P. gingivalis biofilm grew for 48 h and then was plasma-treated. XTT assay and CFU counting were performed. Cytotoxicity was accessed immediately or after 24 h. Plasma was applied for 1, 3, 5 or 7 min. In vivo: Thirty C57BI/6 mice were subject to experimental periodontitis for 11 days. Immediately after ligature removal, animals were plasma-treated for 5 min once-Group P1 (n = 10); twice (Day 11 and 13)-Group P2 (n = 10); or not treated-Group S (n = 10). Mice were euthanized on day 15. Histological and microtomography analyses were performed. Significance level was 5%. Halo diameter increased proportionally to time of exposure contrary to CFU/mL counting. Mean/SD of fibroblasts viability did not vary among the groups. Plasma was able to inhibit P. gingivalis in planktonic culture and biofilm in a cell-safe manner. Moreover, plasma treatment in vivo, for 5 min, tends to improve periodontal tissue recovery, proportionally to the number of plasma applications.

Cold Atmospheric Pressure Plasma Jet Reduces Trichophyton rubrum Adherence and Infection Capacity.

This study aimed to evaluate the effects of cold atmospheric pressure plasma (CAPP) jet on Trichophyton rubrum growth, germination and adherence to nail. The effects of plasma jet on T. rubrum conidia germination and on mycelial growth were evaluated by in vitro assays. An ex vivo nail infection model was used to evaluate the effects on conidia adherence and infection. Biochemical analyses of nail fragments exposed or not to CAPP were performed by attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy. Plasma jet exposure for 10 and 15 min completely inhibited mycelial growth after only one exposure. Fifteen minutes of exposure could reduce conidia germination in suspension. Fungal suspensions exposed to plasma jet for 10 and 15 min were not able to infect nail specimens. These results were corroborated by ATR-FTIR analyses of nail fragments. In conclusion, single exposure to CAPP for 15 min was able to inhibit fungal growth, adherence and infection capacity. The results suggest that cold atmospheric plasma jet can be a promising alternative for the treatment of onychomycoses caused by T. rubrum.

Amplitude-modulated cold atmospheric pressure plasma jet for treatment of oral candidiasis: In vivo study.

The aim of this study was to establish an effective and safe protocol for in vivo oral candidiasis treatment with atmospheric plasma jets. A novel amplitude-modulated cold atmospheric pressure plasma jet (AM-CAPPJ) device, operating with Helium, was tested. In vitro assays with Candida albicans biofilms and Vero cells were performed in order to determine the effective parameters with low cytotoxicity. After the determination of such parameters, the protocol was evaluated in experimentally induced oral candidiasis in mice. AM-CAPPJ could significantly reduce the viability of C. albicans biofilms after 5 minutes of plasma exposure when compared to the non-exposed group (p = 0.0033). After this period of exposure, high viability of Vero cells was maintained (86.33 ± 10.45%). Also, no late effects on these cells were observed after 24 and 48 hours (83.24±15.23% and 88.96±18.65%, respectively). Histological analyses revealed significantly lower occurrence of inflammatory alterations in the AM-CAPPJ group when compared to non-treated and nystatin-treated groups (p < 0.0001). Although no significant differences among the values of CFU/tongue were observed among the non-treated group and the groups treated with AM-CAPPJ or nystatin (p = 0.3201), histological analyses revealed marked reduction in candidal tissue invasion. In conclusion, these results point out to a clinical applicability of this protocol, due to the simultaneous anti-inflammatory and inhibitory effects of AM-CAPPJ with low cytotoxicity.