Work

Morphology of Candida albicans Exposed to Electric Current Treatments and UV Radiation in Sabouraud Broth Analyzed with Scanning Electron Microscopy

Year: 2024

Type: article

Abstract: Antimicrobial resistance has become a global treat that is under constant observation by the WHO [1]. A counter-strategy to address this important problem is the environmental cleaning and disinfection, [2] currently, there is a wide variety of disinfectant chemical products (alcohols, chlorines, hydrogen peroxide, etc.) and physical processes (UV-C light, heat) of high efficiency; however, research is being performed in order to improve disinfection rates in the hospital environment, especially against multi-drug resistant pathogens. However, microorganisms develop resistance against antimicrobial compounds [3,4], and the estimated surface contamination is reported in values of 5 to 30% despite the application of appropriate cleaning and disinfection protocols. The ineffectiveness of existing cleaning and disinfection formulations can be attributed to the presence of dry biofilm, where the survival of vegetative bacteria has been demonstrated for long periods [5]. This problem requires of new disinfection methods to be tested to help control the appearance of drug resistant microorganisms. Electrical current is a disinfection method that is being tested against microorganisms in static fluids and other scenarios that simulate real life situations [6]; according to results on recent reports, it’s a promising method that needs to be further studied in order to analyze the effects in cellular metabolism and structural components. In this research work the effect of electrical current, alone or in combination with UV-C light, was tested on Candida albicans, a fungus of great interest due to the high rates of systemic infections and mortality it causes in immunocompromised patients [7]. Aliquots of 2mL of 1.5x108CFU/mL C. albicans ATCC 10231 in Sabouraud Dextrose broth were exposed to 5 minutes of UV radiation (λ254nm) in a EsteriUV disinfection chamber (American Medical Systems), to electrical current of 50 volts (PowerEase® 90W), or the combination of both. After the treatments, 1mL of each aliquot was taken and centrifuged at 14,000g for 10 minutes to obtain a cell pellet, in which 1mL of 2.5% glutaraldehyde was added; samples were left to rest for 1 hour after homogenizing them. Later, samples were centrifuged again to remove the glutaraldehyde by decanting the samples, two washes were performed with 1x PBS at 4,000 RPM for 5 minutes. The next step consisted in dehydration of the samples by exposing them for 15 minutes to each of the ethanol solutions of the following concentrations: 75%, 85%, 95% and absolute; centrifuging (4,000 RPM for 5 minutes) and decanting before applying the next solution. Once dehydration was completed, the samples were resuspended in absolute ethanol, poured onto sterile filter paper, folded to prevent sample loss, and left to dry in a vacuum chamber (JEOL EMDSC-U10A) for 24 hours. Samples were then transferred to a sample holder and immobilized on a copper adhesive tape. The characterization was carried out using scanning electron microscopy (SEM) using a JEOL model JSM-7401f equipment at the Advanced Materials Research Center (CIMAV). The images obtained by the SEM equipment show the different morphological changes caused to the cells of Candida albicans after the treatments. Fig. 1A consist of the control sample C. albicans not exposed to either treatment, therefore normal morphology can be appreciated. Cells exposed to a electrical current (50V) are shoed in Fig.1B, minor changes in morphology are observed; some cells show a collapse in their walls. Fig.1C consists in yeast cells exposed to UV-C showed irregular yeast shapes and cellular collapse as well as a higher number of pseudohyphae. Fig.1D showed the same characteristics as Fig.1B and 1C, but also the breaking of cellular walls and cellular debris are present in the sample. A) Morphology of C. albicans, control sample. B) Sample of C. albicans exposed to 5 minutes of 50V current. C) Samples irradiated with UV-C for 5 minutes. D) C. albicans exposed to 5 minutes of UV-C and 50V of electrical current.

Source: Microscopy and Microanalysis

DOI: 10.1093/mam/ozae044.401