Due to the increase in life expectancy, new treatments have emerged which, although palliative, provide individuals with a better quality of life. Artificial saliva is a solution that contains substances that moisten a dry mouth, thus mimicking the role of saliva in lubricating the oral cavity and controlling the existing normal oral microbiota. This study aimed to assess the influence of commercially available artificial saliva on biofilm formation by Candida albicans. Artificial saliva I consists of carboxymethylcellulose, while artificial saliva II is composed of glucose oxidase, lactoferrin, lysozyme and lactoperoxidase. A control group used sterile distilled water. Microorganisms from the oral cavity were transferred to Sabouraud Dextrose Agar and incubated at 37 °C for 24 hours. Colonies of Candida albicans were suspended in a sterile solution of NaCl 0.9%, and standardisation of the suspension to 10 6 cells/mL was achieved. The acrylic discs, immersed in artificial saliva and sterile distilled water, were placed in a 24-well plate containing 2 mL of Sabouraud Dextrose Broth plus 5% sucrose and 0.1 mL aliquot of the Candida albicans suspension. The plates were incubated at 37 °C for 5 days, the discs were washed in 2 mL of 0.9% NaCl and placed into a tube containing 10 mL of 0.9% NaCl. After decimal dilutions, aliquots of 0.1 mL were seeded on Sabouraud Dextrose Agar and incubated at 37 °C for 48 hours. Counts were reported as CFU/mL (Log10). A statistically significant reduction of 29.89% (1.45 CFU/mL) of Candida albicans was observed in saliva I when compared to saliva II (p = 0.002, considering p ≤ 0.05).
The resident microbiota in the human body, such as the oral cavity, gastrointestinal tract and genitourinary tract, is able to provide resistance to disease. However, imbalances in the microbial components can promote the growth of opportunistic microorganisms, such as yeasts of genus Candida. Fungal infections present as a major cause of infectious diseases and the microorganisms of genus Candida are the most frequently isolated pathogenic fungi in human fungal infections. Bacillus spp. and Lactobacillus spp. are bacteria that have probiotic effects used in commercially available products and in studies that aim for the development of probiotics able to inhibit the microbial pathogenicity and restore the balance of resident microbiota. Thus, with increasing fungus resistance to the use of antifungal agents, which are capable of causing serious side effects to the host organism unable to destroy the target microorganism, it becomes important to develop therapeutic and/or prophylactic alternatives that have a different and an effective mechanism of action with capacity to combat fungal infections without harming the patient. Probiotic bacteria provide an alternative strategy for the prevention and treatment of candidiasis and other infectious diseases.
The increase in survival and resistance of microorganisms organized in biofilms demonstrates the need for new studies to develop therapies able to break this barrier, such as photodynamic therapy, which is characterized as an alternative, effective, and non-invasive treatment. The objective was to evaluate in vitro the effect of antimicrobial photodynamic therapy on heterotypic biofilms of Candida albicans and Bacillus atrophaeus using rose bengal (12.5 μM) and light-emitting diode (LED) (532 nm and 16.2 J). We used standard strains of B. atrophaeus (ATCC 9372) and C. albicans (ATCC 18804). The biofilm was formed in the bottom of the plate for 48 h. For the photodynamic therapy (PDT) experimental groups, we added 100 μL of rose bengal with LED (P+L+), 100 μL of rose bengal without LED (P+L-), 100 μL of NaCl 0.9 % solution with LED (P-L+), and a control group without photosensitizer or LED (P-L-). The plates remained in agitation for 5 min (pre-irradiation) and were irradiated with LED for 3 min, and the biofilm was detached using an ultrasonic homogenizer for 30 s. Serial dilutions were plated in BHI agar and HiChrom agar and incubated at 37 °C/48 h. There was a reduction of 33.92 and 29.31 % of colony-forming units per milliliter (CFU/mL) for C. albicans and B. atrophaeus, respectively, from the control group to the group subjected to PDT. However, statistically significant differences were not observed among the P+L+, P+L-, P-L+, and P-L- groups. These results suggest that antimicrobial photodynamic therapy using rose bengal (12.5 μM) with a pre-irradiation period of 5 min and LED for 3 min was not enough to cause a significant reduction in the heterotypic biofilms of C. albicans and B. atrophaeus.
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