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Table of Contents   
ORIGINAL RESEARCH  
Year : 2015  |  Volume : 26  |  Issue : 2  |  Page : 158-162
Evaluation of effect of topical ozone therapy on salivary Candidal carriage in oral candidiasis


Department of Oral Medicine and Radiology, Hitkarini Dental College and Hospital, Jabalpur, Madhya Pradesh, India

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Date of Submission26-Jan-2014
Date of Decision31-Mar-2014
Date of Acceptance17-Apr-2015
Date of Web Publication22-Jun-2015
 

   Abstract 

Background and Aim: Ozone is highly valued for various therapeutic applications such as antimicrobial, antihypoxic, analgesic, and immunostimulating for more than a century in the medical profession. Ozone therapy is now gaining a strong foothold in dentistry. Ozone has bactericidal, fungicidal, and virucidal properties. Oral candidiasis is one of the most common opportunistic fungal infections of the oral cavity. Hence, a study was conducted to evaluate and compare the ability of ozonated water and topical clotrimazole in reducing the Candidal species colony-forming unit (CFU) count in oral candidiasis.
Materials and Methods: The study included 40 candidiasis patients of either sex aged between 18 and 60 years attending the Department of Oral Medicine and Radiology. The patients were randomly assigned to either topical ozone therapy or topical clotrimazole groups. Salivary Candidal CFU counts were assessed during and after the treatments.
Results and Conclusion: There was gradual but significant reduction in Candidal CFU count in both groups. At the end of the treatment, Candidal CFU count reduction in ozone group (60.5% reduction) was more than the clotrimazole group (32.3% reduction). 14 patients (70%) with candidiasis in ozone group were reduced to 6 (30%) whereas only 8 patients (40%) out of 13 (65%) in clotrimazole group, although intergroup comparison was not statistically significant. Ozone therapy was much more effective in reducing the patients with candidiasis to a state of carriers. These findings suggest that ozonated water might be useful to treat oral candidiasis.

Keywords: Candidiasis, oral, ozone, saliva

How to cite this article:
Khatri I, Moger G, Kumar N A. Evaluation of effect of topical ozone therapy on salivary Candidal carriage in oral candidiasis. Indian J Dent Res 2015;26:158-62

How to cite this URL:
Khatri I, Moger G, Kumar N A. Evaluation of effect of topical ozone therapy on salivary Candidal carriage in oral candidiasis. Indian J Dent Res [serial online] 2015 [cited 2020 Oct 22];26:158-62. Available from: https://www.ijdr.in/text.asp?2015/26/2/158/159146
Ozone is highly valued for various effects like antimicrobial, antihypoxic, analgesic, immunostimulating etc., on biological systems. It is commonly used in the treatment of external ulcers, skin lesions, arterial circulatory disorders, immunodeficiency states, fungal infections etc., for more than a century in the medical profession. [1],[2],[3] Its simplicity of performance, good tolerance by patients, antibiotic-resistant infections, and high medico-social and economic efficiency are the factors responsible for its widespread popularity. Ozone therapy is now gaining a strong foothold in dentistry. There have been several reports on the beneficial effects of the same in various periodontic, pedodontic, oral surgery, and endodontic conditions. It's use as standard dental therapy is less known, because it has mostly been used in an empirical fashion without rational basis and appropriate controls. [2]

Ozone has bactericidal, fungicidal, and virucidal properties. Oral candidiasis is one of the most common opportunistic fungal infections of the oral cavity. Rising incidences of candidiasis have been reported throughout the world in the past two decades. It can be a frequent and significant source of oral discomfort, pain, loss of taste, and aversion for food. Oral candidiasis is managed initially with topical treatment, before systemic antifungal drugs. Polyene antibiotics are the initial choice of drugs and azoles have been developed more recently. Milder episodes respond to topical therapy with nystatin, clotrimazole or ketoconazole. Fluconazole and clotrimazole are effective in diffuse and systemic candidiasis. Fluconazole develops resistance, especially with advanced HIV disease. The use of miconazole is limited because of potential side-effects such as vomiting and diarrhea. Ketoconazole in elderly patients is not recommended due to drug interactions and hepatotoxicity.

With the widespread use of antifungal drugs, the epidemiology of fungal pathogens has changed. These emerging fungi are characterized by resistance or lower susceptibility to standard antifungal agents. [4] Hence, in this study, an attempt was made to evaluate and compare the ability of ozonated water with topical clotrimazole in the reduction of Candidal colony-forming unit (CFU) in oral candidiasis cases.

Aim

To evaluate the effect of topical ozone therapy on the salivary Candidal carriage in candidiasis patients.

Objectives

  • To evaluate salivary Candidal count before and after topical ozone therapy
  • To study the effect of clotrimazole in the salivary Candidal count before and after topical application
  • To compare the efficacy of topical ozone therapy and topical clotrimazole in salivary Candidal carriage count after the treatment.



   Materials and Methods Top


Forty candidiasis patients of either sex aged between 38 and 73 years attending the Department of Oral Medicine and Radiology were included in the study. Candidiasis was diagnosed on clinical and microbiological examinations. The study was approved by Ethical Committee of the college. Both verbal and written informed consent was taken from all subjects participated in the study.

Subjects were randomly allocated to either:

Group I or Study Group: 20 patients treated with topical ozone therapy. Each patient in this group was instructed to rinse with 5-10 mL of ozonated water for 1 min in the morning once daily for 5 days. Patient was advised to avoid drinking or eating for 15-20 min after the rinse.

Group II: 20 patients treated with topical clotrimazole. Each patient in this group was treated with candid mouth paint (Clotrimazole 1%). Patients were advised to apply the candid mouth paint to affected areas thrice daily for 2 weeks. Patients were instructed to avoid consuming water and food for at least 2 h after the application.

Patients in both groups were recalled after completion of the treatments, and mycological assessment was carried out by repeating the saliva collection.

Salivary samples of both groups before and after the treatment were inoculated using Sabouraud's dextrose agar media. [4],[5] This was then incubated for 48 h at 37°C to assess colony resembling yeast/mL of saliva (CFU). Colony resembling yeast growth was removed from the plates and processed further for identification using gram staining. Colony counting was done. The microbiological changes before, during, and after the treatments were assessed.

Patients who had received any antifungal therapy in the last 6 months, patients on any chemotherapeutic mouth rinses in the past 6 months, pregnant and lactating mothers, presence of any respiratory infection, recent myocardial infarction, hemorrhages in any organ, hyperthyroidism and thrombocytopenia, acute alcohol intoxication, and subjects having zone allergy [6] were excluded from the study.


   Results Top


The study population comprised of 46 candidiasis patients out of which only 40 patients were considered for the study. The remaining six patients were excluded because they could not be followed-up. There were 20 male and 20 female patients. They were in the age group ranging from 38 to 73 years. There were 20 patients treated with ozone therapy and 20 patients treated with clotrimazole.

Colony-forming unit count of salivary Candidal species before and after topical ozone therapy and CFU count of salivary Candidal species before and after topical clotrimazole were assessed and analyzed. Appropriate univariate and bivariate analysis were carried out using the "Student t-test" for the continuous variable (age) and "two-tailed Fisher exact test" or "Chi-square (χ2 ) test" for categorical variables. The comparison between groups, that is, according to the severity of the diseases was done using "ANOVA."

Antifungal effect of topical ozone therapy

The mean salivary Candidal species CFU count for the total 20 patients at the baseline visit in this group was 1871.85. The mean CFU count reduced gradually over 5 days of the therapy. At the end of the therapy, it was 738.95 (60.5% reduction), and the reduction was statistically highly significant (P < 0.001) [[Table I] and Graph 1].
Table 1: Comparison of antifungal activity of topical ozone therapy and topical clotrimazole after treatments


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Antifungal effect of topical clotrimazole treatment

The mean salivary Candidal species CFU count for the total 20 patients at the baseline visit in this group was 1768.20 [[Table I] and Graph 1]. The mean CFU count reduced gradually over 7-14 days of topical application of clotrimazole. At the end of the treatment, it was 1197.55 (32.3% reduction), and the reduction was statistically highly significant (P < 0.001).

Comparison of antifungal activity of ozone therapy and clotrimazole after treatments

Intergroup comparison at the baseline visit, the mean salivary Candidal species CFU count for ozone therapy group before treatment was 1871.85 and for clotrimazole group was 1768.20 [[Table I] and Graph 1]. There was a gradual reduction in mean CFU during the treatment period in both groups. At the end of therapy, 60.5% reduction in ozone group and 32.3% reduction in clotrimazole group was observed, but the difference between the groups was not statistically significant.

Comparison of Candidal colony-forming units before and after the treatments

Intergroup comparison at the baseline visit patients with salivary Candidal species CFU count above 400 in ozone group was 14 and in clotrimazole group 13 [Table 2]. After the treatment, patients with CFU count above 400 in ozone group were reduced to 6 and were statistically significant. In clotrimazole group, it was reduced to 8 and was statistically not significant. Although ozone therapy was much more effective than clotrimazole group in reducing the Candidal species count, the differences between the groups were not statistically significant.

Both topical ozone and clotrimazole mouth paint were well-tolerated. None of the patients in either group I or group II had any adverse effects of the treatments.


   Discussion Top


In the present study, "Medical Ozone Generator," Model 0023-GW-P [Figure 1] was used to prepare ozonated water. It is possible to supply a high dose of ozonated water using this apparatus. [7] Ozonated water was stored at 4°C whenever required. Both the activity and the concentration of ozone in the aqueous phase decrease faster at 22°C than 4°C, indicating that the effectiveness of ozone immediately decreases when ozonated water is stored at room temperature. [8]
Figure 1: Ozone Generator

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The advantages of ozone in the aqueous phase are its potency, ease of handling, lack of mutagenicity, rapid microbicidal effects, and suitability for use as a soaking solution for medical and dental instrument. [8],[9],[10] The bactericidal, fungicidal, and virucidal properties of ozone are the result of its intense oxidizing capacity, with the formation of free radical and direct destruction of almost all microorganisms. In addition, ozone favors tissue healing and increases blood perfusion. [11],[12]

The standard deviation was found to be high for both the groups in our study. This is due to the fact that some subjects had a relatively low whereas others had a high initial CFU count. For future research in this area, establishing a minimum number of initial CFUs as inclusion criteria probably would lower the standard deviation to a more acceptable level. [13]

Saliva collection was done between 9.00 am and 12.00 noon to avoid diurnal variations. Whole saliva collection was done by spitting method. [14]

Ozonated water was prepared by ozone generator. Ozone at a concentration of 25 μg/mL is allowed to pass in the form of minute bubbles through 1 L distilled water for 1 min 50 s and it was collected in container made exclusively of glass and well-sealed with teflon cap. By using distilled water and a high-quality "Medical Ozone Generator," Model: 0023-GW-P [7] a maximum saturation of approximately 25 μg ozone/mL at room temperature was obtained. [15]

In Group I patients [[Table I] and Graph 1], at the end of the therapy, the Candidal CFU was reduced to 60.5% which was highly significant statistically (P < 0.001). In the present study, we found that ozonated water was highly effective in killing Candidal species. Similar to our results, other investigators also observed a reduction in the presence of oral fungi and bacterial count after ozonized water treatment. [8],[10],[16]

Nagayoshi, et al. also reported the antimicrobial property of ozonated water. It was highly effective in killing Gram-positive and Gram-negative oral microorganisms. Among them, the Gram-negative bacteria were substantially more sensitive than the Gram-positive oral streptococci and Candidal albicans. Furthermore, they found that ozonated water had strong bactericidal activity against the bacteria in plaque biofilm. [8]

Kshitish and Laxman in their randomized, double-blind, crossover split-mouth design study found that the use of both ozonated water as well as chlorhexidine technique had fungicidal and bactericidal effect in periodontitis. The plaque, gingival inflammation, and bleeding reduction were higher in ozone group. The antibacterial and antifungal efficacy of ozone on  Actinobacillus actinomycetemcomitans Scientific Name Search itans and C. albicans was appreciable. [16]

However, Candidal species were not completely eliminated after exposure to 25 μg ozone/mL in the present study. This is in accordance with the study of Nagayoshi et al., who also reported incomplete eradication after exposure to 2-4 mg/L of ozonated water. [8]

De Faria, et al. in their study demonstrated that ozone exposure of Candidal isolates led to the significant yeast counts reduction that was significantly correlated to the time of exposition. The total inactivation of C. albicans type-strain occurred after 5 min of ozone exposition at the concentration of 3.3 mg/L. However, these exposition parameters were not effective for total inactivation of clinical C. albicans isolates. These results suggest that clinical strains are more resistant to ozone in relation to the type-strain. [5]

Arita, et al. found that the combination of ozonated water and ultrasonication had a strong effect on the viability of C. albicans. Almost no viable C. albicans cells were detected after the denture plates were exposed to flowing ozonated water (2-4 mg/L) at a flow rate of 2 L/min for 1 min. [17] Complete elimination of candida species in their study could be due to the higher concentration of ozone in addition to ultrasonication and prewash of the dentures with saline.

Huth, et al. in their study of effectiveness of ozone against endodontopathogenic microorganisms in root canal biofilm model revealed that the highest concentration of ozonated water (20 μg/mL) lead to a near eradication of the microorganisms in the 3-week-old biofilm and a complete elimination by gaseous ozone at a concentration of 32/g−3 for 1 min or a lower concentration (4/g−3 ) for contact times of at least 2.5 min. [18]

Clotrimazole has been extensively used for the prophylaxis and treatment of oral and vaginal candidiasis. [4],[19] It is known to be very effective locally, and only a small percentage of the drug applied to the oral mucosa can be detected in the serum or urine. [19] Clotrimazole topical preparations are generally well-tolerated, but local irritation has necessitated withdrawal of therapy in a few cases. [20]

Hence, clotrimazole was considered as a positive control when testing the efficacy of ozone. The patients in Group II were treated with candid mouth paint (clotrimazole 1%). At the end of the treatment, the Candidal CFU count was reduced to 32.3% which was statistically highly significant (P < 0.001) [[Table I] and Graph 1]. Similar to our findings Murray, et al. reported 32% mycological eradication with clotrimazole troches for the treatment of oropharyngeal candidiasis. [21] However, in the study by Sholapurkar et al. recorded 85.71% of patients with negative culture at the end of the treatment with clotrimazole mouth paint. [22]

The result of this study showed that at the end of the treatment, out of 14 patients of group I (n = 20), only six had CFU >400. But in group II, out of 13 patients only eight had CFU >400 [Table 2]. The intergroup comparison was not statistically significant. However, ozone therapy was found to be much more effective in reducing the number of patients with Candidiasis to a state of carriers. Patients with candidiasis had >400 CFU/mL of saliva, whereas carriers of C. albicans had <400 CFU/mL. Thus, quantitative cultures of saliva may aid in the diagnosis of oral candidiasis. [23]

Clotrimazole has certain side-effects such as hypersensitivity, local irritation, low compliance, and less tolerance of the drug. [24] Hence, ozone should serve as a good alternative antifungal agent for oral candidiasis as there have not been any reported harmful effects of topical ozone in the literature. [3],[19]

Considering the various beneficial effects on the biological system and low cytotoxicity of ozonated water, [3],[5] our findings may represent a promising alternative for control of C. albicans levels in vivo. However, many more long-term studies are needed to substantiate the use of ozonated water for professional and home care, particularly regarding the optimal ozone concentration, mode of use, frequency and duration of application of the same. The small sample size of the present study requires replication of these findings in a larger sample, possibly using a crossover design to minimize the effects of confounding factors and to maximize assay sensitivity for detecting difference between different treatment methods.


   Conclusion Top


The study was conducted to compare the efficacy of topical ozone therapy and clotrimazole mouth paint in the treatment of oral candidiasis. The results of our study should provide a guideline for further researches as present findings suggest that ozonated water might be useful to control oral infectious microorganisms, particularly C. albicans. Topical ozone therapy, when given in therapeutic doses through a controlled device, is the safest known therapy.

 
   References Top

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Correspondence Address:
Dr. Ganapathi Moger
Department of Oral Medicine and Radiology, Hitkarini Dental College and Hospital, Jabalpur, Madhya Pradesh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0970-9290.159146

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    Tables

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