Effects of Different Disinfectants
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Dentists, dental equipments and dental laboratories are exposed to different types of pathogenic microorganisms. The aim of this study was to investigate the effect of spraying three different types of disinfectant agents: sodium hypochlorite 0.525 %, Epimax and Deconex, on condensational silicon after 5 and 10 minutes.
Method and materials:
In this in vitro experimental study, 66 circular samples of condensational silicon impression materials of 1cm diameter and 2mm thickness (1.5 mm thickness of puty and 0.5 mm thickness of wash) were contaminated with Staphylococcus aureus ,(ATCC29213) Pseudomonas aeruginosa (ATCC27853) and Candida albicans fungus (PTCC5027). Except for control samples all of them were disinfected with sodium hypochlorite 0.525%, Deconex and Epimax by way of spraying. Afterwards, they were kept in plastic bags filled with humid cotton rolled for 5 and 10 minutes. In order to isolate bacteria the samples were immersed in 2% trypsin for one hour and then the solution was diluted with normal salin in portion of 1,1/2 and 1/4. The trypsin suspensions were transferred to culture plates and the number of colonies was counted after 24 and 48 hours for bacteria and after 72 hours for fungus. All data was analyzed by Mann Whitney test in SPSS software program (=0.05).
There was a significant difference between disinfection capability of Epimax and Deconex for all mentioned microorganisms after 5 minutes (P value=0.034). Also the difference was significant between disinfection ability of hypochlorite sodium 0.525% and Epimax for Staphylococcus aureus (P value=0.043) and Pseudomonas aeruginosa (P value=0.046) after 5 minutes. There was a significant difference between disinfection effects of Epimax and Deconex(P value=0.034) and hypochlorite sodium0.525% and Epimax(P value=0.034) for Pseudomonas aeruginosa after 10 minutes. By changing time from 5 minutes to 10 minutes, disinfection efficacy of Epimax and Hypochlorite sodium 0.525% changed dramatically. Deconex was completely efficient in eliminating 100 % of microorganisms in both 5 and 10 minutes.
This study revealed that condensational silicon can be effectively disinfected by three types of disinfectant agents by spraying method, although Deconex showed the best results compared to the other agents which indicate that this agent can be selected as a first choice to disinfect condensational silicon impression materials.
Condensational silicone, Disinfection, Impression materials,
Dentists and dental laboratories are exposed to different types of pathogen microorganisms. The main sources of transition of oral infections from patients to laboratory are impression materials, impression trays and poured stone casts. (1)
New studies have shown that, 67% of materials which are sent to dentallaboratories are infected by various microorganisms (2). The most frequently identified microorganisms are Streptococcus species, Staphylococcus species, Escherichia coli species, Actinomyces species, Antitratus species, Pseudomonas species, Enterobacter species, Klebsiella pneumonia and Candida species.(3). therefore, an effort should be made to eliminate most of these microorganisms and reduce the rate of infection transmission to dentistry laboratories. The International Dental Federation consequently have insisted on disinfecting all impressions from patients before sending them to laboratories (4). Also the American Dental Association has advised all dental workers to disinfect all patients’ impression trays (5). In some studies, it has been declared that washing the impression materials with tap water; only removes 40% of bacteria but other studies have reported that it has the capacity to reduce microorganisms up to 90 %( 6). The most common chemical disinfectants used by dentists are Alcohols, Aldehydes, Chlorine combinations, phenols, Biguanides, Iodide combinations and Ammonium (7). Based on the type of chemical disinfectant, there are two common methods to disinfect dental materials: 1) immersion 2) spraying (6).
Disinfection by immersing in the chemical materials has been proved to cover all surfaces in one time (8) while spraying is not capable of disinfecting all surfaces effectively and also cannot cover all undercuts, it significantly reduces the amount of distortion that take place in immersing method (6).
Silicon impression materials are the first group of polymeric impression materials (9). These materials have the best dimensional stability. Polyvinyl impression materials are the only impression materials which can be disinfected without any dimensional changes (10). Also, different methods such as soaking in glutaraldehyde for 30 minutes have been suggested to disinfect these materials. Use of hypochlorite sodium and phenol combinations with the soaking time less than pouring time have been suggested by Some studies (11).
J.Bustos investigated the effect of 0.05 % hypochlorite sodium and glutaraldehyde after 5 and 10 minutes on silicon impression materials. He declared that both of these disinfectants can efficiently prevent the bacterial growth on these impressions materials (12).
Ghahremanloo A et al also conducted a research in 2009 and investigated the antimicrobial effect of sodium hypochlorite 0.525 %, Deconex and Sanosil. They concluded that the use of 0.525 % sodium hypochlorite spray on the surface of alginate; effectively disinfects 96.6 % of the samples (13).
Since none of the mentioned disinfectant methods and agents have been accepted as a gold standard for disinfecting dental materials, finding an appropriate way seems rational.
The aim of this study was to investigate the disinfectant ability of spraying Deconex, Sodium hypochlorite 0.525 % and Epimax on condensational silicon impression material in 5 minutes and 10 minutes.
Methods and Materials
This randomized experimental and microbiological study was carried out with the cooperation of dental school and microbiology department of the Isfahan University of Medical Sciences, aiming to evaluate the disinfection effect of: Sodium Hypochlorite0.525%, Deconex and Epimax on the condensational silicon impression material.
In order to prepare samples, the heavy body impression material (putty) was mixed with the catalyst according to manufacturer instructions. The mixture was placed in a syringe with one centimeter diameter so that samples with 1.5 millimeter thickness were gained. Then the light body impression material (Wash) was mixed with the catalyst on a paper pad with a sterile spatula and was placed in the upper 0.5 millimeter of the syringe.
Eventually 66 samples, with 2mm thickness and 1 cm diameter, were prepared. In order to ensure that samples were kept sterile during preparation, three samples were selected as negative controls (blank) and were incubated on TSB culture for 24 to 48 hours; after which the bacterial growth was examined. For each bacterial type, 21 samples were used. Sodium hypochlorite0.525% was used to disinfect three of them for five minutes and three others for 10 minutes. Three samples were disinfected with Deconex for five minutes and three others for 10 minutes. And three samples were disinfected with Epimax for five minutes and three others for 10 minutes. At Last, three more samples were used as positive controls to check for any microbial pollution.
Preparation of Microbial Solution and Yeast
For many types of susceptibility testing, a standard inoculum of bacteria must be used. The standard inoculums were prepared according to 0.5 McFarland (1.5×108cfu/ml) by transferring 1-2 colonies of 18-24 hours cultures to TSB medium and incubated at 35°c until 0.5 McFarland turbidity of media was gained. For Candida albicans fungus, the sample was taken from 48 hour Saborose and Dextrose Agar cultures.
Contamination of Samples
To evaluate the disinfection effect of three mentioned substances, samples were separately polluted with microbial suspensions of Staphylococcus aureus (ATCC29213), Pseudomonas aeruginosa (ATCC27853) and Candida albicans fungus (PTCC5027). The impressions were put in sterile test tubes separately with 2 milliliter of microbial suspension in each tube and then all samples incubated at 35º C for one hour.
Disinfection of Samples and microbiological Surveys:
After contamination, all samples were rinsed with sterile distilled water for 30 seconds. In order to disinfect all samples, except controls, Sodium Hypochlorite 0.525%, Deconex and Epimax were separately sprayed 10 times on each sample for 15 seconds. Then the samples were put into sterile plastic bags containing sterile cotton, humidified with sterile distilled water to form a moisturized environment for 5 and 10 minutes.
Protease Trypsin, which is able to isolate the microbes from contaminated environments, was used. The ideal time and dilution for effective use of Trypsin is 60 minutes and 2% respectively. This time and dilution are based on the maximum microorganisms than can be isolated from the samples. After washing the samples with sterile distilled water for 30 seconds, they were put in Trypsin 2% solution for 60 minutes. Trypsin Suspensions with 1, and dilution were then prepared. Using 100 micro liter samplers, these samples were transferred to Muller Hinton Agar for Pseudomonas aeroginosa and Staphylococcus aureus bacteria and Saborow Dextrose Agar culture was selected for Candida albicans fungus. Using a Pasteur pipet bent with heat at 90 degrees, the samples were spread on cultures. After 24 and 48 hours incubation, the grown bacterial colonies on culture were counted. The grown fungus colonies of Candida albicans on Saborow culture were counted after 72 hours. All results, were analyzed by Mann Witney test in SPSS software program.(α=0.05)
According to the table 1, there is a significant difference between disinfection ability of Deconex and Epimax in 5 minutes (P Value=0.034). This difference is also significant for hypochlorite sodium and Epimax just for pseudomonas aeruginosa (p value=0.046) and staphylococcus Aureus (p value=0.043). In other cases, there is no significant difference in disinfection ability of materials in 5 minutes. (P value>0.05)
According to the table 2, significant difference between disinfection capability of hypochlorite sodium – epimax and deconex- epimax was just seen after 10 minutes for pseudomonas aeruginosa.
According to the table 3, Deconex completely eradicated all three kinds of microorganisms after 5 and 10 minutes. This is not true for hypochlorite sodium, as this material just eradicated staphylococcus Aureus and pseudomonas aeruginosa after 10 minutes 98.68 % eradication of pseudomonas aeruginosa was seen after 5 minutes.
After 5 minutes, epimax could eradicate 95.78 % of pseudomonas aeruginosa. This agent can completely eradicate candida Albicans and Staphylococcus Aureus after 10 minutes.
By increasing time from 5 to 10 minutes, disinfection ability of all agents increased, except for Deconex which is 100 % for all microorganisms in both 5 minutes and 10 minutes.
When there are large numbers of bacterial colonies, which are not countable, we can count the number of colonies using inverse dilution coefficient. Since real number of colonies was countable in dilution#1, and all results from other dilutions were similar to mentioned results, the results regarding to the number of colonies in other two dilutions were not reported.
Dentists practicing dentistry, encounter potentially harmful microorganisms. Patients are the most common source of microorganisms (14). Studies indicate that the surface of impressions taken out of the mouth is polluted with bacteria (15-18). As impressions and occlusal records cannot be sterilized by heat, chemical disinfection is still the common practicable method to eradicate microorganisms (19-21). So far there is no global way to disinfect impression materials (22). The American Dental Association (ADA) recommends soaking impression materials in disinfectant solutions for less than 30 minutes (23). Muller Bolla et al found that in European schools of dentistry, the soaking method is applied for 63% and 73% of Alginate impressions and silicon impressions respectively. The approximate time of disinfection was 10.3±6.3 minutes (20).
But Hiroshi Egusa and colleagues in 2008 showed that impressions from patients' mouths contain hazardous microorganisms like Streptocci, Staphylococcus aureus, Methicillin resistant Staphyloccocus, , Candida, Pseudomonas aeruginosa with rate of 100%, 55.6% 25,9%,5.6 % and 5.6 % respectively(22). These are opportunist pathogens that spread and transfer through the oral cavity (22). Candida causes common opportunist infections known as oral candidiasis, found in patients with immune deficiency (23). Pseudomonas aeruginosa is a deadly infectious agent that exists epidemically in hospital appliances and instruments (22) However, studies show that among population the spreading rate of S. pyogenes, S. pneumonia and S. aureus to pharynx is 10%, 20 to 32% and 30% respectively (14). This is the reason that in this survey, Staphylococcus aureus, Candida albicans and Pseudomonas aeruginosa were selected to investigate the disinfection capability of disinfectant agents.
By the year 1991, washing the impression materials with running water was the common way to remove microorganisms (22). This method can reduce about 90% of bacteria. (24) Running water can wash up saliva, blood and debris. But recent studies indicate that such methods cannot eliminate microorganisms from impression materials completely. Therefore washing the impression materials with running water, without disinfectants is not rational (22).
In this survey three common disinfectant agents were used. One of them is Hypochlorite sodium 0.525 %. which is used in housework too. This disinfectant can efficiently prevent microorganism's growth and disinfect the impression materials.
In a study by J.Bustos (12), it was shown that immersing silicon impressions in 0.5% hypochlorite sodium after 5 and 10 minutes dramatically prevent the bacterial growth in compare to the control group. Although in the recent study spraying method was used, results of the recent study are consistent with the mentioned study.
On the other hand Westerholm, Reuggeberg and colleagues also showed that spraying Sodium hypochlorite can effectively disinfect the impression materials (25-26). The Westerholm and et al study showed that Sodium hypochlorite can almost completely (99.99%) prevent the growth of S. aureus and this rate is about 99.60 % after 5 minutes and 100 % after 10 minutes for s.aureus in the present study. In another study by Ghahramanloo et al, spraying Sodium Hypochlorite 0.525 % could disinfect samples effectively (96.6%) after 10 minutes which is a good indicator of high capability of this agent (13). In studies by Westerholm, Rueggeberg and Ghahramanloo disinfection effect of these agents were assessed on irreversible hydrochloride (Alginate), but in this survey this effect has been assessed on condensational silicon. The results showed that there is no difference in disinfection capability of hypochlorite sodium regardless of impression material and this is a good proof for high penetration of this agent into impression materials porosities.
Decnex is an alcoholic based disinfectant agent, which in this study could effectively disinfect impression materials after 5 and 10 minutes (100 %). But in Ghahremanloo A et al study, this agent could eradicate 70.4 % of microorganisms (13). Maybe the main reason for this difference is that Ghahramanloo A used irreversible hydrochloride, which has more porosities and cause deep penetration of microorganism into this impression material and can define the lesser capability of disinfectant agent in eradicating microorganisms. The specific feature about this agent is that there is no difference in disinfection ability of Deconex after 5 minutes and 10 minutes.
In the present study, for the first time the antimicrobial effect of Epimax on impression materials was investigated. This agent could not effectively eradicate microorganism after 5 minutes in compare to two other agents, but after 10 minutes it completely (100 %) eradicated candida albicans and staphylococos aureos but this rate was 97.89 for Pseudomonas aeroginosa which shows the importance of using this agent for longer duration.
However, it should be mentioned that the results of the present study are not completely consistent with the results of other studies, due to different impression materials brands and usage time.
One of the shortcomings of the present study is that it is an in-vitro experimental study which is different from clinical and in-vivo situations. Usually impression materials remain 3 to 5 minutes in patient's mouth, while in our study it took 60 minutes to attach all the bacterial types to the samples as 60 minutes is an effective time for bacterial adherence. Also pressure while taking an impression and saliva can alter bacterial adherence capacity.
This study investigated the effect of three common disinfectant agents on two types of bacteria and one fungus. As so many dentists are concerned about viruses such as HIV and HBV, further studies should be conducted to find an effective way to eradicate these kinds of pathogens.
It can be concluded that sodium hypochlorite, Deconex and Epimax can effectively disinfect condensational silicon. This capability is higher for Deconex and it is recommended that Deconex can be the first choice to disinfect condensational silicon impression materials.
We would like to express our sincere acknowledgement in the support and help of microbiology department of Isfahan medical university and research department of dentistry faculty of Isfahan University of medical science.
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