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ORIGINAL RESEARCH
Year : 2021  |  Volume : 12  |  Issue : 3  |  Page : 149-154

Antimicrobial Efficacy of Sonic v/s Ultrasonic Activation on Disinfection of Root Canal System: An In Vitro Study


1 Department of Conservative and Endodontics, A B Shetty Memorial Institute of Dental Sciences, NITTE (Deemed to be University), Deralakatte, Mangalore, India
2 Department of Microbiology, K S Hegde Medical College NITTE (Deemed to be University), Deralakatte, Mangalore, India

Date of Submission25-Nov-2020
Date of Decision16-Mar-2021
Date of Acceptance29-Mar-2021
Date of Web Publication2-Nov-2021

Correspondence Address:
Shruthi H Attavar
Department of Conservative and Endodontics, A B Shetty Memorial Institute of Dental Sciences, NITTE (Deemed to be University), Deralakatte, Mangalore - 575018
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/denthyp.denthyp_178_20

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  Abstract 


Introduction: The key for long-term success in root canal therapy is the effective debridement with chemical irrigants before obturation. The purpose of this study was to compare the antimicrobial effect of sodium hypochlorite and chlorhexidine along with activation using sonic and an ultrasonic device in teeth when used in teeth infected with Enterococcus faecalis. Material and Method: A total of 90 single rooted mandibular premolar were infected with E. faecalis and placed in the incubator for 24 hours .The specimens were divided into six groups as follows: Group I: positive control; Group II: negative control; Group III: sodium hypochlorite + passive ultrasonic activation for 20, 40, and 60 seconds; Group IV: chlorhexidine + passive ultrasonic activation; Group V: sodium hypochlorite + endoactivator; and Group VI: chlorhexidine + endoactivator activation. After disinfection, dentinal shavings were collected using H file and subcultured in brain heart infusion agar (BHI) broth further streaked in brain heart agar plates to check the colony counting. Result: Bonferroni post hoc test was done to compare the significant difference between different experimental groups. Group III showed a significant difference in the antimicrobial activity compared to Group IV, V, and VI with P < 0.05. One-way analysis of variance (ANOVA) analysis was done to analyze the intergroup comparison of the bacterial count at various time intervals (20, 40, and 60 seconds). It was observed that there was a statistically significant difference with P < 0.01 between the time interval of different experimental groups. Conclusion: Within the limitation of this study, sodium hypochlorite along with passive ultrasonic irrigation showed a reduction in E. faecalis count compared to the other irrigating protocols used in the study and the results were statistically significant.

Keywords: Chlorhexidine, endo activator, passive ultrasonic activation, E. faecalis, sodium hypochlorite


How to cite this article:
Attavar SH, Hegde MN, Shetty V. Antimicrobial Efficacy of Sonic v/s Ultrasonic Activation on Disinfection of Root Canal System: An In Vitro Study. Dent Hypotheses 2021;12:149-54

How to cite this URL:
Attavar SH, Hegde MN, Shetty V. Antimicrobial Efficacy of Sonic v/s Ultrasonic Activation on Disinfection of Root Canal System: An In Vitro Study. Dent Hypotheses [serial online] 2021 [cited 2021 Dec 7];12:149-54. Available from: http://www.dentalhypotheses.com/text.asp?2021/12/3/149/329759




  Introduction Top


The removal of microorganisms, toxins, and their irritants from the infected root canal system determines the success of endodontic treatment so as to best obtain a host-manageable bioburden.[1] The disinfection of root canal system by chemomechanical preparation plays an critical role in reducing the bacterial load.[2] The mechanical instrumentation of the root canal along with the synergetic effect of antimicrobial property of irrigants enhances the elimination of microorganisms.[3] The mechanical preparation of the root canal system is an essential phase since it aims to facilitate the easy flow of irrigants within the canal space.[4]

E faecalis is gram-negative facultative anaerobic cocci found in 4% to 40% of primary endodontic infection and nine times more likely in failed root canal cases. The prevalence of E faecalis in root filled teeth with periradicular lesions is from 24% to 77%.[5]

Sodium hypochlorite is a potent antimicrobial agent introduced by Walker in the year 1936. It is used in the concentration between 05% and 6%. The most important property of sodium hypochlorite is that it dissolves the organic pulp tissue and collagen.[6] Chlorhexidine gluconate a bisguanide derivative shows its antimicrobial property by destroying the microbial cell wall and attacks the bacterial cytoplasm causing coagulation of intracellular components.[7] The application of a final irrigating solution that remains active not only at the time of application but also over a fairly long period of time thereafter stands as one strategy for preventing bacterial recolonization or eliminating the bacteria that persist after the root canal treatment.[7]

Traditionally, irrigants were delivered into the root canal using syringe and metal needles of various sizes and tip design. Clinically, this method was ineffective because irrigants could not penetrate into peripheral areas such as fins, isthmus, and deltas. In an attempt to improve the penetration and efficacy of irrigants, various mechanical devices have been developed to enhance the cleaning efficacy of the root canal system.[8]

Alternatively, different solution agitation methods and devices have been proposed to increase effectiveness of irrigating solutions, including agitation with hand files, plastic instruments, and sonic and ultrasonic devices. Sonic activation generates mechanical oscillation, mainly at the file tip, at 1 to 6000 Hz frequency, while ultrasonic devices generate microstreaming along the file with 40,000 to 45,000 Hz frequency.

Passive ultrasonic irrigation was first described by Weller et al. in 1980. The term “passive” does not adequately describe the process, as it is in fact active; however, when it was first introduced, the term “passive” related to the “noncutting” action of the ultrasonically activated file. passive ultrasonic irrigation (PUI) relies on the transmission of acoustic energy from an oscillating file or smooth wire to an irrigant in the root canal. The energy is transmitted by means of ultrasonic waves and can induce acoustic streaming and cavitation of the irrigant.[9]

The synergetic effect of sodium hypochlorite and chlorhexidine along with ultrasonic activation and endoactivator in disinfection of the root canal system has not been evaluated. Therefore, the aim of the present study is to determine the efficacy of sonic v/s ultrasonic device in eradication of E. faecalis from the root canal system.


  Material and Method Top


Sample size determination was done based on 5% level of significance and 80% power effect size of 0.4. The total sample size for six groups is 90, that is, 25, in each group.

Selection and standardization of specimen

Ninety noncarious single rooted mandibular premolars were collected and sterilized according to occupational safety hazard (OSHA) regulation. The presence of single root was confirmed radiographically using digital radiograph. The exclusion criteria were teeth with open apex, resorption, caries, and fractured roots. The crown of all teeth was sectioned using high speed carbide disk at the level of cemento enamel juntion (CEJ). Working length was determined using size 15 k file (Dentsply Sirona) 1 mm short of the apical foramen. Cleaning and shaping of the root canal was done with the protaper gold rotary file system using the crown down technique till file size F 3 (Dentsply Sirona). The apical foramen was sealed using epoxy resin to prevent bacterial leakage.

Specimen sterilization

The tooth samples were sealed in a plastic pouch and sterilized using ethylene oxide sterilization.

Cultivation of E. faecalis and specimen contamination

A suspension of 50 mL of E. faecalis [American type culture collection (ATCC) 29212] was incubated in 5 mL of brain heart infusion agar (BHI) broth at 37°C for 24 hours. The concentration of the inoculum was adjusted to McFarland scale, which corresponds to the concentration of 3 × 108 cells/mL and optical density (OD) of 550 nm. The root canal of all teeth was inoculated with the bacterial strain of E faecalis using sterile insulin syringe and incubated for 21 days at 37°C.

Experimental groups

Group I: N = 15–Positive control group − Brain heart infusion broth with inoculation of E. faecalis

Group II: N = 15–Negative control − Brain heart infusion broth without inoculation of E. faecalis

Group III: N = 15–Sodium hypochlorite + Passive ultrasonic activation (Irrisafe tips, Aceteon Satelec)

Subgroup A: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Passive ultrasonic − activation − 20 seconds

Subgroup B: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Passive ultrasonic activation − 40 seconds

Subgroup C: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Passive ultrasonic activation − 60 seconds

Group IV: N = 15–Chlorhexidine + Passive ultrasonic activation (Irrisafe tips, Aceton Satelec)

Subgroup A: N = 5–5 mL of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Passive ultrasonic − 20 seconds

Subgroup B: N = 5–5 ml of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Passive ultrasonic − 40 seconds

Subgroup C: N = 5–5 mL of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Passive ultrasonic − 60 seconds

Group V: N = 15–Sodium hypochlorite + Endoactivator (Dentsply Sirona) activation

Subgroup A: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Endoactivator − 20 seconds

Subgroup B: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Endoactivator − 40 seconds

Subgroup C: N = 5–5 mL of 3% sodium hypochlorite (Vensons, India) + Endoactivator − 60 seconds

Group VI: N = 15–Chlorhexidine + Endoactivator activation

Subgroup A: N = 5–5 mL of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Endoactivator − 20 seconds

Subgroup B: N = 5–5 mL of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Endoactivator − 40 seconds

Subgroup C: N = 5–5 mL of 2% chlorhexidine (DC Chlor, Red Gold Mines) + Endoactivator − 60 seconds

Microbial sampling

After disinfection of the root canal, the dentinal shavings were collected using H file (Dentsply Sirona) and placed in BHI broth in a micro tube and incubated for 24 hours. The samples were streaked on brain heart infusion agar plate and incubated at 37°C for 24 hours [Figure 1].[10],[11]
Figure 1 Stages in the experimental procedure.

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Statistical analysis

Statistical analysis was done using Statistical Package for the Social Sciences (SPSS) 23.0 software for windows. The quantitative data obtained from the mean colony forming unit of the Bonferroni post hoc test was done to compare the significant difference between different experimental groups. One-way analysis of variance (ANOVA) was done to analyze the intergroup comparison of the bacterial count at various time intervals (20, 40, and 60 seconds).


  Results Top


A pairwise comparison was done using Bonferroni post hoc test to analyze the level of significance between groups activated with different time intervals (20, 40 and 60 sec) [Table 1]. P < 0.05 was considered statistically significant. Bonferroni post hoc test was done to compare the significant difference between different experimental groups [Table 2]. Group III showed a significant difference in the antimicrobial activity compared to Group IV,V, and VI with P < 0. 05.
Table 1 Pairwise comparison of log10 CFU of bacteria between different time intervals

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Table 2 Pairwise comparison of log10 CFU of bacteria between different study groups

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One-way ANOVA analysis was done to analyze the intergroup comparison of the bacterial count at various time intervals (20, 40, and 60 seconds) [Table 3]. It was observed that there was a statistically significant difference with P < 0.01 between the time interval of different experimental groups. Bonferroni post hoc test was done to compare the bacterial count between the groups at different time intervals [Table 4]. The results of the study showed that there was no statistical significant difference in group group III activated for duration of 20 and 40 sec, but there was a statistically significant difference between groups activated for 20 and 60 seconds with P value of 0.02. In Group IV, there was a statistically significant difference between activation time of 20 seconds when compared to 40 and 60 seconds with P value of 0.02 and 0.04, respectively, but there was no statistically significant difference between activation time of 40 and 60 seconds. In Group V, there was a statistically significant difference between groups irrigated for 20, 40, and 60 seconds. In Group VI, there was no statistical significance between groups irrigated for 20 and 40 seconds, but there was a significant difference between groups irrigated between 40 and 60 seconds.
Table 3 Comparison of log10 CFU of bacteria between different study groups at each time interval

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Table 4 Comparison of log10 CFU of bacteria between different time intervals in each study groups

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  Discussion Top


The aim of endodontic treatment is the elimination of bacteria and the prevention of reinfection. This is accomplished by effective mechanical instrumentation and the use of irrigants and intracanal disinfectants. Sodium hypochlorite is the most effective antimicrobial solution; the property is attributed to the presence of hypochlorous acid. Chlorhexidine is a biguanide with a broad spectrum antimicrobial activity against both gram-positive and gram-negative organisms with the property of substantivity.[12]

Passive ultrasonic activation is one of the predominant irrigation regimens in endodontic practise with the advantage of acoustic streaming that removes the intraradicular biofilm by increasing the shear stress on the walls of the root canal. Endoactivator is a sonic device that operates at a frequency of 1 to 8 kHz with the flexible plastic tips that do not stop when it comes in contact with root canal wall, and hence it can be used in curved canals.[13]

Among irrigant agitation equipment, sonic and ultrasonic devices have been extensively tested (De Gregorio et al. 2009, Gu et al. 2009, Gulabivala et al. 2010). Sonic activation generates mechanical agitation primarily on the tip of files, whereas ultrasonic activation generates microstreaming around the file and a secondary acoustic streaming (Ahmad et al. 1987, De Gregorio et al. 2009, Gu et al. 2009, Gulabivala et al. 2010). Ex vivo studies in root canal systems with restricted flow through the apical foramen, which better simulates fluid flow in clinical conditions (Tay et al. 2010), revealed that penetration of solutions is enhanced by ultrasonics and, to a lesser degree, sonic activation (De Gregorio et al. 2009, Paragliola et al. 2010, Sainz-Pardo et al. 2014).[14]

In the present study, a long-term incubation of E. faecalis for 21 days was done since a short duration of 3 days would lead to the formation of young planktonic cells compared to 21 days incubation that leads to the penetration of microorganisms into dentinal tubules. Since we focused on the in-depth penetration of irrigants into the dentinal tubule, a long duration of incubation was selected.[15]

In this study, the activation time was used at an interval of 20, 40, and 60 seconds. As a general rule, the greater the time of activation, the greater the canal cleanliness [Table 1].[16] Ultrasonic activation of NaOCl from 30 seconds to 1 minute for each canal with three cycles of 10 to 20 seconds (with constant irrigant renewal) seems to be a sufficient to obtain cleaned canals at the end of the preparation. Sodium hypochlorite along with passive ultrasonic activation, that is, Group III, showed an increased antimicrobial activity compared to the Group IV, V, and VI and the results were statistically significant [Table 2]. One of the reasons could be the higher frequency of 30,000 Hz and lower amplitude of 11 mm that during oscillation causes shear stress in root canal walls. The bacterial cell walls get teared off due to this high velocity and energy produced by the ultrasonic device. Furthermore, these ultrasonic devices can cause degassing of the solution and decomposition of molecules. In addition to these factors, the ultrasonic wave that vibrates in the irrigating solution can convert sound energy into heat, which leads to heat generation and increases in the temperature.[17] In general, it is believed that sonic activation is seemingly less effective than the use of ultrasound as a more velocity fluid stream is induced with the latter. Furthermore, instrument size, tip diameter, instrument taper, confinement of the instrument within the canal, and the type of irrigants have an influence on the cleaning and disinfection efficacy associated with enhanced fluid streaming.[18]

The present study also evaluated the effect of sodium hypochlorite in disinfection of root canal system and it was shown that it had a better antimicrobial efficacy compared to chlorhexidine. This can be attributed to the fact that sodium hypochlorite, when it comes in contact with organic debris, leads to the formation of hypochlorous acid that interferes with the oxidizing sulfhydryl group of the bacterial enzyme, thus disrupting the microbial metabolism and killing of bacterial cells.[19] It has been demonstrated that ultrasonic activation of sodium hypochlorite dramatically enhances the effectiveness of cleaning the root canal space. Besides, it greatly increases the flow of liquid and improves both the solvent and antibacterial capacities and the removal effect of organic and inorganic debris from root canal walls.


  Conclusion Top


Within the limitation of this study, sodium hypochlorite along with passive ultrasonic irrigation showed a reduction in E faecalis count compared to the other irrigating protocols used in the study and the results were statistically significant.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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    Figures

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    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

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