Abstract
Objective
To evaluate the effectiveness of three rotary file systems on the elimination of Enterococcus faecalis (E. faecalis) biofilms in single-rooted teeth.
Methods
A total of 70 mandibular premolars were included in this study in which 60 root canals were inoculated with E. faecalis. The positive control group (n=5) received bacterial inoculation without mechanical preparation. The negative control group (n=5) was neither infected nor mechanically prepared. The teeth were randomly assigned to 3 experimental groups (n=20 per-group) according to application of the rotary files: Group 1: XP-endo Rise Shaper (FKG, Le Crêt-du-Locle, Switzerland); Group 2: XP-endo Shaper (FKG, Le Crêt-du-Locle, Switzerland); Group 3: ProTaper Gold (Dentsply Sirona, Erlangen, Germany). Samples were collected using paper points before (S1) and after (S2) mechanical preparation. Colony-forming units (CFUs) were used to quantify bacterial loads.
Results
Significant differences in bacterial counts were observed from S1 to S2 in all tested groups (p<0.001). Intergroup comparisons revealed no significant differences between the XP-endo Rise Shaper and XP-endo Shaper groups (p>0.05) and between XP-endo Shaper and ProTaper Gold groups (p>0.05). Significant differences were observed between the XP-endo Rise Shaper and ProTaper Gold groups (p<0.05). The XP-endo Rise shaper group showed the largest reduction in bacterial loads (98.2%), followed by the XP-endo Shaper (96.0%) and ProTaper Gold (93.8%) groups.
Conclusion
All tested rotary files were able to reduce bacteria but the novel single-file system (XP-endo Rise shaper) was superior in reducing bacterial load compared to XP-endo Shaper and the multiple file system (ProTaper Gold).
Keywords: Disinfection, Enterococcus faecalis, root canal instrumentation, rotary files
HIGHLIGHTS
This study evaluated the effects of XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold in bacterial load reductions of E. faecalis in single-rooted teeth.
Antimicrobial testing was performed using the tube dilution method and CFU counts performed using a Promega Colony Counter.
The XP-endo Rise shaper promoted the most significant reduction in bacterial counts compared to the other two files after root canal preparation.
INTRODUCTION
The main goal of endodontic treatment is to eliminate bacteria, their byproducts, and debris from the root canal system, which are key factors in the development of apical periodontitis (1). Achieving this goal requires a combination of effective chemo-mechanical debridement techniques and thorough disinfection protocols to ensure successful treatment outcomes (2). The microbial interactions within the root canal shape a dynamic bacterial community, where biological variety influences both nature and the progression of infection (3). Among these microorganisms, Enterococcus faecalis (E. Faecalis) has reaped substantial attention due to its role in endodontic failures. It is a gram-positive, facultative anaerobe capable of resisting the actions of chemo-mechanical agents and can persist following endodontic treatment (4, 5). Its genetic adaptability and ability to penetrate the dentinal tubules significantly contribute to endodontic failures and secondary infections (6). The pathogenic nature of E. Faecalis, along with the intricate anatomy of the root canal system poses a significant challenge to achieving complete disinfection of the root canal system (7). However, thorough chemo-mechanical preparation can reduce more than 90% of bacterial endotoxin in infected root canals (8).
Mechanical preparation of the root canals is a cornerstone of endodontic therapy, and significant advancements in instrumentation have revolutionized this process. The advent of Nickel Titanium (NiTi) rotary instruments, particularly those with progressively tapered designs, has notably improved the efficiency and precision of canal preparation (9). Among these, the ProTaper Gold (Dentsply Sirona, Erlangen, Germany) represents an established multi-file system that incorporates gold heat treatment technology. The system includes three shaping files labeled SX, S1, and S2 and five finishing files named F1, F2, F3, F4, and F5, enhance its cutting efficiency and flexibility. Despite these advancements, complete bacterial elimination from root canals remains unattainable (10). In recent years, single rotary file systems have gained popularity for their ability to simplify instrumentation while maintaining effectiveness. The XP-endo Shaper (FKG, Le Crêt-du-Locle, Switzerland), a next-generation adaptive single-file system, employs MaxWire technology that enables the instrument to expand and conform to the canal's morphology by transitioning between martensitic and austenitic phases. This file, with its triangular cross-section booster tip and three cutting edges, has shown promise in reducing bacterial counts within root canals (11, 12). Building on this design, the XP-endo Rise Shaper incorporates a newly engineered booster tip with six cutting edges, enhancing both control and reproducibility (12). Although various in vitro studies have highlighted the efficacy of the XP-endo Shaper in reducing bacterial loads (13–15), comparative evidence regarding the effectiveness of the XP-endo Rise Shaper remains scarce.
A recent meta-analysis underscored the inability of both rotary and reciprocating systems to eliminate bacteria from root canals, emphasizing the need for further research into advanced file systems (16). Addressing this gap, the present in-vitro study aims to evaluate the efficacy of XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold in reducing bacterial loads in single-rooted teeth. By comparing these systems, this study seeks to contribute valuable insights into the effectiveness of three-dimensional rotary file systems in enhancing endodontic outcomes.
MATERIALS AND METHODS
Ethical Approval
The study was approved by our Institutional Review Board (IRB-COD-STD-106-May-2023) and was conducted in accordance with the principles of the Declaration of Helsinki.
Sample Size Calculation
Sample sizes were calculated using G* Power software requiring a minimum of 45 samples (15 per group). The sample size was increased to 20 per group to account for potential losses, resulting in 70 mandibular premolars including 10 teeth for positive and negative controls (5 for each). Sixty premolars were contaminated with E. faecalis for antimicrobial testing. An alpha error of 0.05 and a power of 0.8 were used for all calculations.
Selection and Categorization of Samples
Mandibular first and second premolars were subjected to periapical radiographic evaluation. A total of 70 premolars with complete root development, single canals, and root curvature ≤5° were selected for this in vitro study. For allocation concealment, a total of 60 samples were sequentially numbered and placed in an opaque sealed envelope. The samples were randomly allocated into three groups using randomization.org. Group 1 consisted of preparations using the XP-endo Rise Shaper (n=20); Group 2 included the XP-endo Shaper (n=20); and Group 3 comprised the ProTaper Gold (n=20). The external surfaces of all the teeth underwent ultrasonic scaling to remove any plaque or debris before autoclave sterilization at 121°C for 20 minutes. Traditional access cavity preparation was performed on all teeth using a round bur (Johnson-Promident, New York, USA) and a high-speed handpiece (NSK, Tokyo, Japan). Teeth were decoronated using a diamond disk (Horico, Berlin, Germany) under water cooling. and standardized to a length of 20 mm by placing a K-file (size #10; Dentsply Sirona, Ballaigues, Switzerland) in the canal from the reference point to the apex. The glide path was established by advancing a #10 K-file to the pre-determined working length at the apical constriction. The apical third was shaped to accommodate a master apical K-file (size #30). All the teeth were subjected to continuous manual irrigation with 2 mL of distilled water. 17% EDTA and 5.25% sodium hypochlorite (1 mL each) were used to eliminate the smear layer after which the samples were rinsed in phosphate-buffered saline (2 mL). The apical foramen of prepared canals was occluded with cyanoacrylate to prevent bacterial leakage from the apex (17). Once the samples were completely dried, they were stored in a container filled with distilled water until bacterial inoculation was performed.
Inoculation for Biofilm Formation
Positive control teeth (n=5) were inoculated with bacteria without rotary file preparation, whereas the negative control group (n=5) received no bacterial inoculation nor rotary file preparation to confirm strict aseptic conditions. The bacterial inoculations and the specimen contamination protocol were adapted from Carvalho et al. (13). Three to four colonies of E. faecalis (ATCC29212) were sub-cultured from blood agar plates onto BHI agar plates at 37°C for 24 h. After 24 h of incubation, 2–3 colonies from the sub-cultured agar plate were transferred using a sterile plastic spreader (Copan, Mantua, Italy) to a flask containing 1ml of BHI broth and incubated at 37°C for 24 h to achieve exponential growth. Cultures were adjusted to McFarland standard #1 (3×108 CFU/mL). Bacterial suspensions (20 µl) were inoculated into the samples using a sterile pipette of size #0.5 and the canal was circumferentially filed for 10 seconds using a #15 K file within 1 mm of working length to ensure coverage of the entire root canal extension. The samples were transferred into Eppendorf tubes containing 980 μL of BHI broth and incubated at 37°C for 10 days. To ensure a steady supply of medium and to eliminate excess bacterial cells, the culture medium was replenished every alternative day throughout the incubation period. On the tenth day, the samples were extracted from the Eppendorf tubes, the excess culture medium was discarded, and the external surfaces of the roots were cleaned with sterile gauze.
Pre and Post-preparation Sample Collection
Pre-preparation (S1) bacteriological sample collection was collected from the canals using sterilized paper points (Diadent Group International Inc., Songjeong-dong, Korea; size #30). A first paper point was introduced along the dentinal wall, reaching the complete working length for a total of 60 s. Two further paper points were introduced in succession. All were transferred to 0.5 mL of BHI broth, vortexed for 30 s, and shaken for 60 s. This dilution was labeled as the ‘S1 dilution’. The main objective of the S1 sample is to determine the total number of bacteria present within the canals before rotary file intervention. Samples collected post-intervention (S2) were obtained from the canals, following the methodology outlined for S1. This dilution was labeled as the ‘S2 dilution’ as it contained the bacterial sample in its full concentration post-instrumentation.
Rotary File Preparation
XP-endo Rise Shaper Group: A total of 20 teeth were treated using the XP-endo Rise Shaper 30/.04 at a rotational speed of 800 rpm and a torque setting of 1 Ncm, facilitated by an electric motor (VDW. Silver Reciproc, Dentsply Sirona, Ballaigues, Switzerland). Teeth were shaped using a master apical file (size #30). The XP-endo Rise Shaper was used in a vertical motion until the desired working length was achieved. Movements were repeated on 10 occasions up to the working length. Teeth were subjected to manual irrigation with distilled water through a syringe equipped with a 30-gauge NaviTip needle (Ultradent, Utah, USA) positioned 3 mm short of the working length to prevent binding of the needle in the canal. A standardized 20 ml of irrigating solution was utilized across all specimens, equating to 1 ml per tooth as previously described (18). The irrigating solution was removed using 0.36-mm capillary tips (Ultradent, Utah, USA). The duration of contact between the irrigants and the walls of the root canal was 2 min, constituting the irrigation period.
XP-endo Shaper Group: A total of 20 teeth were treated using the XP-endo Rise Shaper 30/.04 at a rotational speed of 800 rpm and a torque setting of 1 Ncm, facilitated by a VDW.Silver Reciproc electric motor. Each tooth was shaped to a master apical file size #30. The XP-endo Shaper was used in a vertical motion up to ten times until the working length was reached. The irrigation protocol was identical to that outlined for the XP-endo Rise Shaper group.
ProTaper Gold Group: A total of 20 teeth were treated using the ProTaper Gold file with a continuous rotational speed of 300 rpm and a torque setting of 2.5 Ncm, powered by an electric motor (NSK Endo-mate TC2, NSK, Tokyo, Japan). The shaping of each tooth was achieved from S1 to F3 using a vertical motion. The irrigation protocol utilized was identical to that outlined for the XP-endo Rise Shaper group.
Each rotary kit was limited to use in a single canal after which the file was sterilized. Each rotary file was used to prepare a maximum of four canals in total (13).
Subculturing on BHI Agar Plates
A series of 10-fold dilutions of S1 and S2 were prepared, extending to a dilution factor of 10−4, by combining 0.1 ml of the sample with 0.9 ml of BHI broth. A total volume of 20 μL of S1 and S2 master dilutions was subcultured using sterile plastic spreaders onto BHI agar plates at 37°C for 24 h (Fig. 1). CFUs were counted after 24 h as previously described (18) using Promega Colony Counter (Figs. 2, 3). (Promega, Madison, WI, USA) (19). The procedures included root canal preparation, bacterial inoculation, sample collection, and colony count were performed by a single operator to avoid inter-operator variability.
Figure 1.
Master dilutions sub-cultured onto BHI agar plates
BHI: Brain heart infusion
Figure 2.
Representative captures of CFU counts of S1 using Promega Colony Counter
CFU: Colony forming units
Figure 3.
Representative captures of CFU counts of S2 using Promega Colony Counter
Statistical Analysis
Data were analyzed using “SPSS for Windows (Chicago, IL, USA) version 29.0.2. Numerical data are shown as the mean±standard deviation (SD), accompanied by 95% confidence intervals. Bacterial loads were analyzed at S1, S2, and S1-S2. A one-way ANOVA was used to compare parametric data. Pairwise comparisons between groups were performed using Tukey’s post-hoc test. Intragroup comparisons were assessed through paired t-test. P-values<0.05 were deemed significant.
RESULTS
Bacterial Loads
The mean bacterial loads at S1 are shown in Table 1 and Figure 4. Intergroup comparisons at S2 indicated significant differences (p<0.05) across all groups (Table 2) and significant differences were observed between the XP-endo Rise Shaper and ProTaper Gold groups (p<0.05). The mean bacterial count at S2 is shown in Table 3 and Figure 5. Figure 6 shows mean bacterial loads at S1-S2, in which all groups exhibited significant reductions (p<0.05). Intergroup analysis revealed significant differences between the XP-endo Rise Shaper and ProTaper Gold groups (p<0.05) (Table 4). The XP-endo Rise Shaper group showed the greatest reduction in bacteria (98.2%), followed by the XP-endo Shaper (96%) and ProTaper Gold (93.8%) groups (Fig. 7).
TABLE 1.
Descriptive statistics of CFU bacterial count before rotary file preparation (S1) for XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold
| S1 | Mean | Median | Range | SD |
|---|---|---|---|---|
| XP-endo Rise Shaper | 55.5 | 54 | 45 | 14.4 |
| XP-endo Shaper | 49.6 | 46 | 50 | 14.84 |
| ProTaper gold | 43.2 | 45 | 44 | 11.52 |
CFU: Colony forming units, SD: Standard deviation
Figure 4.
Mean bacterial count before rotary file preparation (S1)
TABLE 2.
Descriptive statistics and Intergroup comparisons of CFU bacterial count after rotary file preparation (S2) for XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold
| S2 | Mean | Median | Range | SD |
|---|---|---|---|---|
| XP-endo Rise Shaper | 1.7 | 1.5 | 4 | 1.21 |
| XP-endo Shaper | 2.25 | 2 | 5 | 1.11 |
| ProTaper gold | 2.95 | 3 | 6 | 1.53 |
| One-way Anova F-test value | F=4.618 | |||
| P value | 0.014* | |||
| Pairwise comparison | ||||
| Pairwise comparison – Tukey’s post hoc test | Mean difference | p | ||
| XP-endo Rise Shaper Vs XP-endo Shaper | 0.55 | 0.382 | ||
| XP-endo Rise Shaper Vs ProTaper Gold | 1.25 | 0.01* | ||
| XP-endo Shaper Vs ProTaper Gold | 0.7 | 0.215 | ||
: Represents statistically significant differences (p<0.05). CFU: Colony forming units, SD: Standard deviation
TABLE 3.
Mean, SD, and Intergroup comparisons of CFU bacterial count at S1-S2 for XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold
| S1-S2 | Mean | SD |
|---|---|---|
| XP-endo Rise Shaper | 54.80 | 13.89 |
| XP-endo Shaper | 47.35 | 13.53 |
| Pro taper gold | 40.25 | 11.08 |
| One way Anova F-test | F=4.945 | |
| P value, significance | p=0.01* | |
| Pairwise comparison – Tukey’s post hoc test | Mean Difference | p |
| XP-endo Rise Shaper Vs XP-endo Shaper | 5.15 | 0.422 |
| XP-endo Rise Shaper Vs ProTaper Gold | 12.75 | 0.008* |
| XP-endo Shaper Vs ProTaper Gold | 7.6 | 0.159 |
: Represents statistically significant differences (p<0.05). S1-S2: Absolute mean difference
Figure 5.
Mean bacterial count after rotary file preparation (S2)
Figure 6.
Mean bacterial count at S1-S2
TABLE 4.
Intragroup comparison and reduction percentage of CFU bacterial count for XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold
| Groups | S1 Mean (SD) | S2 Mean (SD) | Paired t-test | p | Reduction % |
|---|---|---|---|---|---|
| XP-endo Rise Shaper | 55.5±(14.4) | 1.7±(1.21) | t=16.791 | <0.001** | 98.2% |
| XP-endo Shaper | 49.6±(14.84) | 2.25±(1.11) | t=13.976 | <0.001** | 96% |
| ProTaper Gold | 43.2±(11.52) | 2.95±(1.53) | t=16.016 | <0.001** | 93.8% |
: Represents highly statistically significant differences (p<0.001). CFU: Colony forming units, S1: Before rotary file preparation, S2: After rotary file preparation, SD: Standard deviation
Figure 7.
Mean bacterial reduction percentage at S1-S2
DISCUSSION
This study evaluated the effects of XP-endo Rise Shaper, XP-endo Shaper, and ProTaper Gold in bacterial load reductions of E. faecalis in single-rooted teeth. Results revealed significant differences in mean bacterial loads from S1 to S2 between groups. Extracted human mandibular premolars were utilized to simulate clinical performance (20). The rotary file preparation included distilled water as an irrigant, permitting a focused assessment of the mechanical action of the files without the influence of antimicrobial agents (17).
The PCR technique is more specific, sensitive, and faster when compared to the culture plate method. The culture plate technique was used in this in vitro study as the bacteria under investigation were species-specific and cultivable (21–24). The contamination protocol was validated as samples at S1 had 100% culturable bacteria. CFUs were counted using Promega Colony Counter, a smartphone-based application with a fully automated colony counter to minimize errors (25, 26).
The XP-endo Rise Shaper demonstrated superior bacterial load reduction compared to the ProTaper Gold system, with no significant differences between XP-endo Rise Shaper and XP-endo Shaper or between XP-endo Shaper and ProTaper Gold. However, a significant difference between groups ProTaper Gold and XP-endo Rise Shaper was achieved. The notable difference between ProTaper Gold and XP-endo Rise Shaper can be attributed to their design features. The XP-endo Rise Shaper's triangular cross-section with a consistent 0.01% taper allows it to occupy more canal space. At the same time, its high rotational speed and elasticity enable dynamic movement within the canal, effectively preventing debris accumulation and eradicating bacterial biofilms (27). In contrast, ProTaper Gold’s design may compact dentine into lateral walls, potentially obstructing bacteria removal from dentinal tubules (23).
In contrast to this study, significant differences in the ability to reduce bacterial loads has been reported between ProTaper Gold and XP-endo Shaper (23). These differences are likely due to the supplemental disinfection using XP-endo Finisher (FKG, Le Crêt-du-Locle, Switzerland) (12).
Single-file instruments are thought to lessen disinfection in root canals due to simplified mechanical preparation, resulting in less instrumentation time and decreased antibacterial irrigant volume. However, recent research confirms that these NiTi rotary systems effectively clean and shape root canals, reducing procedural times by 40% compared to conventional methods that use continuous motion (28, 29). A major drawback of these instruments is the increased fracture risk due to repetitive use and the possibility of cross-contamination. One study showed no significant differences in bacterial loads across different instrument sizes and tapers (30). However, Siddique et al. (23) confirmed that the XP-endo Shaper significantly lowered bacterial load compared to ProTaper Next and Gold, aided by ultrasonic irrigation with 17% EDTA and 5.25% NaOCl.
Due to a lack of studies on the effectiveness of XP-endo Rise Shaper in reducing E. faecalis biofilm, a comparison with the literature could not be attained. Our results indicated that while bacterial load decreased, no instrument completely eliminated bacteria from the canal. This highlights the need for advanced instrumentation and irrigation systems to improve disinfection and host response for enhancing the predictability of endodontic treatment outcomes.
The mono-species biofilm model in this in vitro study provides simplicity, standardization, and control, but poses challenges in applying findings to clinical practice due to the presence of in vivo multispecies biofilms, potentially affecting results. Limitations also include the inability of paper points to effectively retrieve bacteria from within the dentinal tubules. Although results show statistical differences, they may not indicate clinical significance, as periapical infection development depends on bacterial numbers and the host's defense mechanisms (31). Further clinical trials are essential to validate these findings, as interactions with the host components remain unobserved.
CONCLUSION
All the examined rotary files were able to reduce bacteria but the novel single-file system (XP- endo Rise shaper) was superior in reducing E. faecalis bacterial count when compared to its predecessor and the multiple file system (ProTaper Gold).
Footnotes
Please cite this article as: Kannan R, Eid B, Adtani P, Elsewify T, Elemam R. Effectiveness of Three Rotary File Systems on Enterococcus Faecalis in Single Rooted Teeth: An In Vitro Study. Eur Endod J
Disclosures
Ethics Committee Approval
The study was approved by the Gulf Medical University Ethics Committee (no: IRB-COD-STD-106-May-2023, date: 01/06/2023).
Authorship Contributions
Concept – R.K., B.E.; Design – B.E., R.E.; Supervision – B.E., P.A., R.E.; Funding – R.K.; Materials – P.A.; Data collection and/or processing – R.K., T.E.; Data analysis and/or interpretation – R.K., R.E., P.A.; Literature search – R.K., B.E., R.E.; Writing – R.K., T.E.; Critical review – T.E.
Conflict of Interest
All authors declared no conflict of interest.
Use of AI for Writing Assistance
The authors declared that no artificial intelligence-assisted technologies were used during the submission of this work.
Financial Disclosure
The authors declared that this study received no financial support.
Peer-review
Externally peer-reviewed.
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