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Year : 2015  |  Volume : 26  |  Issue : 4  |  Page : 406-410
Quantitative evaluation of apical extrusion of intracanal bacteria using rotary ProTaper, K3XF, twisted and hand K-file system: An ex vivo study

1 Department of Conservative Dentistry and Endodontics, Peoples College of Dental Sciences and Research Centre, Bhopal,Madhya Pradesh, India
2 Department of Prosthodontics, MGM Dental and Medical Hospital, Navi Mumbai, Maharashtra, India
3 Department of Medicine, NSCB Medical College Hospital, Jabalpur, Madhya Pradesh, India

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Date of Submission22-Feb-2014
Date of Decision23-Mar-2014
Date of Acceptance12-Aug-2015
Date of Web Publication20-Oct-2015


Aims: The aim of this study was to evaluate the number of intracanal bacteria extruded apically during root canal preparation using rotary ProTaper, K3XF, twisted, and hand K-file system. Subjects and Methods: Seventy extracted single-rooted human mandibular premolar teeth were used. Access cavities were prepared and the teeth were mounted in glass vials. Root canals were then contaminated with a pure culture of Enterococcus faecalis (ATCC 29212) and incubated at 37°C for 24 h. The contaminated roots were divided into four experimental groups of 15 teeth each and one control group of 10 teeth. Group 1: ProTaper; Group 2: K3XF; Group 3: Twisted file; Group 4: Hand K-file; Group 5: Control group. Bacteria extruded from the apical foramen during instrumentation were collected into vials. The microbiological samples were incubated in culture media for 24 h. Colonies of bacteria were counted and the results were given as number of colony-forming units (CFU)/ml. Statistical Analysis Used: The obtained data were analyzed using the Kruskal–Wallis one-way analysis of variance and Mann–Whitney U-tests. Results: There was a significant difference between the rotary and hand instrumentation system related to the apically extruded intracanal bacteria. Conclusions: Both the rotary and hand instrumentation systems extruded intracanal bacteria through the apical foramen. K3XF file system showed least bacterial extrusion amongst all instrumentation groups.

Keywords: Enterococcus faecalis, K3XF, K-file, ProTaper, twisted

How to cite this article:
Ghogre P, Chourasia HR, Agarwal M, Singh M P, Gurav S, Ghogre R. Quantitative evaluation of apical extrusion of intracanal bacteria using rotary ProTaper, K3XF, twisted and hand K-file system: An ex vivo study. Indian J Dent Res 2015;26:406-10

How to cite this URL:
Ghogre P, Chourasia HR, Agarwal M, Singh M P, Gurav S, Ghogre R. Quantitative evaluation of apical extrusion of intracanal bacteria using rotary ProTaper, K3XF, twisted and hand K-file system: An ex vivo study. Indian J Dent Res [serial online] 2015 [cited 2019 Sep 19];26:406-10. Available from:

Complete preparation of the root canal space is one of the most important stages in endodontic treatment. Despite strict length control of endodontic instruments during root canal preparation the dentin chips, pulpal fragments, necrotic debris, residual pulp tissue, and microorganisms can be forced towards the periapical area. The extruded material referred to as “worm of necrotic debris” has been related to periapical inflammation and post instrumentation flare-ups.[1]

The inter-appointment flare-up is a complication characterized by the development of pain, swelling or both, which commences within a few hours or days after root canal procedures. A true flare-up is of sufficient severity to require an unscheduled visit for emergency treatment.[2] Studies have reported varying frequencies of flare-ups, ranging from 1.4% to 16%.[3],[4],[5],[6],[7],[8],[9],[10]

The causative factors of inter-appointment flare-ups comprise mechanical, chemical and/or microbial injury to the pulp or periradicular tissues.[1],[4] Mechanical and chemical injuries are often associated with iatrogenic factors. However, microbial injury caused by microorganisms and their products that egress from the root canal system to the periradicular tissues is conceivably the major and the most common cause of inter-appointment flare-ups.[1],[11]

In asymptomatic chronic periradicular lesions associated with infected canals, there is a balance between microbial aggression from the infecting canal microbiota and the host defences in the periradicular tissues. During chemomechanical preparation, if the bacteria are extruded apically, the host will face a challenge from a larger number of irritants than initially. Consequently, there will be a transient disruption in the balance between aggression and defence in such a way that the host will mobilize an acute inflammatory response to re-establish the equilibrium.[2] Therefore, it is logical to assume that minimizing the amount of apically extruded debris should minimize postoperative reactions.

All preparation techniques and instruments have been reported to be associated with some amount of apical extrusion of debris, even when preparation is maintained short of the apical terminus.[12],[13],[14],[15] The choice of an instrumentation technique should also be taken into consideration, to limit the periapical extrusion of intracanal materials during treatment.

To date, there has been few literature published on the apical extrusion of intracanal bacteria during root canal instrumentation.[16],[17],[18],[19],[20] Considering the clinical unreliability of the studies measured the amount of apically extruded debris, it seems that counting the number of colony-forming units (CFU) of extruded bacteria during ex vivo evaluations be a more reliable index.

The purpose of this ex vivo study is to compare the number of intracanal bacteria extruded apically after root canal instrumentation with three rotary nickel-titanium instrumentation system using ProTaper, K3XF and twisted files and manual technique with stainless steel K-file.

   Subjects and Methods Top

In this experimental ex vivo study, seventy freshly extracted human single-rooted mandibular premolar teeth with mature apices and curvatures between 0° and 10° were selected. Buccal and proximal radiographs were taken with radiovisiograph to ensure that the teeth were single-rooted and single canal. Teeth with immature apex and calcified canals were excluded. The test microorganism used in this study was Enterococcus faecalis ATCC 29212 strain (MediMark, Europe).

The teeth were cleaned of debris and soft tissue remnants and were stored in physiological saline solution until required. Endodontic access cavities were prepared with Endo-Access Bur (Dentsply, Maillefer, Switzerland) in a high-speed handpiece. Pulp remnants were extirpated with a fine barbed broach, with care taken not to push the broach through the apical foramen. Teeth with apical foramen in which No. 10 K-file should pass through the apex without any resistance and size No. 15 K-file cannot pass through easily were selected.

Glass vials of 5 ml with rubber stoppers were taken, holes were created in the center of the rubber stopper by using heated instrument. The tooth was inserted under pressure into the rubber stopper, which was fixed to the cementoenamel junction by means of cyanoacrylate (Pidilite, India). Two coats of nail varnish were applied to the external surface of all roots to prevent bacterial microleakage through lateral canals or other discontinuities in the cementum. The rubber stopper with the tooth was then fitted into the mouth of the vial. The apical part of the root was suspended within the vial, which acted as a collecting container for apical material extruded through the foramen of the root. The vial was vented with a 26-gauge needle (Dispovan, India) alongside the rubber stopper during insertion to equalize the air pressure inside and outside the vial and used to be an electrode for the electronic working length determination during canal instrumentation [Figure 1]. The entire model system was sterilized in Ethylene Oxide Gas Sterilizer (3M, USA) for a 12-h cycle.
Figure 1: The systematic presentation of test apparatus

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A pure culture of E. faecalis (ATCC 29212) was used to contaminate root canals. A suspension was prepared by adding 1 ml of a pure culture of E. faecalis grown in brain–heart infusion broth for 24 h, to fresh brain–heart infusion broth. Then, McFarland standard number 0.5 was used to evaluate the broth to ensure that the number of bacteria was 1.5 × 108 CFU/ml.

The test apparatus with tooth on the rubber stopper was sealed with Parafilm Tape (Pechiney Plastic Packaging, Chicago, USA) to avoid any contamination of the vials. Each root canal was filled entirely with the E. faecalis suspension using sterile pipettes and a size 10 K-file used to carry the bacteria down the length of the canals. The contaminated root canals were then dried in an incubator at 37°C for 24 h.

Before the experiment, the glass vials were entirely filled with 0.9% normal saline via another needle without opening the vials. A hole was created in the nail varnish that covered the apical foramen using No. 10 K-file. During this procedure, only 1–2 mm of instrument was placed beyond the apical foramen. In this way, a standard size of foramen and apical patency was achieved. The contaminated roots were divided into four experimental groups of 15 teeth each and one control group of 10 teeth.

Single operator under aseptic conditions, carried out the canal preparation and sampling procedures on each specimen under a Class I laminar airflow cabinet to prevent airborne bacterial contamination. Working length determination in all teeth was achieved using DentaPort ZX Electronic Apex Locator (J Morita, USA), until it gives reading 1 mm short of apex. Engine-driven instruments were used with X-Smart Endodontic Motor (Dentsply, Maillefer, Switzerland) using the automatic reverse function mode. One ml of 0.9% normal saline was used per instrument for each root canal as an irrigant because of the different numbers of the files in groups. The irrigant was delivered by disposable plastic syringe with a 26-gauge stainless steel needle that had been placed passively down the canal, up to 3 mm from the apical foramen without binding.

Group I: ProTaper rotary file group

ProTaper rotary files (Dentsply, Maillefer, Switzerland) were used in a crown-down manner according to manufacturer's instructions using a gentle in-and-out motion. Instruments were withdrawn when resistance was felt and then changed into the next instrument. The shaping file (S1) was used first and moved apically to 3–5 mm short of the working length. SX files were then used sequentially until resistance was encountered, followed by S1 and S2 to the working length for shaping the coronal two-thirds of the canal. The apical one-third was finished by using F1, F2, F3 sequentially to the working length.

Group II: K3XF file group

K3XF instruments (SybronEndo, West Collins, CA, USA) were used in a crown-down manner according to the manufacturer's instructions. Instruments were withdrawn when resistance was felt and changed into the next instrument. File sequences were: Size 25, 0.10 and size 25, 0.08 taper was used for coronal flaring until resistance was felt, then sequential use of size 40, 0.06, followed by size 35, 0.06, and then size 30, 0.06 taper was used upto the working length.

Group III: Twisted file group

Twisted file instruments (SybronEndo, West Collins, CA, USA) were used in crown-down manner according to manufacturer's instructions. Instruments were withdrawn when resistance was felt and changed for next instrument. File sequences were: Size 25, 0.08 taper for coronal-third enlargement until the resistance was felt, followed by size 25, 0.06 taper for middle-third to apical-third preparation, and then size 30, 0.06 taper for apical-third preparation.

Group IV: Hand K-file group

The step-back technique was used to prepare the root canal. K-files (Dentsply, Maillefer, Switzerland) were used first with a quarter clockwise rotation followed by a pull-back motion and used repeatedly until the working length was reached. Apical preparation was continued up to size 30 and the step-back was done with a reduction of 1 mm for each file until size 45.

Group V: Control group: No instrumentation done

At the end of canal preparation, 0.1 ml of normal saline was taken from the experimental vials to count the bacteria; the suspension was plated on brain–heart infusion agar at 37°C for 24 h. Colonies of bacteria were counted using a classical bacterial counting technique (Collin's method)[21] and the results were given as number of CFU/ml.

Statistical tests were performed using SPSS (SPSS Inc., Chicago, IL, USA). Data were analyzed statistically using Kruskal–Wallis one-way analysis of variance and Mann–Whitney U-tests. P < 0.05 was set as level of statistical significance.

   Results Top

  • Data regarding the mean number of intracanal bacteria extruded are presented in [Table 1] and [Graph 1]. The results indicated that both the rotary (crown-down) and hand (step-back) instrumentation techniques extruded intracanal bacteria through the apical foramen
  • K-file extruded maximum number of CFU's and showed significant differences (P < 0.05) with ProTaper, K3XF and twisted file [Table 2]
  • Among the rotary instrumentation system, ProTaper file system showed maximum number of extruded CFU's apically with significant differences (P > 0.05)
  • K3XF file system showed least bacterial extrusion amongst all instrumentation groups
  • Though no significant difference was seen between K3XF and twisted rotary file system, K3XF showed less CFU's compared to the twisted file system.
Table 1: The mean number of extruded bacteria for each instrumentation system

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Table 2: Inter-comparison between groups by using Mann-Witney U-test

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   Discussion Top

The aim of the present ex vivo study was to assess the apical extrusion of intracanal bacteria as a result of root canal shaping by using rotary nickel-titanium ProTaper, K3XF, and twisted files and a stainless steel hand K-file.

The teeth were selected carefully according to tooth type, canal size at the working length, and canal curvature. This ensured that the number of apically extruded bacteria was due to the instrumentation technique and not due to the tooth morphology. Pulpal tissue remnants were removed before the preparation and had no influence on the results.[18] A standardized tooth model was used to decrease the number of variables as described by Er et al.[16]

E. faecalis was chosen as the bacteriological marker in this study, because it is a nonfastidious, easy-to-grow aerobic bacterium of significant clinical importance.[22] Common to all techniques were the amount and type of irrigant and the operator. 0.9% of normal saline was used for irrigation in the present study to rule out any reduction in the bacterial count as a result of medication.[23]

In the present study, working length measurements were made 1 mm short of the apical foramen. Martin and Cunningham [24] demonstrated the greater debris extrusion when canals were instrumented at a length where the file was observed to just protrude through the apical foramen versus 1 mm short of the apical foramen. The apical diameter of the master apical instrument in all the groups was standardized at ISO size 30 to avoid any variations in the amount of extruded bacteria due to the size of apical diameter.[25],[26] Therefore, apically extruded debris from root canal specimens could be attributed to the design and technique of the respective instrument used in that particular group.

The results of this study demonstrated that all the instrumentation systems tested, extruded intracanal bacteria apically. However, all engine-driven instruments using the crown-down technique extruded less intracanal bacteria than manual instrumentation using the step-back technique, in correlation with the results of Kustarci et al.[18] and Ghivari et al.[20]

Reddy and Hicks [13] suggested that rotation during instrumentation, in engine-driven techniques, tended to pack dentinal debris into flutes of the instruments and directed them towards the orifice whereas in step-back hand instrumentation technique, the reason for more apical extrusion of bacteria may be that the K-file in the apical one-third acts as a piston, that tends to push debris through the foramen and less space was available to flush out debris coronally.

The results from the present study showed that least amount of bacteria were extruded with K3XF compared to ProTaper and twisted file could be related to its unique design feature of variable pitch which helps to prevent the screwing-in effect of instrument. It has increasing variable helical flute angle from tip to handle which helps to dislodge the dentin chips from working area and carried coronally to the orifice. Blade relief areas controls the depth at which the flutes engage the dentin and protecting the file from over-cutting.[27]

Fayyad et al.[28] reported that twisted files remove less tooth structure than ProTaper file because of its constant taper, highly symmetrical smaller triangular cross-section with regard to flute width and depth resulting in uniform removal of dentin than ProTaper files providing more room for debris to be held in the flutes until they can be wiped.[29]

ProTaper rotary system showed the highest number of intracanal bacteria extruded apically may be due to the progressive taper sequence from tip to coronal portion, along with sharp and larger convex triangular cross-sectional design lead to aggressive cutting of dentinal walls resulting in substantial amount of dentin removal in short period of time.[28] The taper favors the preparation of the apical-third in early stages. The wear occurs early throughout the whole root canal because the instruments reach the working length in the beginning of the preparation, causing greater apical extrusion of bacteria.[30]

Clinically, every effort should be made to limit the periapical extrusion of intracanal material during the root canal treatment. Further in vivo research in this direction could provide more insight into the biologic factors associated with the correlations and consequences of apically extruded bacteria.


The authors wish to thank the Department of Microbiology, Peoples College of Medical Sciences and Research Centre, Bhopal, Madhya Pradesh, India for their involvement and support in the present study.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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[PUBMED]  Medknow Journal  
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Fayyad DM, Elhakim Elgendy AA. Cutting efficiency of twisted versus machined nickel-titanium endodontic files. J Endod 2011;37:1143-6.  Back to cited text no. 28
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Correspondence Address:
Priyanka Ghogre
Department of Conservative Dentistry and Endodontics, Peoples College of Dental Sciences and Research Centre, Bhopal,Madhya Pradesh
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0970-9290.167625

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