Download - Comparison of antibacterial efficacy of intracanal medicaments

18 J INDIAN SOC PEDOD PREVENT DENT | Jan - Mar 2010 | Issue 1 | Vol 28 |

Original article

Comparison of antibacterial efficacy of intracanal medicaments in multiple visit pulpectomies in primary molars-an in vivo study

Lele GS, Subba Reddy VV1

Professor, Department of Pedodontics and Preventive Dentistry, Sinhagad Dental College and Hospital, Pune - 411 043, 1Professor and Head, College of Dental Sciences, Davangere - 577 004, India

Correspondence:Dr. Gauri S Lele, Department of Pedodontics and Preventive Dentistry, Sinhagad Dental College and Hospital, Pune- 411041, India. E-mail: [email protected]

Abstract

Antibacterial efficacy of formocresol, 2% gluteraldehyde and iodine-potassium iodide was assessed by obtaining cultures at consecutive appointments in multiple visit pulpectomies in primary molars. Formocresol and 2% gluteraldehyde were more effective as intracanal medicaments and caused significant reduction in the counts of aerobic and anaerobic microorganisms, thereby supporting the need for placing intracanal medicaments with antibacterial properties, in multiple visit pulpectomies in primary molars.

Key words

Formocresol, intracanal medicaments, iodine-potassium iodide, multiple visit pulpectomies, 2% gluteraldehyde

DOI: 10.4103/0970-4388.60482 PMID: ***

Introduction

In endodontic therapy of primary teeth, due to the tortuous and complex nature of root canals and the change in their morphology with root resorption, ‘biomechanical preparation’ is restricted merely to debridement of the root canals. When this root canal environment is left empty, it allows for microbial recolonization in the pulp space, making it a privileged sanctuary for microorganisms, their by-products and degradation products of both the micro-organisms and pulpal tissue. [1]

Thus, for optimal success, it becomes imperative to place intracanal medicaments within the pulp chamber or canals, which exert their antimicrobial effect by direct contact with the organisms or by way of vapour

action of the volatile components that reaches all the irregularities within the canals. Of these, formocresol, 2% gluteraldehyde and iodine-potassium iodide are reported to have excellent antimicrobial activity and vapor-forming effect with minimal toxicity and tissue irritation[2-4] based on observations made in in vitro studies. Hence the present study was aimed at evaluating and comparing the antibacterial efficacy of these intracanal medicaments in primary teeth multiple visit pulpectomies.

Materials and Methods

The study was carried out in the Departments of Pedodontics and Preventive Dentistry and Oral Pathology and Microbiology, in children requiring multiple visit pulpectomy procedures for their primary teeth.

Selection procedureForty healthy children of both genders were selected from those who reported to the department. Informed consent was obtained from parents showing willingness for their child’s participation. The inclusion criteria were:~ Age of the child was between 4 and 11 years.

19J INDIAN SOC PEDOD PREVENT DENT | Jan - Mar 2010 | Issue 1 | Vol 28 |

Lele and Reddy: Intracanal medicaments in multiple visit pulpectomies in primary molars

~ The child had a primary molar with the one of the following diagnosis:(i) chronic pulpitis, (ii) chronic periapical abscess, (iii) acute exacerbation of chronic pulpitis, (iv) acute exacerbation of chronic periapical abscess.

~ The primary molar had minimal mobility and restorable crown

~ Minimal root resorption or bone resorption, as seen radiographically.

The subjects selected were divided in four groups of 10 children each, wherein for groups I, II and III the intracanal medicaments placed were formocresol, 2% gluteraldehyde and iodine-potassium iodide, respectively. Group IV children formed the control group that received a placebo (distilled water).

A standardized protocol, as described below, was followed for the pulpectomy procedure and collection of bacterial cultures.

Clinical methodFor each tooth in every group at the first visit, local anesthetic was administered and under rubber dam isolation, access cavity preparation was done. The pulp tissue was extirpated from the canals using barbed broaches and K-files,[5-7] canals were irrigated using physiologic saline solution[7-9] and the first sample for culture was obtained using sterile paper points.

On the basis of their radiographic length, the canals were filed, irrigated and dried with paper points. A small cotton pellet, moistened with the respective medicament was placed in the pulp chamber and covered with another plain cotton pellet as an open dressing at the end of the first visit.

At the second visit after 5-7 days, again under rubber dam isolation, the treatment pellet was removed, canals were irrigated, dried with sterile paper points and a second sample was obtained. Following copious irrigation and drying of canals, cotton pellet moistened with the respective medicament was placed in the pulp chamber and it was sealed with zinc oxide-eugenol cement.

At the third visit, after initial drying a third culture of the root canals was taken. The tooth was then irrigated, dried and obturated with zinc oxide-eugenol cement. The same procedure was repeated for all the patients with their respective medicaments.

Collection of bacterial samplesUsing sterile paper points, bacterial samples were

collected at the first, second and third visit (i.e. ‘a’, ‘b’ and ‘c’) from all canals of each tooth included in the study. These were first placed to remove excess of saline solution from canals and were discarded. To obtain the culture, fresh sterile paper points were again inserted in all canals and kept there for a minute so as to absorb as much of exudate along the canal walls as possible [Figure 1]. During this time, the test tube containing fluid thioglycollate medium prepared in the laboratory and sealed with cotton plug was held in readiness. The absorbent points were removed with tweezers and dropped into the tube [Figure 2] and the plug was replaced.[10] The tube was placed back in the stand, after checking that the paper points had dropped into the medium and that it was correctly labeled [Figure 3]. Similarly, at the second visit, after removal of treatment pellet and at the third visit, after removal of zinc oxide-eugenol cement and treatment pellet, copious irrigation of the canals was done and the second and third cultures were obtained. The labeled test tubes were then transported to Department of Oral Pathology and Microbiology.

Laboratory methodIodine-potassium iodide medicament, fluid thioglycollate and blood agar media were prepared in the laboratory.

Laboratory culturing methodUnder strict conditions of asepsis, using a platinum loop, the homogenously dispersed material in the test tubes was streaked on to two petriplates containing blood agar. Of these, the test tube and one petriplate were incubated under aerobic conditions and the other petriplate anaerobically for 48 hours using Gaspak chamber and anaerobic jar.

Figure 1: Collection of bacterial sample



Assessment of microbial growthAssessment of microbial growth in the test tubes and petriplates was carried out after 48 hours The test tubes were assessed for presence or absence of turbidity and were scored as follows [Figures 4 and 5].- Clear (C), i.e. no growth seen- Slight Turbidity (S)

- Moderate Turbidity (M)- Dense Turbidity (D)

The petriplates were assessed under both aerobic and anaerobic conditions for presence or absence of growth; growth if seen was quantified by measuring the number of colony forming units (CFUs) with a magnifying lens [Figures 6 and 7].

Figure 2: Transfer of bacterial sampleFigure 3: Sample in transport medium

Figure 4: Test tube scores: ‘Slight’ and ‘Clear’ Figure 5: Test tube scores: ‘Dense’ and ‘Moderate’

Figure 6: Microbial growth assessment on petriplates: No growth Figure 7: Microbial growth assessment on petriplates: Growth present



The data so collected was grouped, tabulated and statistically analyzed.

Results

Root canal samples from each tooth were obtained at three consecutive visits, wherein ‘a’, ‘b’ and ‘c’ represent the 1st, 2nd and 3rd visit, 5-7 days apart.

Comparison of test tube scores at each visit and at different time intervals for each of the four groups was carried out using the Fischer’s Exact Probability test. For all four groups, the difference in scores from Moderate/Dense to Clear/Slight between the 1st and 2nd visits (a-b) was not statistically significant. However, the difference in scores between 2nd and 3rd visits (b-c) and 1st and 3rd visits (a-c) were found to be statistically significant for groups I and II [Table 1].

Comparison of test tube scores between different groups at each visit was carried out using the Fischer’s Exact Probability test as shown in Table 2 and none of the differences in scores were found to be statistically significant.

Microbial growth in the aerobic culture at different time intervals ‘a’, ‘b’ and ‘c’ for each of the four groups was recorded as shown in Table 3 and these values were analyzed using the Wilcoxon’s Signed Rank test. For group I, the differences in CFUs between the 2nd and 3rd visits (b-c) and 1st and 3rd visits (a-c) were statistically significant with P , 0.01 and 0.05, respectively. For group II value of b-c was statistically significant with P , 0.05. In groups III and IV, none of the differences between time intervals showed any statistical significance.

Inter-group comparison of changes in microbial growth in the aerobic culture for the three time intervals a-b,

Table 1: Comparison of test tube scores at different time intervals

Group Time of assessment Total (Nos)

Test tube score Significance of difference||**

Clear/Slight Total (C*/S†) Moderate/Dense Total (M‡/D§) a-b b-c a-c

I a†† 10 1 1 2 5 3 8 NS Sig. P = 0.011 Sig. P = 0.011

b‡‡ 10 - 2 2 6 2 8

c§§ 10 3 5 8 2 - 2

II a 10 - - - 4 6 10 NS Sig. P = 0.027 Sig. P = 0.005

b 10 - 1 1 3 6 9

c 10 - 6 6 4 - 4

III a 10 - 3 3 4 3 7 NS NS NS

b 10 - 2 2 6 2 8

c 10 1 4 5 5 - 5

IV a 10 1 1 2 3 5 8 NS NS NS

b 10 - 3 3 5 2 7

c 10 1 5 6 4 - 4

C* - Clear; S† - Slight turbidity; M‡ - Moderate turbidity; D§ - Dense turbidity; Fisher’s exact probability test; NS: Not significant; **Comparison has been done for C/S vs. M/D;a†† - First visit; b‡‡ - Second visit; c§§ - Third visit

Table 2: Inter-group comparison of test tube scores at different time intervals

Time of assessment Group Total (Nos.)

Test tube score Significance of difference||**

Clear/Slight Total (C*/S†) Moderate/Dense Total (M‡/D§)

a†† I 10 1 1 2 5 3 8 Not significant

II 10 - - - 4 6 10

III 10 - 3 3 4 3 7

IV 10 1 1 2 3 5 8

b‡‡ I 10 - 2 2 6 2 8 Not significant

II 10 - 1 1 3 6 9

III 10 - 2 2 6 2 8

IV 10 - 3 3 5 2 7

c§§ I 10 3 5 8 2 - 2 Not significant

II 10 - 6 6 4 - 4

III 10 1 4 5 5 - 5

IV 10 1 5 6 4 - 4

C* - Clear; S† - Slight turbidity; M‡ - Moderate turbidity; D§ - Dense turbidity; ||- Fisher’s Exact Probability Test; **- Comparison has been done for C/S vs. M/D; a†† - First visit; b‡‡ - Second visit; c§§ - Third visit



b-c and a-c was carried out using the Fischer’s Exact Probability test and none of the results were found to be statistically significant [Table 4].

Microbial growth in the anaerobic culture at different time intervals ‘a’, ‘b’ and ‘c’, for each of the four groups was recorded as in Table 5 and these values were analyzed statistically using the Wilcoxon’s Signed Rank test. For group I, the differences in CFUs between all three time intervals, i.e., a-b, b-c and a-c were all statistically significant where P , 0.05, 0.01 and 0.05, respectively. For the IInd group, only the difference b-c was significant statistically where P , 0.01. For group III and IV, none of the differences between time intervals showed any statistical significance.

Comparison of changes in microbial growth between different groups in anaerobic culture was carried out using Fisher’s Exact Probability test. For the time interval a-b, statistically significant difference was observed between groups I and II, II and III and II and IV. For the time interval b-c, statistically significant difference was observed between groups I and II and groups II

and III. For the time interval a-c, statistically significant difference was observed only between groups II and III [Table 6].

Discussion

The treatment procedure in this study was carried out in three visits with a period of 5-7 days in between for standardization.[11] At the 3rd visit only if the tooth was asymptomatic, it was obturated with Zinc oxide-eugenol as this material has an antibacterial effect, is resorbable, easily available and economical.[5,6,11-13]

The bacteriologic samples were collected with the help of sterile paper points as per the sampling technique described by Grossman,[10] in which the transfer from the canals to the test tubes took a few seconds and most strains recovered from infected root pulps are known to tolerate substantial oxygen exposure. The time that elapsed between sampling of the root canals and culturing the same in the laboratory was always less than 8 hours since this period is considered adequate for maintaining the viability of the microorganisms.[14]

Table 4: Inter group comparison of changes in microbial growth in aerobic culture

Time interval

Changes in CFUs

I II III IV Difference between groups*No. of cases % No. of cases % No. of cases % No. of cases %

a-b Decreased 4 40 2 20 3 30 3 30 Not significant

Increased 4 40 7 70 4 40 6 60

No growth 2 20 1 10 3 30 1 10

b-c Decreased 7 70 7 70 5 50 7 70 Not significant

Increased - - 2 20 2 20 1 10

No growth 3 30 1 10 3 30 2 20

a-c Decreased 5 50 7 70 4 40 4 40 Not significant

Increased 1 10 2 20 3 30 4 40

No growth 4 40 1 10 3 30 2 20

*Fisher’s exact probability test (Statistical significance was tested on number of cases showing differences in colony forming units) P , 0.05, P , 0.01: Significant (Wilcoxon’s Signed Rank test)NS: Not significant

Table 3: Comparison of microbial growth (CFUs/0.005 ml) at different time intervals in aerobic culture

Group Particulars of CFUs Time of assessment Difference between time intervals*

a b c a-b b-c a-c

I Range 0-28 0-17 0-6 20.3 (NS) 7.7 (P , 0.01) 7.4 (P , 0.05)

Mean 8.0 8.3 0.6

II Range 0-210 0-155 0-70 214.2 (NS) 40.2 (P , 0.05) 26.0 (NS)

Mean 44.1 58.3 18.1

III Range 0-108 0-180 0-60 218.8 (NS) 25.2 (NS) 6.4 (NS)

Mean 20.3 39.1 13.9

IV Range 0-227 0-154 0-114 21.7 (NS) 24.1 (NS) 22.4 (NS)

Mean 49.3 51.0 26.9

*Statistical significance was tested on number of cases showing differences in CFUs P , 0.05, P , 0.01: Significant (Wilcoxon’s Signed Rank test) NS: Not Significant



Statistically significant difference in the test tube scores between the 2nd to 3rd and 1st to 3rd visit was observed only with formocresol and 2% gluteraldehyde, which proves the longer duration of effectiveness of these medicaments under both open and sealed conditions of the pulp chamber.

The mean CFUs in aerobic culture for all the four groups showed an increase in count from ‘a’ to ‘b’ probably because the medicaments were placed as open dressings and thus, micro-organisms from saliva could have freely diffused through the cotton into the root canal system. Naidorf (1985) had also suggested that the introduction of oxygen to the root canal system and periapical area during access preparation might facilitate an aerobic condition, thus encouraging aerobic micro-organisms to multiply preferentially. Subsequently, when the medicaments were sealed with a temporary restoration, statistically significant difference in the reduction of counts between the 2nd and 3rd visits was seen in the formocresol and 2% gluteraldehyde groups, which proves the efficacy of these medicaments.

Comparison of CFUs in anaerobic cultures also showed an increase in the mean values from ‘a’ to ‘b’ with

formocresol, iodine-potassium iodide and the placebo group as against group II, i.e., 2% gluteraldehyde. The increase seen in other groups could be explained on the basis that in the canals infected by saliva, facultative anaerobic bacteria are present in greater proportion than anaerobic bacteria. Subsequently, the relative proportion of anaerobic bacterial strains and cells increases with time. Sundqvist[15] mentioned that anaerobic infections occur when there is a compromised blood supply or antecedent infection by aerobic bacteria, both of which produce a milieu with a low oxygen reduction potential (Eh).

After placing a closed dressing, a decrease in the mean CFUs was observed in all the four groups between 2nd and 3rd visits (b-c). Of these, only with formocresol and 2% gluteraldehyde, this difference was statistically significant i.e., these two medicaments showed better antibacterial efficacy compared to iodine-potassium iodide and similar findings were observed by Collet et al.[16] and O’Hara et al.[17-19] In the present study, iodine-potassium iodide did not show statistically significant difference in the mean CFUs between 2nd and 3rd visits. Kamran E. Safavi, Larz Spangberg and K. Langeland[20] similarly found iodine-potassium iodide to have longer

Table 6: Inter-group comparison of changes in microbial growth in anaerobic culture

Time interval Changes in CFUs

I II III IV Difference between groups*

No. of cases % No. of cases % No. of cases % No. of cases % Groups Significance*

a-b Decreased 1 10 8 80 1 10 2 20 I vs. II P 5 0.007

Increased 7 70 2 20 9 90 5 50 II vs.III P 5 0.003

No growth 2 20 - - - - 3 30 II vs IV P 5 0.048

b-c Decreased 8 80 9 90 5 50 5 50 I vs II P 5 0.029

Increased - - 1 10 5 50 2 20 II vs III P 5 0.065

No growth 2 20 - - - - 3 30

a-c Decreased 3 30 7 70 2 20 4 40 II vs III P 5 0.047

Increased 1 10 3 30 7 70 3 30

No growth 6 60 - - 1 10 3 30

Other comparisons are not significant. *Fisher’s exact probability test. P , 0.05, P , 0.01: Significant

Table 5: Comparison of microbial growth (CFUs/0.005 ml) at different time intervals in anaerobic culture

Group Particulars of CFUs Time of assessment Difference between time intervals*

a b c a-b b-c a-c

I Range 0-141 0-173 0-25 254.4 (P , 0.05) 3.0 (P , 0.01) 22.9 (P , 0.05)

Mean 25.9 80.3 3.0

II Range 9-110 34-145 10-72 1.1 (NS) 28.6 (P , 0.01) 29.7 (NS)

Mean 63.6 62.5 33.9

III Range 0-84 0-58 0-54 217.1 (NS) 2.0 (NS) 215.1 (NS)

Mean 14.5 31.6 29.6

IV Range 0-270 0-360 0-158 229.3 (NS) 26.4 (NS) 2.9 (NS)

Mean 53.2 82.5 56.1

*Statistical significance was tested on number of cases showing differences in colony forming units. P , 0.05, P , 0.01: Significant (Wilcoxon’s Signed Rank test) NS: Not significant



bacteriocidal effect but its duration of antimicrobial efficacy was very short. Conversely, when Ellerbruch and Murphy[21] checked antimicrobial activity of medicaments based on the vapor-forming activity after 24 hours, iodine-potassium iodide was found to be a better medicament than 2% gluteraldehyde. This difference may be due to the fact that iodine-potassium iodide is mainly bactericidal, while 2% gluteraldehyde is bacteriostatic and also that the efficacy was assessed for the period of 24 hours whereas in this study, it was assessed after 5-7 days.

For group IV, the placebo group, in which distilled water was used as control, the reduction in counts observed was mainly attributed to extirpation and irrigation carried out during the procedure.

Conclusions

Based on the present study, in case of multiple visit pulpectomies in vivo:• Theantibacterialefficacyof formocresoland2%

gluteraldehyde was higher when compared to iodine-potassium iodide;

• Use of formocresol as intracanalmedicamentresulted in statistically significant reduction (P , 0.01) in counts of aerobic and anaerobic microorganisms after placement of closed dressing.

• Iodine-potassiumiodidedidnotshowanysignificantantibacterial efficacy against both aerobes and anaerobes.

References

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15. Sundqvist G. Ecology of the root canal flora. J Endodon 1992;18:427-30.

16. Masillamoni CR, Kettering JD, Torabinejad M. The bio compatibility of some root canal medicaments and irrigants. Int J Endod 1981;14:115-20.

17. Hill SD, Berry CW, Seale NS, Kaga M. Comparison of antimicrobial and cytotoxic effects of glutaraldehyde and formocresol. Oral Surg Oral Med Oral Pathol 1991;71:89-95.

18. Marquez-Aviles JR, Miller J. Beechwood-creosote, formocresol, bactericidal and irritant properties for endodontia. Brit Dent J 1980;149:105-8.

19. O’hara P, Torabinejad M, Kettering JD. Antibacterial effects of various endodontic medicaments on selected anaerobic bacteria’. J Endodon 1993;19:498-500.

20. Safavi KE, Spangberg LS, Langeland K. Root canal dentinal tubule disinfection. J Endodon 1990;16:207-10.

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Source of Support: Nil, Conflict of Interest: Nil

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