|Year : 2017 | Volume
| Issue : 4 | Page : 296-300
Comparison of Candida species isolated from children with and without early childhood caries: A descriptive cross-sectional study
MS Beena1, Faizal C Peedikayil1, MB GufranAfmed2, TP Chandru1, K Soni1, N Dhanesh1
1 Department of Pediatric and Preventive Dentistry, Kannur Dental College, Kannur, Kerala, India
2 Department of Microbiology, Kannur Medical College, Kannur, Kerala, India
|Date of Web Publication||15-Sep-2017|
M S Beena
Department of Pediatric and Preventive Dentistry, Kannur Dental College, Anjarakandy, Kannur, Kerala
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Early childhood caries (ECC) is characterized by the presence of one or more decayed, missing (due to caries), or filled teeth surfaces in any primary tooth, in a child below 6 years of age. Although ECC is primarily associated with high levels of maternal Streptococcus mutans, there has been an increased interest in finding the relationship between oral fungal flora and dental caries. Objective: The aim of the study is to identify and characterize the Candida species and to compare the candidal isolates in children with ECC and without ECC. Materials and Methods: The study was conducted on children below 6 years of age, who were categorized into ECC and non-ECC groups of fifty children each. Samples were collected using sterile cotton swabs and were inoculated on Sabouraud's Dextrose Agar and incubated at 37°C for 24 h. Candidal colonies were isolated, species identified and virulence factors tested for both ECC and non-ECC groups. Results: The candidal carriage among the ECC children was found to be 84%, which was significantly higher than the non-ECC group of 24%. Candida albicans and non-albicans Candida (NAC) were isolated in both ECC and non-ECC groups. Phospholipase production was significantly high in ECC group whereas hemolysin production and germ tube formation showed no significant difference between the two groups. Conclusion: A significant correlation was found between the presence of Candida and ECC. NAC also plays an important role in the development of ECC. The virulence factors such as phospholipase may be responsible for the pathogenicity of Candida in the development of ECC.
Keywords: Candida species, dental plaque, early childhood caries, speciation
|How to cite this article:|
Beena M S, Peedikayil FC, GufranAfmed M B, Chandru T P, Soni K, Dhanesh N. Comparison of Candida species isolated from children with and without early childhood caries: A descriptive cross-sectional study. J Indian Soc Pedod Prev Dent 2017;35:296-300
|How to cite this URL:|
Beena M S, Peedikayil FC, GufranAfmed M B, Chandru T P, Soni K, Dhanesh N. Comparison of Candida species isolated from children with and without early childhood caries: A descriptive cross-sectional study. J Indian Soc Pedod Prev Dent [serial online] 2017 [cited 2020 Nov 23];35:296-300. Available from: https://www.jisppd.com/text.asp?2017/35/4/296/214916
| Introduction|| |
Early childhood caries (ECC) is a destructive form of tooth decay that afflicts young children. It is considered to be a common chronic childhood disease that affects the normal health and well-being of the child. ECC has a very rapid and widespread progression which makes its prevention and management a challenge. The etiology of ECC is multifactorial. Although mutans streptococci is frequently associated with the carious lesion, the presence of other microorganisms such as Candida albicans is found to enhance its pathogenicity.,
Candida is a normal commensal in the oral cavity of healthy individuals. The percentage of Candidal colonization ranges from 20% to 40% in healthy individuals, whereas it is about 60% in immunocompromised people. There are many species of Candida, of which C. albicans is the most prevalent one recovered from the oral cavity. However, there is due attention needed for non-albicans Candida (NAC) due to their enhanced resistance to certain antifungal agents. Identification of infecting strains of Candida is important because isolates of Candida species differ widely in their ability to cause infection  and their susceptibility to antifungal agents. They also express various virulence factors responsible for its pathogenicity.
At present, literature showing the role of NAC in the development of ECC and the virulence of these organisms are few. Hence, the present study was undertaken to characterize different Candida species in ECC and to compare its existence as a normal commensal in the oral cavity of non-ECC individuals.
| Materials and Methods|| |
The present study was carried out on children who came to the Department of Pediatric and Preventive Dentistry, Kannur Dental College. The sample for the study consisted of 100 children who were below six years of age. The research protocol was reviewed and approved by the Institutional Ethical Committee and Review Board, and the study was in compliance with the Code of Ethics (Declaration of Helsinki). The study was explained in detail to the parents, and an informed consent was obtained. The categorization of ECC group consisted of children having one or more decayed (cavitated or noncavitated), missing (due to caries), or filled tooth surfaces in any primary tooth who were randomly selected. Non-ECC group consisted of children who were caries free. The exclusion criteria consisted of usage of systemic or topical antibiotics in the past one month, oral candidiasis, systemic, or congenital diseases.
Samples collected from the buccal, lingual, proximal, and cervical portions of the tooth using sterile cotton swabs were immediately transferred to the laboratory for microbiological analysis. The samples were inoculated for culture on Sabouraud's Dextrose Agar (SDA) plates supplemented with 1% chloramphenicol and incubated at 37°C for 24–72 h. Isolates were identified by colony morphology on SDA plates. Culture is said to be negative if there is no growth even after 72 hours of incubation. Isolates were speciated based on the conventional methods of germ tube testing and Dalmau plate culture on corn meal agar [Figure 1]. The samples were also streaked on HiCrome Agar media (Himedia, India) and incubated at 37°C for 24 h. Colonies were identified depending on their color and pattern of growth. The virulence markers such as hemolysin, phospholipase, and germ tube formation were tested on all Candida isolates.
The data were statistically analyzed using “Chi-square test,” “Mann–Whitney U-test,” and “Fisher exact test” in SPSS Version 17.0 (SPSS Inc. Chicago USA). The results were considered statistically significant at P ≤ 0.05.
| Results|| |
The growth of Candida on SDA appeared in 1 to 2 days as creamy, smooth, convex pasty colonies with a moldy odor [Figure 2]. When species isolation was done by conventional methods, C. albicans showed the presence of germ tube formation. In Dalmau plate culture, chlamydospore formation was observed under microscope for C. albicans [Figure 3]. When cultured on HiCrome agar, colonies of C. albicans were seen as light green-colored smooth colonies; Candida tropicalis appeared as metallic blue-colored raised colonies. Candida glabrata colonies appeared as cream smooth colonies, while Candida krusei appeared as purple fuzzy colonies [Figure 4].
|Figure 3: Chlamydospore formation of Candida albicans - microscopic view|
Click here to view
Candidal carriage among the ECC group was found to be 84%, whereas 24% in non-ECC group is statistically significant [Figure 5]. Whereas the difference in proportion of C. albicans and NAC among ECC and non-ECC groups was found to be statistically not significant.
The species distribution of Candida found in the ECC group and non-ECC group is shown in [Table 1]. The difference of the distribution of the species among the ECC and non-ECC is statistically found to be nonsignificant (P > 0.05).
|Table 1: Comparison of species distribution of Candida among the early childhood caries and nonearly childhood caries groups|
Click here to view
Virulence factors such as hemolysin, phospholipase, and germ tube formation were expressed by the Candida species isolated in the study. Phospholipase production showed a statistically significant difference among the ECC and non-ECC groups. The other virulence factors tested such as hemolysin and germ tube formation showed no statistically significant difference among the groups as seen in [Table 2].
|Table 2: Comparison of virulence factors of Candida among early childhood caries and nonearly childhood caries group|
Click here to view
| Discussion|| |
The microbiology of dental plaque resulting in ECC has been of great interest to dental researchers. Various studies have shown Candida to have a role in microbial adherence to dental surfaces in coaggregation with Streptococcus mutans. Although C. albicans is widely known as an etiological agent for dental caries, the various other species of Candida and its virulence factors responsible for the cariogenicity are not clearly understood. Hence, the purpose of the study was to isolate and characterize Candida species in the ECC group and to compare it with the non-ECC group.
The present study showed a high candidal carriage in ECC individuals when compared to non-ECC group. Candida exerts a synergistic action along with mutans streptococci enhancing its cariogenicity. The presence of Candida has been shown to increase the adherence of mutans streptococci to the oral biofilm and carious tooth substance in vitro. Other factors such as adherence of Candida to saliva proteins and S. mutans, its acid-producing capability, its ability to penetrate into dentinal canals, and its enzymatic activity to degrade collagen may also be responsible for its cariogenic ability , However, some studies could not find a correlation between Candida and dental caries,,, and it is attributed to factors such as difference in saliva rate, composition, buffering capacity, and the difference in the technical methods employed for yeast detection by those authors.
C. albicans showed the highest prevalence among the candidal isolates. This may be due to its capacity to form germ tubes, facilitating adhesion. Its pathogenic potential can be attributed to factors such as molecules which mediate adhesion to and invasion into host cells, the secretion of hydrolases, the yeast-to-hypha transition, contact sensing and thigmotropism, biofilm formation, phenotypic switching, and a range of fitness attributes.
However, the presence of significant amount of NAC such as C. glabrata, C. guilliermondii, C. krusei, C. kyfer, C. tropicalis could be appreciated in our study. There is a growing concern about the presence of NAC as they exhibit a higher azole resistance when compared to C. albicans. Butola et al. have reported a higher production of virulence factors such as phospholipase among NAC than C. albicans. However, the cariogenic potential of NAC is yet to be understood clearly. Makihira et al. reported that the adhesion of C. albicans to intact and denatured type I collagen was significantly greater than those of other species and suggested that C. albicans possessed the ability to adhere specifically to extracellular matrix as compared to other Candida species.
Virulence of Candida species is a significant factor that contributes to its colonization, pathogenicity, and infection of tissues. In the present study, virulence factors such as formation of germ tubes, hyphae, hydrolytic enzymes such as phospholipases and hemolysin were expressed by the Candida species. The candidal isolates of ECC group showed phospholipase production whereas it was absent in the non-ECC group. Phospholipase acts by degrading the cell membrane of tissues and epithelial cells, and its presence among the isolates of ECC group alone may suggest its role in the pathogenicity. Hemolysin allows Candida to acquire iron from host tissues, which then is used by the fungus for metabolism, growth, and invasion during host infection.
| Conclusion|| |
The current study substantiates previous evidence of the association between candidal carriage and dental caries. In addition to C. albicans, NAC also can be considered to be a major pathogen associated with ECC. Various virulence factors such as phospholipase, hemolysin, and germ tube formation seem to affect its pathogenicity. However, further studies emphasizing the various other virulence factors such as protinase production and phenotypic switching responsible for the virulence of the NAC need to be researched.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Klinke T, Kneist S, de Soet JJ, Kuhlisch E, Mauersberger S, Forster A, et al.
Acid production by oral strains of Candida albicans
and lactobacilli. Caries Res 2009;43:83-91.
Barbieri DD, Vicente VA, Fraiz FC, Lavoranti OJ, Svidzinski TI, Pinheiro RL. Analysis of the in vitro
adherence of Streptococcus mutans
and Candida albicans
. Braz J Microbiol 2007;38:624-31. Available from: http://www.scielo.br/scielo.php
. [Last accessed on 2017 Jul 14].
Srivastava B, Bhatia HP, Chaudhary V, Aggarwal A, Kumar Singh A, Gupta N. Comparative evaluation of oral Candida albicans
carriage in children with and without dental caries: A microbiological in vivo
study. Int J Clin Pediatr Dent 2012;5:108-12.
Marchant S, Brailsford SR, Twomey AC, Roberts GJ, Beighton D. The predominant microflora of nursing caries lesions. Caries Res 2001;35:397-406.
Thomas A, Mhambrey S, Chokshi K, Chokshi A, Jana S, Thakur S, et al.
Association of oral Candida albicans
with severe early childhood caries – A pilot study. J Clin Diagn Res 2016;10:ZC109-12.
Fragkou S, Balasouli C, Tsuzukibashi O, Argyropoulou A, Menexes G, Kotsanos N, et al. Streptococcus mutans
, Streptococcus sobrinus
and Candida albicans
in oral samples from caries-free and caries-active children. Eur Arch Paediatr Dent 2016;17:367-75.
Maijala M, Rautemaa R, Järvensivu A, Richardson M, Salo T, Tjäderhane L. Candida albicans
does not invade carious human dentine. Oral Dis 2007;13:279-84.
Peretz B, Mazor Y, Dagon N, Bar-Ness Greenstein R. Candida, mutans streptococci, oral hygiene and caries in children. J Clin Pediatr Dent 2011;36:185-8.
Ratson T, Greenstein RB, Mazor Y, Peretz B. Salivary Candida
, caries and Candida
in toothbrushes. J Clin Pediatr Dent 2012;37:167-70.
Nikawa H, Yamashiro H, Makihira S, Nishimura M, Egusa H, Furukawa M, et al
. In vitro
cariogenic potential of Candida albicans
. Mycoses 2003;46:471-8.
Mayer FL, Wilson D, Hube B. Candida albicans
pathogenicity mechanisms. Virulence 2013;4:119-28.
Deorukhkar SC, Santosh S. Species distribution and antifungal susceptibility profile of Candida species isolated from bloodstream infections. J Evol Med Dent Sci 2012;1:241-3.
Butola R, Agwan V, Thakuria B, Madan M. A comparative study of virulence factors in clinical isolates of Candida
species. Int J Curr Microbiol Appl Sci 2015;4:716-22.
Makihira S, Nikawa H, Tamagami M, Hamada T, Nishimura H, Ishida K, et al.
Bacterial and Candida
adhesion to intact and denatured collagen in vitro
. Mycoses 2002;45:389-92.
Sitheeque MA, Samaranayake LP. Chronic hyperplastic candidosis/candidiasis (candidal leukoplakia). Crit Rev Oral Biol Med 2003;14:253-67.
Udayalaxmi J, Shenoy N. Comparison between biofilm production, phospholipase and haemolytic activity of different species of Candida
isolated from dental caries lesions in children. J Clin Diagn Res 2016;10:DC21-3.
Almeida RS, Wilson D, Hube B. Candida albicans
iron acquisition within the host. FEMS Yeast Res 2009;9:1000-12.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]
[Table 1], [Table 2]