Medicine in Microecology 15 (2023) 100076

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Medicine in Microecology
journal homepage: www.journals.elsevier.com/medicine-in-microecology

Fungal composition in saliva and plaque in children with caries: Differences

and influencing factors
Meixiang Yin a, b, 1, Yang You a, 1, Xiao Zheng a, Qiuying Liang a, Buling Wu a, *, Wen'an Xu a, **
a
Department of Pediatric Dentistry, Shenzhen Stomatology Hospital (Pingshan), Southern Medical University, 143 Dongzong Road, Pingshan District, Shenzhen, 518118,
China
b
Department of Stomatology, Shenzhen Samii Medical Center, No.1 Jinniu West Road, Pingshan District, Shenzhen, 518118, China

A R T I C L E I N F O A B S T R A C T

Keywords: Caries is a dental disease that can affect oral and psychological health and has a high incidence in children. The
Mycobiome role of fungal flora in childhood caries has not been fully described. In this study, we aimed to investigate the
Dental caries fungal composition differences and the influencing factors in unstimulated saliva (S) and supragingival plaque (P)
Internal transcribed spacer
samples in childhood caries. S and P samples were collected from 63 children with caries. The ITS2 region in the
Saliva
Plaque
fungal genome was then amplified and sequenced. Subsequently, we quantified and analyzed the fungal com-
positions in the samples. Cryptococcus was the most abundant genus in the S and P subgroups. The relative
abundances of Cryptococcus and Wickerhamomyces significantly differed between the S and P subgroups (p <
0.05). Significant differences were also observed in alpha and beta diversities between the two subgroups (p <
0.05). However, no significant differences were observed between the mycobiome of the SFe and SMa subgroups
or the PFe and PMa subgroups (p > 0.05). Conversely, species differences were detected between the SDD and
SMD subgroups (p < 0.05) but not in the PDD and PPD subgroups (p > 0.05). Our findings revealed significant
differences in the mycobiome of unstimulated saliva and supragingival plaque. Dentition period and oral hygiene
behaviors may have affected these differences.

revealed the presence of fungal communities in children with and

1. Introduction
Dental caries is a microbially mediated multifactorial disease of hard
tissues [1]. The incidence of dental caries is high among childre, More-
over, a lack of timely treatment and prevention can affect chewing
function and psychological health, thereby causing a serious burden to
society [2].
Oral bacteria play an important role in the etiology of dental caries
[3–7]. For example, researchers have detected Streptococcus mutans in
childhood caries and conducted comprehensive research on its patho-
genic mechanisms [8]. Previous studies on oral microorganisms have
mainly focused on the present bacterial groups; however, the mycobiome
community and its relevance in oral health and disease has been mini-
mally investigated. Ghannoum et al. [9] identified the presence of fungi
in the oral cavity of 20 healthy adults, and for the first time, to charac-
terized the “basal fungal flora” in healthy people. Fechney et al. [10]
without dental caries. Among these, Candida albicans has been widely
studied, and an increasing amount of evidence suggests its associated
with dental caries [11–13]. In addition, several studies have attempted to
investigate the effects of gender, age, and geographic location on the
composition of oral fungi [9,14].
Typically, unstimulated saliva and supragingival plaque are consid-
ered suitable for studying the oral microbiome [15–17]. Many studies
have focused on fungal communities in saliva or plaque samples sepa-
rately [18–20], whereas the relationship and phenotypic effects between
salivary and supragingival mycobiomes remain poorly understood.
Recently, high-throughput sequencing technology has promoted the
discovery of highly diverse oral microbial communities [17,21–23]. Most
ITS-based fungal studies have focused on the ITS1 or ITS2 subregion.
Compared with the ITS1 subregion, ITS2 is more commonly used owing
to its small length variation, more common primer sites, and less classi-
fication bias [24].

* Corresponding author.
** Corresponding author.
E-mail addresses: Yinmeixiang782130@163.com (M. Yin), youyang_claire@outlook.com (Y. You), ixzheng@163.com (X. Zheng), 313495715@qq.com (Q. Liang),
bulingwu@smu.edu.cn (B. Wu), xu_wenan@smu.edu.cn (W. Xu).
1
These authors contributed equally to this work and should be considered co-first authors.

https://doi.org/10.1016/j.medmic.2023.100076
Received 20 December 2022; Received in revised form 16 January 2023; Accepted 19 January 2023
Available online 20 January 2023
2590-0978/© 2023 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-
nc-nd/4.0/).
M. Yin et al.
Abbreviations
internal transcribed spacer (ITS)
International Caries Detection and Assessment System
II (ICDASII)
no significance (ns)
principal coordinate analysis (PCoA)
Severe early childhood caries (S-ECC)
Decayed teeth (DT)
Decayed teeth surfaces (DS)
unstimulated saliva (S)
supragingival plaque (P)
Male in S (SMa)
Male in P (PMa)
Female in S (SFe)
Female in P (PFe)
Deciduous dentition in S (SDD)
Deciduous dentition in P (PDD)
Mixed dentition in S (SMD)
Mixed dentition in P (PMD)
In this study, we used high-throughput sequencing technology to
target the ITS2 region of fungi in the S and P samples from 63 children
with caries. We aimed to explore the relationship between the myco-
biomes of the S and P samples, as well as investigate the effects of gender,
dentition stage, decayed teeth (DT), decayed teeth surfaces (DS), and oral
hygiene behaviors on the fungal composition of both samples. The
findings of this study will serve as a foundation for future investigations
on the relationship between oral fungal communities and caries in chil-
dren, and how to select appropriate research samples.
2. Material and method
2.1. Population study
This study was approved by the Ethics Committee of the Shenzhen
Samii Medical Center (Ethics Number: SSMC-202011-R01) in Shenzhen,
China. Sixty-three children with caries aged 2–9 years, who were patients
of the dental outpatient clinic of Shenzhen Samii Medical Center, were
encouraged to participate in this study. The guardians signed the
informed consent form. Individuals who had systemic diseases, used
antibiotics within three months, suffered from periodontitis or oral
mucosal disease, received fluoridation within three months, used fixed
orthodontic devices, or could not cooperate with clinical examination
and sample collection were excluded. The general and lifestyle charac-
teristics and oral health of children were examined using questionnaires
answered by their guardians. The gender, dentition stage, DT, and DS
data of each participant are listed in Supplementary Table 1.
2.2. Clinical examination
Two experienced dentists performed the oral examinations according
to the International Caries Detection and Assessment System II (ICDASII)
standard [25]. Examiners were rigorously trained and regularly recali-
brated at the measurement point to reduce errors. Standard oral scopes,
headlights, and community periodontal index (CPI) probes were used in
the examinations.
2.3. Sample collection
The children were informed to stop brushing the night before sam-
pling and refrain from eating and drinking 2 h before sampling. Sampling
was generally scheduled at 2:00–5:30 p.m. The spit method was
2
Medicine in Microecology 15 (2023) 100076
performed to collect unstimulated saliva samples, while sterile cotton
swabs were used to collect the supragingival plaque samples from all
tooth surfaces. A total of 63 S and 63 P samples were collected. All
samples were collected in 1.5 ml sterile sampling tube (Beyotime,
FCT217), placed on dry ice after collection, and immediately frozen at

80  C for further analysis. The samples were grouped according to
sampling type, gender, and dental period. The first grouping scheme was
S and P subgroups. The two groups were further categorized by gender,
as follows: male in S (SMa), male in P (PMa), female in S (SFe), and fe-
male in P (PFe). In addition, the S and P subgroups were divided ac-
cording to dentition period as follows: deciduous dentition in S (SDD),
deciduous dentition in P (PDD), mixed dentition in S (SMD), and mixed
dentition in P (PMD).
2.4. DNA extraction and ITS2 rRNA amplicon sequencing and analysis
Total DNA was extracted using a bacterial DNA extraction mini kit
(Mabio, Guangzhou, China), and the DNA quality and concentration
were detected using a Thermo NanoDrop One (Thermo Fisher Scientific,
MA, USA). Briefly, the bead milling step was performed, and the lysis
time was extended by 15 s to dissolve the thick cell wall of fungi and
improve the success rate of DNA extraction. The ITS2 region of fungi was
amplified using PCR. The forward and reverse primer sequences were
GCATCGATGAAGAACGCAGC and TCCTCCGCTTATTGATATGC, respec-
tively. Primers were synthesized by Invitrogen (Carlsbad, CA).
For amplicon sequencing, PCR products were mixed in an equal
density ratio according to the Gene Tools Analysis software (version
4.03.05.0; SynGene). The pooled PCR products were purified using an E.
Z.N.A. Gel Extraction Kit (Omega, USA). The NEBNext® Ultra™ II DNA
Library Prep Kit for Illumina® (New England Biolabs, USA) was used for
library construction. The constructed amplicon library was sequenced for
PE250 using the Illumina Nova 6000 platform (Guangdong Magigene
Biotechnology Co., Ltd., Guangzhou, China).
2.5. Mycobiome analysis
Raw data were filtered to remove primers and junction contamina-
tion, the clean reads were sorted into tags based on the overlap in the
reads, and sequence splicing was performed using Fast Length Adjust-
ment of Short reads (FLASH, v1.2.11). The tags were clustered into OTUs
and compared with the ITS chimera reference database UNITE (v20147
03). Species annotation was performed based on the OTU and annotation
results, species complexity analysis, and species difference analysis be-
tween groups.
2.6. Statistical analysis
Species compositions are presented in bar charts and crop species
abundance histograms for each sample at the phylum, genus, and species
levels. All species with abundance below 0.5% in all samples and no
annotation were categorized as Others. R software (v3.4.1) was used.
Independent sample t-test was used to compare the mean age, DT, and
DS between the male and female groups, and Chi-Square was used to
evaluate sample population differences. The top 10 species based on
relative abundance were compared using the Wilcoxon rank-sum test.
Significant differences were set as p-value＜0.0001, marked with “****“,
0.0001  p-value <0.001, marked with “***“, 0.001  p-value <0.01,
marked with “**“; 0.01  p-value <0.05, marked with “*“, and p-value＞
0.05, marked with no significance (ns). False discovery rate (FDR) values
were calculated for the p-values. The results were screened for significant
differences according to p < 0.05 and FDR <0.05.
Alpha diversity was measured using the Wilcoxon Rank-Sum Test
performed in the R software (v3.2.1). To compare the overall variability
in the microbial community structure between the two groups, beta di-
versity was analyzed using principal coordinate analysis (PCoA) and a
box chart in QIIME (v1.80) using unweighted UniFrac distances.
M. Yin et al. Medicine in Microecology 15 (2023) 100076

Functional annotation was performed using the FUNGuild software. 3.2. Differential analysis between unstimulated saliva and supragingival
Redundancy analysis (RDA) was performed using the vegan package [26] plaque
in R (http://cran.r-project.org/) with the normalized OTU abundances of
fungal data. The length of the lines represented the magnitude of each 3.2.1. Fungal community composition and diversity
factor ‘s relative influence. Angles between lines <90 , indicated factors In both the S and P subgroups, 7, 8, and 10 taxa were detected at the
with the same influence, angles >90 , indicated factors with opposite phylum, genus, and species level, respectively; the remaining taxa were
influence. classified as Others (Fig. 1A).
In the S subgroup, at the phylum level, Basidiomycota (48.7%) was the
3. Results most abundant, followed by Ascomycota (28.6%). At the genus level,
Cryptococcus (47.9%) was the most abundant, followed by Wick-
3.1. Fundamental analysis erhamomyces (8.5%), Candida (7.7%), Cladosporium (3.4%), Aspergillus
(1.8%), and Hyphopichia (1.4%). At the species level, Cryptococcus_longus
A total of 63 children (36 males and 27 females; 24 with deciduous (47.9%) was the most abundant, followed by Wickerhamomyces_anomalus
dentition and 39 with mixed dentition) with a mean age of 80.27  21.06 (8.5%), Candida_albicans (1.7%), Candida_intermedia (2.2%), Candida_-
months (ranging from 2 to 9 years) were included in this study. The sake (2.2%), Saccharomyces_cerevisiae (1.5%), and Hyphopichia_burtonii
number of DT in each of the 63 children ranged from 1 to 20, with a mean (1.4%).
value  SD of 7.41  4.27, and the number of DS ranged from 1 to 48, In the P subgroups, at the phylum level, Ascomycota (30.3%) was the
with a mean value  SD of 13.51  10.82. No significant differences were most abundant, followed by Basidiomycota (26.7%). At the genus level,
observed in the number of samples of deciduous and mixed dentition Cryptococcus (25.9%) was the most abundant, followed by Candida
between the male and female groups (Table 1). (14.6%), Wickerhamomyces (4.4%), Cladosporium (4.1%), and Hypho-
pichia (2.2%). At the species level Cryptococcus_longus (26.1%) was the
most abundant, followed by Candida_albicans (9.8%), Wick-
Table 1 erhamomyces_anomalus (4.5%), Hyphopichia_burtonii (2.2%), Candida_
Demographic and clinical characteristics of the study participants. intermedia (1.5%), Candida_sake (1.4%), and Candida_dubliniensis (1.4%)
Variable Male (n ¼ 36) Female (n ¼ 27) p- (Fig. 1A).
value The Sobs, Chao, and ACE indices of the S subgroup were significantly
Age in months (mean  SD) 78.61  21.66 82.48  21.10 0.475a lower (p < 0.05) than those of the P subgroup. In contrast, the coverage
Dentition period 0.131b indices of the S subgroup were significantly higher (p < 0.05) than those
Deciduous dentition 16 7 of the P subgroup (Fig. 1B, Supplementary Table 3). This indicated that
Mixed dentition 20 20
the alpha diversity of the P subgroup was significantly higher than that of
Decayed teeth (mean  SD) 8.05  4.71 6.48  3.42 0.344a
Decayed teeth surfaces (mean  SD) 14.61  11.62 11.92  9.60 0.158a the S subgroup. PCoA plots showed that the samples were clustered by
a
plaque or saliva, indicating variability in fungal composition between
Independent-Sample T Test.
b plaque and saliva samples (p < 0.05) (Fig. 1C).
Chi-Square.

Fig. 1. Composition and diversity of the fungal community. (A) Composition of the fungal community at the phylum, genus, and species level (species abundance
below 0.5% and no annotation in all samples were combined into Others). (B) Alpha diversity analysis of the S and P subgroups. (C) Beta diversity analysis (PCoA
analysis) of the S and P subgroups.

3
M. Yin et al.
3.2.2. Differential species analysis
The top 10 species, in terms of their relative abundances, were
selected at the phylum, genus, and species levels to show the mean
relative abundance and variability of each group (Fig. 2A). The relative
abundance and p-values of the species are presented in Supplementary
Tables 2A and B, and C. At the genus level, the first and second most
abundant taxa in the S subgroup were Cryptococcus (47.9%) and Wick-
erhamomyces (8.5%), respectively, whereas Candida (7.7%) was third,
which was only 1/6th the abundance of Cryptococcus. Meanwhile, the
first most abundant taxa at the genus level in the P subgroup was Cryp-
tococcus (26.1%), which was approximately twice as abundant as Candida
(14.6%).
Species differences analysis suggested that Basidiomycota at the
phylum level; Cryptococcus, Wickerhamomyces, Aspergillus at the genus
level; and Cryptococcus_longus and Wickerhamomyces_anomalus at the
species level were significantly different between the S and P subgroups.
In contrast, no significant differences were observed for Candida
(Fig. 2B).
3.3. Influence of gender and dentition period on S and P subgroups
At the genus level, species differences analysis indicated no signifi-
cant differences between the mycobiome of the SFe and SMa subgroups
or the PFe and PMa subgroups (Fig. 3A, C). Alpha diversity analysis
showed that the Shannon and Simpson indices in the SFe and SMa sub-
groups, as well as in the PFe and PMa subgroups were not significantly
different (p > 0.05) (Fig. 3B, D). Furthermore, Chao1, Ace, Coverage, and
Sobs indices were not significantly different (p > 0.05) (Supplementary
Table 3). Beta diversity unweighted-unifrac index was also not significant
(p > 0.05) (Fig. 3B, D). These results indicate that there was no signifi-
cant difference between alpha and beta diversity. Overall, gender exerted
no effect on the fungal composition of unstimulated saliva and plaque.
Species differences were detected between the SDD and SMD sub-
groups for Ascomycota and Basidiomycota (Fig. 4A), The Shannon and
Simpson indices, as well as the beta diversity unweighted-unifrac index,
in the SDD and SMD subgroups were significantly different (p < 0.05)
(Fig. 4B). In contrast, no significant differences were observed between
the PDD and PMD subgroups (Fig. 4C and D).
Fig. 2. Differential species analysis. (A) Key species differences analysis of the top 10 fungal species at the phylum, genus, and species level in the S and P subgroups.
(B) Species differences analysis at the phylum, genus, and species level in the S and P subgroups.
4
Medicine in Microecology 15 (2023) 100076
3.4. RDA of the relationship between fungal communities and function and
phenotypes
The use of fluoride toothpaste, the frequency of teeth brushing, and
the frequency of sweet consumption may affect the distribution of oral
fungi [27]. An increase in the number of DT and DS may also affect the
composition of the microorganisms. In this study, we collectively cate-
gorized the following as phenotypes: frequency of eating sweets (Eat_-
freq), frequency of brushing teeth (Brush_freq), frequency of fluoride
toothpaste use (Fluoride freq), DT, and DS. We performed RDA to explore
the relationship between these phenotypes and the top 10 oral fungal
species in childhood caries. Among the five factors, Eat_freq had the
relatively lowest effect on the top 10 fungi. Eat_freq, Brush_freq, and the
use of handiwork toothpaste (Fluoride) exhibited the same trend of
positive or negative effects on the flora, and the effects of DT and DS on
fungal flora showed the same trend (Fig. 5A). These results suggest
measures to prevent childhood caries at the microbiological level by
improving dietary and oral hygiene.
Fungal functional annotations were performed according to the
guidelines of the guild prediction system in the FUNGuild software [28]
(Fig. 5B). The annotation function was the sum of the relative abun-
dances of all samples within the subgroup.
4. Discussion
The role of fungi in the microbial composition of dental caries has
recently become a popular topic, and high-throughput sequencing is now
commonly used for fungal detection [24]. This study was conducted to
investigate the differences in fungal communities between unstimulated
saliva and supragingival plaque in childhood caries, and to determine the
effect of gender, dental stage, DT, DS, and oral hygiene behaviors on the
fungal composition of the two groups.
In this study, Cryptococcus was the most abundant genus in both the S
and P subgroups, which differs from previous studies that reported
Candida to be the most dominant fungal genus detected [9,19,29]. In the
S subgroup, the second and third most abundant genera were Wick-
erhamomyces and Candida, respectively, whereas the two microbes were
reversely ranked in the P samples. Cryptococcus and Wickerhamomyces are
less frequently mentioned in caries-related literature. Only a few studies
M. Yin et al.
Fig. 3. Influence of gender on the S and P subgroups. (A) Species differences analyses of the SFe and SMa subgroups at the phylum, genus, and species level. (B) Alpha
and beta diversity of the SFe and SMa subgroups. (C) Species differences analyses of the PFe and PMa subgroups at the phylum, genus, and species level. (D) Alpha and
beta diversity of the PFe and PMa subgroups.
have reported the presence of Cryptococcus in the oral cavity [9,19,22]. A
thick cell wall is a typical feature of the genera Cryptococcus and Wick-
erhamomyces [30]. The bead milling step was performed in this study,
and the lysis time was extended by 15 s to dissociate the thick cell wall of
the fungi and improve the success rate of DNA extraction. This may
explain the high abundance of Cryptococcus and Wickerhamomyces
detected in this study.
Research on Wickerhamomyces is still in its infancy, and it has
currently only been identified as a human opportunistic pathogen [31].
Whether they constitute the core fungal flora in children with dental
caries requires further research.
Combining the results of key species differences analysis and species
differences analysis, the abundance of Cryptococcus and Wick-
erhamomyces were significantly higher in the S subgroup than in the P
subgroup. No relevant studies explain this phenomenon, but we specu-
late that salivary microhabitats may be more suitable for the growth of
these two genera; however, this topic warrants further investigation.
In the alpha diversity analysis, the P subgroup were more abundance
and had higher homogeneity than the S subgroup. This may be due to two
factors. First, unstimulated saliva has a neutral PH environment [32],
whereas supragingival plaque in dental caries is usually mildly acidic
owing to acid production by microorganisms, resulting in an acid-base
5
Medicine in Microecology 15 (2023) 100076
imbalance. A mildly acidic environment is favorable for the growth
and reproduction of microorganisms [16]. Second, thiamine salts, io-
dides, and nitrates in saliva exert antibacterial effects and limit the
growth of microorganisms [32]. This suggests that the plaque is an
optimal environment for supporting the diversity of microorganisms. Our
study suggests different fungal compositions and diversities between the
S and P subgroups, which is consistent with the results of many previous
studies [33–35]. Similarly, the beta diversity indices showed significant
differences in fungal composition between the two subgroups.
Gender, age, and other factors may affect the oral fungal population
of childhood caries. Jesus et al. [36] suggested that gender might be a
critical determinant of severe early childhood caries (S-ECC). In contrast,
our study did not observe any effect of gender on the fungal microbiome
in the S and P subgroups. These inconsistent findings may be due to the
following variables: (1) sample geographical location, wherein the
former findings were collected in Canada, whereas the present study was
conducted in China; (2) the age range, wherein the former study popu-
lation was younger than 72 months of age, while that in the present study
was between 2 and 9 years of age; (3) the former study investigated the
influence of gender on the supragingival plaque fungal community in
caries-free and S-ECC children, and the present study focused on whether
gender had any effect on the respective microbial composition of saliva
M. Yin et al. Medicine in Microecology 15 (2023) 100076

Fig. 4. Influence of dentition period on the S and P subgroups. (A) Species differences analyses of the SDD and SMD subgroups at the phylum, genus, and species level.
(B) Alpha and beta diversity of the SDD and SMD subgroups. (C) Species differences analyses of the PDD and PMD subgroups at the phylum, genus, and species level.
(D) Alpha and beta diversity of the PDD and PMD subgroups.

Fig. 5. Redundancy analysis (RDA) of the relationship between fungal and phenotypes and fungal functional annotation. (A) Redundancy analysis (RDA) of the top 10
fungal flora and phenotypes. (B) Functional annotation of the S and P subgroups. (Function on the annotation is the sum of the relative abundance of all samples in
the group.)

6
M. Yin et al.
and plaque in children with caries. These findings suggest that we are far
from fully understanding the influence of gender on dental fungal com-
munities and further research is needed.
Mycobiome differences were detected between the SDD and SMD
subgroups but not between the PDD and PPD groups, suggesting denti-
tion is not an influential factor of supragingival plaque fungi. However, it
does affect the fungal distribution in saliva.
This may be attributed to the low biomass of fungi in the oral cavity
and the fact that the fungal content in supragingival plaque may be lower
than that in saliva; thus, no differences were detected in the plaque [19,
37]. The composition of supragingival plaque may be relatively stable
once it is formed; therefore, it is not significantly affected by different
dental periods.
The FUNGuild software currently lacks data on the fungal genome, so
we can only perform functional annotations but not pathway prediction.
This suggests that most of the current studies have been based on
investigating ITS genes and have not investigated pathway analyses.
Despite the progress in understanding the microbial composition of
oral fungi and their biological functions, several challenges remain and
require urgent resolution. In our study, a significant proportion of fungi
were classified as Others in the S and P subgroups as their taxonomic
levels were not annotated or their abundance was <0.5% in the samples.
We noted at least three problems. First, experimental biases introduced in
key procedures may affect the experimental results [38], which may
suggest that appropriate experimental protocols should be developed for
different samples. Second, the NGS platform used in this experiment,
which only sequences ITS regions, has proven to be technically limited
[24]. The current third-generation sequencing technologies, SGS PacBio
RSII and Sequel, can sequence the complete ITS region and identify mi-
crobial communities more accurately than NGS [39,40]. Finally,
although UNITE is currently the most commonly used taxonomic data-
base for studying fungal biomes in different environments, there is a
growing concern about the inadequate taxonomic coverage of existing
databases, with many species being detected and unable to be classified
to a meaningful taxonomic level outside the kingdom level. This limits
studies attempting to describe the human fungal biome [24].
Our findings revealed significant differences in the fungal composi-
tion between unstimulated saliva and supragingival plaque. Dentition
period and oral hygiene behaviors may have affected these differences.
However, as this was a cross-sectional study, to ensure reliability, the
results need to be supported by longitudinal studies, and the bacterial
and viral communities should also be studied to obtain a more compre-
hensive understanding of the microbial composition of the S and P
samples of childhood caries.
To the best of our knowledge, this is the first study to comprehen-
sively describe the differences and influencing factors in the fungal
composition in unstimulated saliva and supragingival plaque of child-
hood caries. It provides a reference for further studies of fungal micro-
biota of childhood caries to aid the selection of an appropriate sample
source. In addition, it also serves as a foundation for future research of
the correlation between oral fungi and childhood caries.
Author contribution
Mx.Y: Conceptualization, Methodology, Formal analysis, Investiga-
tion, Project administ ration, Writing – original draft; Y.Y: Methodology,
Software, Investigation. X.Z: Fo rmal analysis, Investigation, Review and
Editing. Qy.L: Investigation, Software, Review and Editing. Wa.X:
Conceptualization, Resources, Review and editing, Funding acquisit ion.
Bl.W: Review and Editing, Visualization, Funding acquisition.
Conflict of Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
7
Medicine in Microecology 15 (2023) 100076
Acknowledgments
This work was supported by grants from the President Foundation of
Shenzhen Stomatology Hospital (Pingshan) of Southern Medical Uni-
versity (grant no. 2021A001) and Oral Infectious Disease Mechanism
Research and Clinical Translation Application Innovation team of
Guangdong Province of China (grant no. 2021KCXTD033). We would
like to thank Editage (www.editage.cn) for the English language editing.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https
://doi.org/10.1016/j.medmic.2023.100076.
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