Evaluation of the Effectiveness of Crotoxin as an Antiseptic against Candida spp. Biofilms.

The growing number of oral infections caused by the Candida species are becoming harder to treat as the commonly used antibiotics become less effective. This drawback has led to the search for alternative strategies of treatment, which include the use of antifungal molecules derived from natural products. Herein, crotoxin (CTX), the main toxin of Crotalus durissus terrificus venom, was challenged against Candida tropicalis (CBS94) and Candida dubliniensis (CBS7987) strains by in vitro antimicrobial susceptibility tests. Minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC), and inhibition of biofilm formation were evaluated after CTX treatment. In addition, CTX-induced cytotoxicity in HaCaT cells was assessed by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) colorimetric assay. Native CTX showed a higher antimicrobial activity (MIC = 47 µg/mL) when compared to CTX-containing mouthwash (MIC = 750 µg/mL) and nystatin (MIC = 375 µg/mL). Candida spp biofilm formation was more sensitive to both CTX and CTX-containing mouthwash (IC100 = 12 µg/mL) when compared to nystatin (IC100 > 47 µg/mL). Moreover, significant membrane permeabilization at concentrations of 1.5 and 47 µg/mL was observed. Native CTX was less cytotoxic to HaCaT cells than CTX-containing mouthwash or nystatin between 24 and 48 h. These preliminary findings highlight the potential use of CTX in the treatment of oral candidiasis caused by resistant strains.

Candidemia Candida albicans clusters have higher tendency to form biofilms than singleton genotypes†.

The capacity of Candida spp. to form biofilms allows them to attach either to living or inert surfaces, promoting their persistence in hospital environments. In a previous study, we reported strain-to-strain variations in Candida spp. biofilm development, suggesting that some genotypes may be greater biofilm formers than others. In this study, we hypothesize that isolates pertaining to clusters may be found more frequently in the environment due to their ability to form biofilms compared to singleton genotypes. Two hundred and thirty-nine Candida spp. isolates (78 clusters) from candidemia patients admitted to 16 hospitals located in different cities and countries-and the same number of singleton genotypes used as controls-were tested in terms of biofilm formation using the crystal violet and the XTT reduction assays. Candida albicans clusters showed higher biofilm formation in comparison to singleton genotypes (P < .01). The biofilms formed by intra-hospital C. albicans clusters showed higher metabolic activity (P < .05). Furthermore, marked variability was found among species and type of cluster. We observed that the higher the number of isolates, the higher the variability of biofilm production by isolates within the cluster, suggesting that the production of biofilm by isolates of the same genotype is quite diverse and does not depend on the type of cluster studied. In conclusion, candidemia Candida spp. clusters-particularly in the case of C. albicans-show significantly more biomass production and metabolic activity than singleton genotypes.

Epidemiology and prognostic factors of nosocomial candidemia in Northeast Brazil: A six-year retrospective study.

Candidemia has been considered a persistent public health problem with great impact on hospital costs and high mortality. We aimed to evaluate the epidemiology and prognostic factors of candidemia in a tertiary hospital in Northeast Brazil from January 2011 to December 2016. Demographic and clinical data of patients were retrospectively obtained from medical records and antifungal susceptibility profiling was performed using the broth microdilution method. A total of 68 episodes of candidemia were evaluated. We found an average incidence of 2.23 episodes /1000 admissions and a 30-day mortality rate of 55.9%. The most prevalent species were Candida albicans (35.3%), Candida tropicalis (27.4%), Candida parapsilosis (21.6%) and Candida glabrata (11.8%). Higher mortality rates were observed in cases of candidemia due to C. albicans (61.1%) and C. glabrata (100%), especially when compared to C. parapsilosis (27.3%). Univariate analysis revealed some variables which significantly increased the probability of death: older age (P = 0.022; odds ratio [OR] = 1.041), severe sepsis (P < 0.001; OR = 8.571), septic shock (P = 0.035; OR = 3.792), hypotension (P = 0.003; OR = 9.120), neutrophilia (P = 0.046; OR = 3.080), thrombocytopenia (P = 0.002; OR = 6.800), mechanical ventilation (P = 0.009; OR = 8.167) and greater number of surgeries (P = 0.037; OR = 1.920). Multivariate analysis showed that older age (P = 0.040; OR = 1.055), severe sepsis (P = 0.009; OR = 9.872) and hypotension (P = 0.031; OR = 21.042) were independently associated with worse prognosis. There was no resistance to amphotericin B, micafungin or itraconazole and a low rate of resistance to fluconazole (5.1%). However, 20.5% of the Candida isolates were susceptible dose-dependent (SDD) to fluconazole and 7.7% to itraconazole. In conclusion, our results could assist in the adoption of strategies to stratify patients at higher risk for developing candidemia and worse prognosis, in addition to improve antifungal management.

Effect of quorum sensing molecules and natamycin on biofilms of Candida tropicalis and other yeasts isolated from industrial juice filtration membranes.

AIMS: Cells limit the cell number of dense biofilms by releasing self-inhibitory molecules. Here, we aim to assess the effectiveness of yeast quorum sensing (QS) molecules and the antifungal agent natamycin against yeast biofilms of strains commonly isolated from fruit juice ultrafiltration membranes. METHODS AND RESULTS: Yeast QS molecules, such as tyrosol, 2-phenylethanol and farnesol, were detected by solvent extraction and HS-SPME GC-MS in Candida tropicalis cultures. The effect of QS molecules on mono- and multispecies biofilms formed by Rhodotorula mucilaginosa, C. tropicalis, Candida krusei and Candida kefyr was evaluated by plate count and epifluorescence microscopy. Farnesol caused a decrease in cell number and disrupted mono- and multispecies yeast biofilms during adhesion (0·6 mmol l-1 ). 2-phenyl ethanol 1·2 mmol l-1 stimulated biofilm density and increased cell number in both mono- and multispecies biofilms, while tyrosol did not show effects when tested against C. tropicalis biofilms (0·05-1·2 mmol l-1 ). Natamycin caused a strong decrease in cell number and disruption of biofilm structure in C. tropicalis biofilms at high concentrations (0·3-1·2 mmol l-1 ). The combination of farnesol 0·6 mmol l-1 and natamycin at 0·01 mmol l-1 , the maximum concentration of natamycin accepted for direct addition into fruit juices, effectively reduced cell counts and disrupted the structure of C. tropicalis biofilms. CONCLUSION: Farnesol 0·6 mmol l-1 significantly increased the inhibition exerted by natamycin 0·01 mmol l-1 (~5 ppm) reducing biofilm development from juice on stainless steel surfaces. SIGNIFICANCE AND IMPACT OF THE STUDY: These results support the use of QS molecules as biofilm inhibitors in beverages and would certainly inspire the design of novel preservative and cleaning products for the food industry based on combinatory approaches.

Lipid core nanoparticles as a broad strategy to reverse fluconazole resistance in multiple Candida species.

Fungal resistance is the major problem related to fluconazole treatments. This study aims to develop innovative lipid core nanocapsules and nanostructured lipid carriers containing fluconazole, to study in vitro antifungal activity and to assess the possibility of resistance reversion in Candida albicans, C. glabrata, C. krusei, and C. tropicalis isolates. The action mechanism of nanoparticles was investigated through efflux pumps and scanning electron microscopy studies. The lipid core nanocapsules and nanostructured lipid carriers were prepared by interfacial deposition of preformed polymer and high-pressure homogenization methods, respectively. Both nanostructures presented sizes below 250 nm, SPAN < 1.6, negative zeta potential, pH slightly acid, high drug content and controlled drug release. The nanostructured lipid carriers were unable to reverse the fungal resistance. Lipid core nanoparticles displayed advantages such as a reduction in the effective dose of fluconazole and resistance reversion in all isolates tested - with multiple mechanisms of resistance. The main role of the supramolecular structure and the composition of the nanoparticles on antifungal mechanisms of action were discussed. The results achieved through this study have an impact on clinical therapy, with a potential application in the treatment of fungal infections caused by resistant isolates of Candida spp.

In vitro interaction of Candida tropicalis biofilm formed on catheter with human cells.

Candida tropicalis has emerged as one of the major Candida non-C. albicans species, in terms of epidemiology and virulence. Despite its virulence, C. tropicalis pathogenic mechanism has yet not been fully defined. The current study aimed to demonstrate the interaction of mature C. tropicalis ATCC 750 biofilm formed on catheter with different human cell lines. In vitro mature (72 h) C. tropicalis biofilms were produced on small catheter fragments (SCF) and were mainly composed by blastoconidia. Then, migration of yeast cells from mature biofilm to human cell surfaces (HeLa and HUVEC) was investigated. After contact with both cell lines, the surface of SCF, containing mature C. tropicalis biofilm, exhibited predominantly the filamentous form. Meanwhile, fresh biofilm formed on human cell surfaces also revealed mainly of blastoconidia involved by extracellular matrix. Total biomass and metabolic activity from the remaining biofilm on SCF surface, after direct contact with human cells, exhibited a significant reduction. Mature C. tropicalis biofilm modified its extracellular matrix components, after contact with human cells. Thus, we described for the first time an easy and simple in vitro model with catheter, which could be a powerful tool for future studies that desires to elucidate the mechanisms involved in C. tropicalis biofilm.

γ-irradiation from radiotherapy improves the virulence potential of Candida tropicalis.

AIM: To evaluate if radiation used in radiotherapy can cause changes in the virulence potential of Candida tropicalis ATCC 750. MATERIALS & METHODS: C. tropicalis was exposed in vitro to identical dose and scheme of irradiation would be used in patients with head and neck cancer. Some virulence parameters were analyzed before and after irradiation. RESULTS: Colony morphologies were irreversibly affected by irradiation. Increase in growth rate, filamentation, adhesion on cell lines and phagocytosis process were also observed. Overall the irradiated C. tropicalis cells became more efficient at causing systemic infection in mice. CONCLUSION: γ-radiation induced important changes in C. tropicalis increasing its virulence profile, which could directly affect the relationship between yeasts and hosts.

Antifungal mechanism of [RuIII(NH3)4catechol]+ complex on fluconazole-resistant Candida tropicalis.

Candidiasis, a major opportunistic mycosis caused by Candida sp., may comprise life-threatening systemic infections. The incidence of non-albicans species is rising, particularly in South America and they are frequently drug resistant, causing unresponsive cases. Thus, novel antimycotic agents are required. Here we tested the antifungal activity of [RuIII(NH3)4catechol]+ complex (RuCat), approaching possible action mechanisms on fluconazole-resistant Candida tropicalis. RuCat significantly (P < 0.05) inhibited the growth and viability of C. tropicalis dose-dependently (IC50 20.3 µM). Cytotoxicity of RuCat upon murine splenocytes was lower (Selectivity Index = 16). Scanning electron microscopy analysis showed pseudohyphae formation, yeast aggregation and surface damage. RuCat-treated samples investigated by transmission electron microscopy showed melanin granule trafficking to cell surfaces and extracellular milieu. Surface-adherent membrane fragments and extracellular debris were also observed. RuCat treatment produced intense H2DCFDA labeling, indicating reactive oxygen species (ROS) production which caused increased lipoperoxidation. ROS are involved in the fungicidal effect as N-acetyl-L-cysteine completely restored cell viability. Calcofluor White chitin staining suggests that 70 or 140 µM RuCat treatment for 2 h affected cell-wall structure. PI labeling indicated necrotic cell death. The present data indicate that RuCat triggers ROS production, lipoperoxidation and cell surface damage, culminating in selective necrotic death of drug-resistant C. tropicalis.

Hemolytic capability and expression of a putative haem oxygenase-encoding gene by blood isolates of Candida tropicalis are influenced by iron deprivation and the presence of hemoglobin and erythrocytes.

Although hemolytic activity is known to be a putative virulence factor contributing to candidal pathogenesis, its production by Candida tropicalis, a species closely related to Candida albicans, is poor understood. The present study was undertaken to evaluate the hemolytic activity and the expression level of a putative haem oxygenase encoding gene by blood isolates of C. tropicalis following growth in iron deprivation, and in the presence of hemoglobin and erythrocytes. The lowest values of hemolytic activity were observed in cell-free culture supernatants of isolates growing in iron-restricted medium (RPMI medium and RPMI medium supplemented with iron chelator bathophenanthrolindisulphonic acid). Hemolysis was increased in the presence of either hemoglobin or erythrocytes. Reverse transcriptase PCR analysis showed that the putative haem oxygenase encoding gene (CtHMX1), potentially related with iron uptake, was up-regulated (p < 0.001) following growth in iron deprivation and in the presence of hemoglobin; CtHMX1 was repressed in the presence of human erythrocytes (p < 0.001). Our data suggest that hemoglobin had positive effect in the production of hemolytic factor and gene expression related to iron uptake in C. tropicalis.

Reduction of Candida tropicalis biofilm by photoactivation of a Heterophyllaea pustulata extract.

CONTEXT: Biofilm formation is an important problem, since this growth mode confers resistance to drugs usually used in therapeutics. OBJECTIVE: In vitro antifungal activity of extracts obtained from Heterophyllaea pustulata Hook f. (Rubiaceae) were studied against Candida tropicalis biofilms, evaluating the effect of irradiation and the oxidative and nitrosative stresses as possible mechanisms of action. MATERIALS AND METHODS: Hexane, benzene, ethyl acetate and ethanol extracts were evaluated at three concentrations (0.2, 0.1 and 0.05 mg/mL) over mature biofilm, under darkness and irradiation. After 48 h of incubation, biofilm quantitation was performed by the O'Toole and Kolter method. Reactive oxygen species (ROS) was measured by nitro-blue tetrazolium (NBT) reaction and reactive nitrogen intermediates (RNI) by the Griess reagent. Superoxide dismutase activation (SOD, NBT assay) and total antioxidant system (FRAP test) were studied. RESULTS: Only the benzene extract at 0.2 mg/mL reduced the biofilms formation. The slight decrease achieved in darkness (17.06 ± 2.80% reduction) was increased by light action (39.31 ± 3.50% reduction), clearly observing a photostimulation. This great reduction was confirmed by confocal microscopy. In darkness, biofilm reduction was mediated by an increase in RNI, whereas under irradiation, the ROS action was most important. Although no SOD activation was observed, a strong stimulation of the total antioxidant system was detected. HPLC analysis established a high content of several anthraquinones in this extract. DISCUSSION AND CONCLUSION: Biofilm reduction by benzene extract was mainly mediated by oxidative stress triggered under light action, confirming a photodynamic sensitization, which could be attributed to its high content of photosensitizing anthraquinones.

In Vitro Studies of Chromone-Tetrazoles against Pathogenic Protozoa, Bacteria, and Fungi.

In vitro studies to fourteen previously synthesized chromone-tetrazoles and four novel fluorine-containing analogs were conducted against pathogenic protozoan (Entamoeba histolytica), pathogenic bacteria (Pseudomonas aeruginosa, and Staphylococcus aureus), and human fungal pathogens (Sporothrix schenckii, Candida albicans, and Candida tropicalis), which have become in a serious health problem, mainly in tropical countries.

Microwave irradiation as an alternative method for disinfection of denture base acrylic resins.

AIM: This study evaluated the effect of microwave irradiation as an alternative method for disinfection of different types of denture base acrylic resins. METHODS: Twenty-four samples for each conventional, microwaved and characterized heat-cured acrylic resin were made and subjected to sterilization with ethylene oxide for the groups: 1) irradiated samples; 2) non-irradiated samples; and 3) samples without yeast. Each group was subdivided according to inoculation with C. albicans, C. dubliniensis and C. tropicalis. The samples were inoculated with 100 µL of inoculum of each species of Candida and later placed in an incubator at 37 °C for 1 hr to perform the first adhesion. After this time, each well was supplemented with sterile media and the plate was once again taken to a stove for incubation at 37 °C for 6 hr. The samples were immersed in 100 mL of sterile water and irradiated with microwave at 650 W for 3 min. Control samples were considered as the non-irradiated group. After incubation for 48 hr, irradiated and non-irradiated samples were subjected to a digital colony counter. RESULTS: Control group (non-irradiated) showed microbial growth for resins and the means of ufc/mL were without statistically significant differences. Microwave irradiated samples (experimental group) promoted no viable colonies for all Candida species and types of acrylic resins. The means of ufc/mL were without statistically significant differences. CONCLUSION: Microwave irradiation was an effective method for disinfection of the acrylic resins inoculated with C. albicans, C. dubliniensis and C. tropicalis.

Cellular characterisation of Candida tropicalis presenting fluconazole-related trailing growth.

We assessed fluconazole susceptibility in 52 Candida tropicalis clinical strains using seven antifungal susceptibility methods, including broth microdilution (BMD) [standard M27 A3 (with neutral and acid pH), ATB Fungus 3, Vitek 2 system and flow cytometric analysis] and agar-based methods (disk diffusion and E-test). Trailing growth, detection of cell-associated secreted aspartic proteases (Saps) and morphological and ultrastructural traits of these clinical strains were also examined. The ranges of fluconazole 24 h-minimum inhibitory concentration (MIC) values were similar among all methods. The essential agreement among the methods used for MIC determinations was excellent and all methods categorised all strains as susceptible, except for one strain that showed a minor error. The presence of the trailing effect was assessed by six methods. Trailing positivity was observed for 86.5-100% of the strains. The exception was the BMD-Ac method where trailing growth was not observed. Morphological and ultrastructural alterations were detected in C. tropicalis trailing cells, including mitochondrial swelling and cell walls with irregular shapes. We tested the production of Saps in 13 C. tropicalis strains expressing trailing growth through flow cytometry. Our results showed that all of the C. tropicalis strains up-regulated surface Sap expression after 24 h or 48 h of exposure to fluconazole, which was not observed in untreated yeast strains. We concluded that C. tropicalis strains expressing trailing growth presented some particular features on both biological and ultrastructural levels.

Cellular characterisation of Candida tropicalis presenting fluconazole-related trailing growth

We assessed fluconazole susceptibility in 52 Candida tropicalis clinical strains using seven antifungal susceptibility methods, including broth microdilution (BMD) [standard M27 A3 (with neutral and acid pH), ATB Fungus 3, Vitek 2 system and flow cytometric analysis] and agar-based methods (disk diffusion and E-test). Trailing growth, detection of cell-associated secreted aspartic proteases (Saps) and morphological and ultrastructural traits of these clinical strains were also examined. The ranges of fluconazole 24 h-minimum inhibitory concentration (MIC) values were similar among all methods. The essential agreement among the methods used for MIC determinations was excellent and all methods categorised all strains as susceptible, except for one strain that showed a minor error. The presence of the trailing effect was assessed by six methods. Trailing positivity was observed for 86.5-100 percent of the strains. The exception was the BMD-Ac method where trailing growth was not observed. Morphological and ultrastructural alterations were detected in C. tropicalis trailing cells, including mitochondrial swelling and cell walls with irregular shapes. We tested the production of Saps in 13 C. tropicalis strains expressing trailing growth through flow cytometry. Our results showed that all of the C. tropicalis strains up-regulated surface Sap expression after 24 h or 48 h of exposure to fluconazole, which was not observed in untreated yeast strains. We concluded that C. tropicalis strains expressing trailing growth presented some particular features on both biological and ultrastructural levels.

In vitro activity of Schinus terebinthifolius (Brazilian pepper tree) on Candida tropicalis growth and cell wall formation.

The aim of this study was to evaluate the in vitro antifungal activity of Schinus terebinthifolius (Brazilian pepper tree) tincture on planktonic Candida tropicalis (ATCC 40042), which is a microorganism associated to oral cavity infections. Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) were determined through the microdilution technique. Possible action of the tincture on fungal cell wall formation was also studied by adding an osmotic protector (0.8M sorbitol) to the microplates. Nystatin was used as standard control and tests were performed in triplicate. S. terebinthifolius was found to have MIC and MFC values of 625 microg/mL on the strain assayed, whereas nystatin showed MIC and MFC of 6.25 microg/mL. Results suggest that S. terebinthifolius tincture acts on fungal cell walls, since the sorbitol test indicated a MIC of 1.250 microg/mL. It may be concluded that S. terebinthifolius has potential in vitro antifungal activity against C. tropicalis strains, and probably acts by inhibiting fungal cell wall formation.

Capsicum annuum L. trypsin inhibitor as a template scaffold for new drug development against pathogenic yeast.

A 6,000 Da peptide, named CaTI, was isolated from Capsicum annuum L. seeds and showed potent inhibitory activity against trypsin and chymotrypsin. The aim of this study was to determine the effect of CaTI on Saccharomyces cerevisiae, Candida albicans, Candida tropicalis and Kluyveromyces marxiannus cells. We observed that CaTI inhibited the growth of S. cerevisiae, K. marxiannus as well as C. albicans and induced cellular agglomeration and the release of cytoplasmic content. No effect on growth was observed in C. tropicalis but morphological changes were noted. In the spot assay, different degrees of sensitivity were shown among the strains and concentrations tested. Scanning electron microscopy showed that S. cerevisiae, K. marxiannus and C. albicans, in the presence of CaTI, exhibited morphological alterations, such as the formation of pseudohyphae, cellular aggregates and elongated forms. We also show that CaTI induces the generation of nitric oxide and interferes in a dose-dependent manner with glucose-stimulated acidification of the medium mediated by H(+)-ATPase of S. cerevisiae cells.

Anti-Candida activity of Mentha arvensis and Turnera ulmifolia.

Candidiasis is the most frequent infection by opportunistic fungi, frequently caused by Candida albicans, Candida tropicalis, Candida parapsilosis, Candida glabrata, and Candida krusei. Mentha arvensis L. is a herbaceous plant that occurs throughout South America and is used as a tea and in the folk medicine. Turnera ulmifolia L. is already known to be of medicinal value. Ethanol extracts from M. arvensis and T. ulmifolia were assayed for antifungal activity against strains of C. albicans, C. tropicalis, and C. krusei. No clinically relevant antifungal activity was demonstrated by the extracts; however, a potentiation effect was observed when the extracts were applied with metronidazole against C. tropicalis. M. arvensis and T. ulmifolia could represent a source of natural products with modifying antifungal activity.

Ultrastructural architecture of colonies of different morphologies produced by phenotypic switching of a clinical strain of Candida tropicalis and biofilm formation by variant phenotypes.

Candida tropicalis has been identified as one of the most prevalent pathogenic yeast species of the Candida-non-albicans (CNA) group. Study of switching in C. tropicalis has not been the subject of extensive research. Therefore, we investigated switching event and characterized the ultrastructural architecture of different phenotypes and biofilm produced in a C. tropicalis clinical strain. Cells switched heritably, reversibly, and at a high frequency between four phenotypes readily distinguishable by the shape of colonies formed on agar at 25°C. SEM analysis was used to verify the architecture of whole Candida colonies at ultrastructural level. The smooth phenotype (parental phenotype) colony showed a hemispherical shape character, while the semi-smooth was characterized by the presence of shallow marginal depressions. The ring and rough phenotypes exhibited more complex architecture and were characterized by the presence of deep central and peripheral depressions areas. The biofilm-forming ability varied among the switch phenotypes. After 12h incubation, the smooth phenotype formed less biofilm compared to the other phenotypes (P<0.05). The electron microscopy analysis revealed that filamentation (pseudohyphae) was associated with ring and rough colonies. The ultrastructural analysis allowed the observation of the arrangement of individual cells within the colonies. At the deep central and peripheral depressions areas of the ring and rough colonies extracellular material was seen in different arrangements. The data presented here open new avenues to study a possible role for extracellular material in the formation and maintenance of the architecture of switch phenotypes in C. tropicalis. It is therefore essential that more strains be investigated to determine the biological significance of extracellular material in C. tropicalis phenotypic switching phenomenon.

Reduction of Candida species colonization in the oral cavity of children infected with human immunodeficiency virus after dental treatment.

OBJECTIVES: The objective of this study was to evaluate Candida species in the oral mucosa before and after dental treatment in children infected with human immunodeficiency virus (HIV) and investigate the relation of these yeasts with cavitated carious lesions, immune suppression, and use of antiretroviral drugs. METHODS: Specimens of oral mucosa (M1) and dentin were collected, using a swab and dental curette respectively, from 30 HIV-infected children. Dental treatment (tooth-brushing, fluoride application, restoration of all carious lesions and extractions, if needed) was performed, and at posttreatment (M2, mean 30 days after baseline) another sample of oral mucosa was collected. RESULTS: Candida colony-forming unit (CFU) values were greater in dentin when compared with oral mucosa before dental treatment (M1), and a reduction of these fungi was observed in oral mucosa after dental treatment (M2) (P < .05). Cavitated carious lesions and caries in dentin were positively correlated with CFU in M1 (P < .05). C. albicans was the most prevalent of the Candida species. A relationship between the level of immune suppression and use of antiretroviral drugs with the prevalence of yeasts was not observed (P > .05). CONCLUSION: The decrease in CFU at M2 and the high prevalence of these yeasts in dentin indicates that cavitated lesions in this population can serve as a niche for colonization and proliferation of Candida species in HIV-infected children.