Influence of oral biofilm index, caries experience, and laboratory markers of disease progression on the oral carriage of Candida in HIV-infected and non-infected children: a cross-sectional study.

The present study aimed to compare the oral Candida rate between infected and uninfected children with the human immunodeficiency virus (HIV), as well as analyze the association between Candida spp. and predisposing factors of colonization, like oral biofilm index, caries experience, and laboratory markers of AIDS progression. A cross-sectional study was employed. Candida species were identified and quantified from saliva samples of 50 HIV-infected and 50 uninfected children. Biofilm index and decayed, missing, and filled teeth (dmft/DMFT) indices were assessed by oral clinical examinations. Additionally, CD4+ T lymphocyte count and viral load were obtained from medical records of the HIV-infected children. Candida species were cultured from 74% of the HIV-infected children and 46% of uninfected ones (p = 0.0076). Candida albicans and Candida parapsilosis were the most frequently isolated species in both studied groups. The isolation of Candida species was significantly higher in HIV-infected children with CD4 ≤ 15% (p = 0.0146); it had influence of mature oral biofilm and the caries index (dmft + DMFT ≥ 8) (p < 0.05) and was associated with the plasma viral load. The present data show that the HIV infection, oral biofilm index, caries experience, and laboratory markers of AIDS progression exert an influence on the prevalence of oral Candida in children.

Anti-Leishmania amazonensis activity of the marine sponge Dercitus (Stoeba) latex (Porifera) from São Pedro and São Paulo Archipelago, Pernambuco, Brazil.

The search for new therapeutic strategies for leishmaniasis treatment is essential due to the side effects of available drugs and the increasing incidence of resistance to them. Marine sponges use chemical compounds as a defense mechanism, and several of them present interesting pharmacological properties. The aim of this study was to evaluate the in vitro activity of the aqueous extract of the marine sponge Dercitus (Stoeba) latex against Leishmania amazonensis. MIC and toxicity against mammal cells were evaluated through broth microdilution assays. Transmission electron microscopy analysis was performed to assess possible effects on L. amazonensis ultrastructure. Arginase and proteolytic activities were measured by spectrometric methodologies. The extract of Dercitus (Stoeba) latex displayed antileishmanial activity and moderate toxicity against peritonial macrophages. Ultrastructural changes were observed after the growth of L. amazonensis promastigotes in the presence of the extract at 150 µg.ml-1 (IC50), mainly on acidocalcysomes. The extract was able to inhibit the activity of arginase and serine proteases. This study shows that Dercitus (Stoeba) latex aqueous extract may be a novel potential source of protozoa protease inhibitors and drugs that are less toxic to be used in the treatment of L. amazonensis infections.

Synergistic interactions between ß-lapachone and fluconazole in the inhibition of CaCdr2p and CaMdr1p in Candida albicans.

BACKGROUND: Mortality rate of invasive Candida infections is raising mainly amongst immunocompromised patients. These infections are hard-to-treat mainly due to the increasing incidence of resistance. The overexpression of ATP-binding cassette and major facilitator superfamily transporters is the main responsible for the failure of antifungal therapies. In a Saccharomyces cerevisiae model, ß-lapachone inhibited Pdr5p, a transporter homologous to those found in Candida albicans. AIMS: To determine whether ß-lapachone reverses the resistance phenotype mediated by efflux transporters in C. albicans clinical isolates. METHODS: The antifungal activity of ß-lapachone combined with fluconazole was measured by agarose chemosensitization and microdilution assays. CaCdr2p and CaMdr1p activities were evaluated through fluorescent dyes accumulation. ATPase activity was assessed using transporter-enriched plasma membranes. RESULTS: ß-lapachone reverted antifungal resistance of S. cerevisiae and C. albicans strains overexpressing CaCdr2p and CaMdr1p transporters by inhibiting these proteins activities. CaCdr2p ATPase activity was not impaired by the compound. CONCLUSIONS: ß-lapachone is a promising drug candidate to be used as an adjuvant in the treatment of candidiasis caused by fluconazole-resistant C. albicans strains.

Synergistic interactions between Beta-lapachone and fluconazole in the inhibition of CaCdr2p and CaMdr1p in Candida albicans / Las interacciones sinérgicas entre la beta-lapachona y el fluconazol en la inhibición de CaCdr2p y CaMdr1p en Candida albicans

BACKGROUND: Mortality rate of invasive Candida infections is raising mainly amongst immunocompromised patients. These infections are hard-to-treat mainly due to the increasing incidence of resistance. The overexpression of ATP-binding cassette and major facilitator superfamily transporters is the main responsible for the failure of antifungal therapies. In a Saccharomyces cerevisiae model, Beta-lapachone inhibited Pdr5p, a transporter homologous to those found in Candida albicans. AIMS: To determine whether Beta-lapachone reverses the resistance phenotype mediated by efflux transporters in C. albicans clinical isolates. METHODS: The antifungal activity of Beta-lapachone combined with fluconazole was measured by agarose chemosensitization and microdilution assays. CaCdr2p and CaMdr1p activities were evaluated through fluorescent dyes accumulation. ATPase activity was assessed using transporter-enriched plasma membranes. RESULTS: Beta-lapachone reverted antifungal resistance of S. cerevisiae and C. albicans strains overexpressing CaCdr2p and CaMdr1p transporters by inhibiting these proteins activities. CaCdr2p ATPase activity was not impaired by the compound. CONCLUSIONS: Beta-lapachone is a promising drug candidate to be used as an adjuvant in the treatment of candidiasis caused by fluconazole-resistant C. albicans strains

ANTECEDENTES: Las tasas de mortalidad de infecciones invasivas causadas por Candida están en aumento, principalmente entre los pacientes inmunocomprometidos. Estas infecciones son difíciles de tratar debido a la creciente incidencia de resistencia a los antifúngicos. La sobreexpresión de los transportadores dependientes de ATP y los de la superfamilia de facilitadores principales es el mayor responsable del fracaso de las terapias antimicóticas. En un modelo de Saccharomyces cerevisiae, la beta-lapachona inhibió Pdr5p, un transportador homólogo a los encontrados en Candida albicans. OBJETIVOS: Determinar si la beta-lapachona revierte el fenotipo de resistencia mediado por transportadores de eflujo en aislamientos clínicos de C. albicans. MÉTODOS: Se midió la actividad antifúngica de la beta-lapachona combinada con fluconazol mediante ensayos de quimiosensibilización con agarosa y microdilución. Las actividades CaCdr2p y CaMdr1p se evaluaron mediante la acumulación de colorantes fluorescentes, y la actividad de ATPasa se evaluó usando membranas plasmáticas enriquecidas con transportador. RESULTADOS: La beta-lapachona revirtió la resistencia antifúngica de las cepas de S. cerevisiae y C. albicans que sobreexpresaban los transportadores CaCdr2p y CaMdr1p al inhibir sus actividades. El compuesto no afectó la actividad ATPasa de CaCdr2p. CONCLUSIONES: La beta-lapachona es una candidata prometedora para ser utilizada como adyuvante en el tratamiento de la candidiasis causada por cepas de C. albicans resistentes al fluconazol

Histatin 5 and human lactoferrin inhibit biofilm formation of a fluconazole resistant Candida albicans clinical isolate.

Candida albicans is the most important fungal pathogen that causes infections in humans. Biofilms are hard-to-treat structures due to their high antifungal resistance. Saliva is a fluid that contains antimicrobial substances acting as the first-line of defense against pathogens, and its immune components may be potential tools for the discovery of new treatments against candidiasis. To evaluate the activity of histatin 5 and human lactoferrin against biofilm formation. A fluconazole-resistant Candida albicans clinical isolate was used as the model microorganism. Morphogenesis was evaluated by differential counting. Biofilm quantification was performed by XTT reduction assay. Thickness and topography of biofilms were assessed through confocal laser scanning microscopy (CLSM). Histatin 5 inhibited yeast-to-hyphae transition in a dose-dependent manner, while the effect of human lactoferrin on this process was inversely proportional to its concentration. Both compounds were able to significantly inhibit biofilm metabolic activity. Histatin 5 reduced biofilm thickness. Histatin 5 and human lactoferrin exhibited in vitro cytotoxicity against a fluconazole-resistant Candida albicans biofilm, which points to the potential application of these compounds in the treatment of biofilms formed by this fungus, especially in resistant infections.

Optimization of von Willebrand factor multimer analysis in vertical mini-gel electrophoresis systems: A rapid procedure.

Von Willebrand disease (VWD) is a common cause of bleeding worldwide. Analysis of von Willebrand factor (VWF) multimer distribution (VWF:MD) is essential to properly classify and treat different types of VWD, and it is performed using a SDS agarose gel electrophoresis followed by Western blotting, a handmade technique that demands days to be completed and requires skillful execution. Aiming both to facilitate gel production and to shorten the preparation time, we developed an uncomplicated technique to provide agility in the analysis of VWF:MD, so that it can be easily accomplished in the routine practice of hemostasis laboratories. On that account, we used a commercial vertical mini-gel electrophoresis system for SDS-PAGE and a semi-dry transfer system, which allowed us to analyze VWF:MD of various samples in a period shorter than 12 h. This technique differentiated VWF:MD in human and animal plasmas under normal, congenital and acquired (experimental envenomation by Bothrops jararaca snake) conditions. This optimized method is cheap, rapid, reproducible, easy to be performed, and uses electrophoresis and Western blotting systems available in most laboratories. All these advantages encourage hemostasis professionals to use it in their routine practices. In order to facilitate the setup and accomplishment of the whole procedure step by step, videos were appended to the article.

Optimization of von Willebrand factor multimer analysis in vertical mini-gel electrophoresis systems: a rapid procedure

Von Willebrand disease (VWD) is a common cause of bleeding worldwide. Analysis of von Willebrand factor (VWF) multimer distribution (VWF:MD) is essential to properly classify and treat different types of VWD, and it is performed using a SDS agarose gel electrophoresis followed by Western blotting, a handmade technique that demands days to be completed and requires skillful execution. Aiming both to facilitate gel production and to shorten the preparation time, we developed an uncomplicated technique to provide agility in the analysis of VWF:MD, so that it can be easily accomplished in the routine practice of hemostasis laboratories. On that account, we used a commercial vertical mini-gel electrophoresis system for SDS-PAGE and a semi-dry transfer system, which allowed us to analyze VWF:MD of various samples in a period shorter than 12?h. This technique differentiated VWF:MD in human and animal plasmas under normal, congenital and acquired (experimental envenomation by Bothrops jararaca snake) conditions. This optimized method is cheap, rapid, reproducible, easy to be performed, and uses electrophoresis and Western blotting systems available in most laboratories. All these advantages encourage hemostasis professionals to use it in their routine practices. In order to facilitate the setup and accomplishment of the whole procedure step by step, videos were appended to the article.

Optimization of von Willebrand factor multimer analysis in vertical mini-gel electrophoresis systems: a rapid procedure

Von Willebrand disease (VWD) is a common cause of bleeding worldwide. Analysis of von Willebrand factor (VWF) multimer distribution (VWF:MD) is essential to properly classify and treat different types of VWD, and it is performed using a SDS agarose gel electrophoresis followed by Western blotting, a handmade technique that demands days to be completed and requires skillful execution. Aiming both to facilitate gel production and to shorten the preparation time, we developed an uncomplicated technique to provide agility in the analysis of VWF:MD, so that it can be easily accomplished in the routine practice of hemostasis laboratories. On that account, we used a commercial vertical mini-gel electrophoresis system for SDS-PAGE and a semi-dry transfer system, which allowed us to analyze VWF:MD of various samples in a period shorter than 12?h. This technique differentiated VWF:MD in human and animal plasmas under normal, congenital and acquired (experimental envenomation by Bothrops jararaca snake) conditions. This optimized method is cheap, rapid, reproducible, easy to be performed, and uses electrophoresis and Western blotting systems available in most laboratories. All these advantages encourage hemostasis professionals to use it in their routine practices. In order to facilitate the setup and accomplishment of the whole procedure step by step, videos were appended to the article.

Salivary pH and colonization by oral Candida in children and adolescents submitted to haemodialysis.

BACKGROUND: Fungal infections are a serious problem among haemodialysis patients. AIM: The aims of this study were to estimate the frequency of oral Candida species among children and adolescents undergoing haemodialysis (HD), to identify the isolated species, and to study the relationship between haemodialysis duration, amounts of colony-forming units, and salivary pH. DESIGN: A matched sample of 52 patients undergoing HD and 52 healthy individuals were selected. The samples were obtained from the dorsum of the tongue, and the colonies were identified through a substrate assimilation test. Stimulated whole saliva was collected from each patient for evaluation of salivary pH. RESULTS: The frequency of oral Candida species was 34.6% (18/52) and 46.20% (24/52) in the HD and control groups (P = 0.23), respectively. Candida parapsilosis complex was the most frequently isolated fungi species in the HD group (P = 0.03). A HD therapy duration of more than 1 year was statistically correlated with a higher number of colony-forming units (P = 0.03) but was not statistically related to salivary pH. CONCLUSIONS: Candida parapsilosis complex was the most frequently isolated fungal species in the young HD patients, and the duration therapy was associated with higher oral colonization.

Role of Candida species from HIV infected children in enamel caries lesions: an in vitro study.

OBJECTIVES: This study analyzed the capacity of Candida spp. from dental biofilm of HIV infected (HIV+) children to demineralize primary molar enamel in vitro by Transversal Microhardness (TMH), Polarized Light Microscopy (PLM) and the quantity of calcium ions (Ca2+) released from the enamel. MATERIAL AND METHODS: Candida spp. samples were isolated from the supragingival biofilm of HIV+ children. A hundred and forty (140) enamel blocks were randomly assigned to six groups: biofilm formed by C. albicans (Group 1); mixed biofilm formed by C. albicans and C. tropicalis (Group 2); mixed biofilm formed by C. albicans and C. parapsilosis (Group 3); mixed biofilm formed by C. albicans, C. parapsilosis and C. glabrata (Group 4); biofilm formed by C. albicans ATCC (Group 5) and medium without Candida (Group 6). Enamel blocks from each group were removed on days 3, 5, 8 and 15 after biofilm formation to evaluate the TMH and images of enamel were analyzed by PLM. The quantity of Ca2+ released, from Groups 1 and 6, was determined using an Atomic Absorption Spectrophotometer. The SPSS program was used for statistical analysis and the significance level was 5%. RESULTS: TMH showed a gradual reduction in enamel hardness (p<0.05) from the 1st to 15th day, but mainly five days after biofilm formation in all groups. The PLM showed superficial lesions indicating an increase in porosity. C. albicans caused the release of Ca2+ into suspension during biofilm formation. CONCLUSION: Candida species from dental biofilm of HIV+ children can cause demineralization of primary enamel in vitro.

Role of Candida species from HIV infected children in enamel caries lesions: an in vitro study

Abstract Objectives This study analyzed the capacity of Candida spp. from dental biofilm of HIV infected (HIV+) children to demineralize primary molar enamel in vitro by Transversal Microhardness (TMH), Polarized Light Microscopy (PLM) and the quantity of calcium ions (Ca2+) released from the enamel. Material and Methods Candida spp. samples were isolated from the supragingival biofilm of HIV+ children. A hundred and forty (140) enamel blocks were randomly assigned to six groups: biofilm formed by C. albicans (Group 1); mixed biofilm formed by C. albicans and C. tropicalis (Group 2); mixed biofilm formed by C. albicans and C. parapsilosis (Group 3); mixed biofilm formed by C. albicans, C. parapsilosis and C. glabrata (Group 4); biofilm formed by C. albicans ATCC (Group 5) and medium without Candida (Group 6). Enamel blocks from each group were removed on days 3, 5, 8 and 15 after biofilm formation to evaluate the TMH and images of enamel were analyzed by PLM. The quantity of Ca2+ released, from Groups 1 and 6, was determined using an Atomic Absorption Spectrophotometer. The SPSS program was used for statistical analysis and the significance level was 5%. Results TMH showed a gradual reduction in enamel hardness (p<0.05) from the 1st to 15th day, but mainly five days after biofilm formation in all groups. The PLM showed superficial lesions indicating an increase in porosity. C. albicans caused the release of Ca2+ into suspension during biofilm formation. Conclusion Candida species from dental biofilm of HIV+ children can cause demineralization of primary enamel in vitro.

Candida species from oral cavity of HIV-infected children exhibit reduced virulence factors in the HAART era.

This study aimed to assess, in vitro, the biofilm viability and the phospholipase and protease production of Candida spp. from the saliva of HIV infected children and healthy controls, and to correlate the results with the use of medical data. A total of 79 isolates were analyzed: 48 Candida albicans isolates (33/15) and 20 Candida parapsilosis sensu lato complex isolates (12/8) (from HIV/control patients, respectively), and 8 Candida krusei, 1 Candida tropicalis, 1 Candida dubliniensis and 1 Candida guilliermondii from HIV patients. The XTT (2, 3-bis (2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-Carboxanilide) reduction assay analyzed the biofilm viability. Phospholipase and protease assays were performed using the egg yolk and Bovine Serum Albumin agar plate methods, respectively. All isolates were able to form biofilm with cell viability. Quantitatively, Candida isolates from both groups presented a similar ability to form biofilm (p > 0.05). The biofilm viability activity was higher in C. albicans isolates than in non-albicans Candida isolates (p < 0.05) for both groups. Phospholipase activity was detected in 32 isolates (40.5%) and it was significantly higher in the HIV group (p = 0.006). Protease activity was detected in 66 isolates (84.8%) and most of them were relatively/very strong producers. No statistical association with medical data was found in the HIV group. Although Candida spp. isolates from HIV-positive children presented higher phospholipase production, in vitro they exhibited reduced virulence factors compared to isolates from healthy individuals. This finding may enlighten the role played by immunosuppression in the modulation of Candida virulence attributes.

Antiproliferative and ultrastructural effects of phenethylamine derivatives on promastigotes and amastigotes of Leishmania (Leishmania) infantum chagasi.

Leishmania (Leishmania) infantum chagasi is one of the agents that cause visceral leishmaniasis. This disease occurs more frequently in third world countries, such as Brazil. The treatment is arduous, and is dependent on just a few drugs like the antimonial derivatives and amphotericin B. Moreover, these drugs are not only expensive, but they can also cause severe side effects and require long-term treatment. Therefore, it is very important to find new compounds that are effective against leishmaniasis. In the present work we evaluated a new group of synthetic amides against the promastigote and amastigote forms of L. infantum chagasi. The results showed that one of these amides in particular, presented very effective activity against the promastigotes and amastigotes of L. infantum chagasi at low concentrations and it also presented low toxicity for mammal cells, which makes this synthetic amide a promising drug for combating leishmaniasis.

Association of Candida Species Isolated From the Dental Plaque of HIV-infected Children and Prevalence of Early Carious Lesions.

PURPOSE: The purposes of this study were to: (1) quantify and identify Candida species (spp.) in the dental plaque of children infected with the human immunodeficiency virus (HIV) and compare with noninfected children; and (2) determine the association of Candida spp. with the presence of caries. METHODS: Seventy HIV-infected and 55 non-HIV-infected three- to 12-year-old children were examined to determine caries prevalence. After a visual inspection, supragingival plaque was collected from the cervical region using standard dental curettes. The material was transferred to microtubes and submitted for analysis to identify and quantify the presence of Candida spp. RESULTS: Candida spp. were more prevalent in the HIV-infected group (72.9 percent) than in the control group (20 percent), and the most prevalent specie was Candida albicans. Caries was found in 72.9 percent of the HIV-infected group and in 58.2 percent of the control group, but a significant difference was only found in the presence of active white spot lesions between the groups. CONCLUSIONS: The dental plaque of HIV-infected children was colonized by Candida species to a much greater extent than that of non-HIV-infected children, and this colonization was significantly associated with the prevalence of early carious lesions in enamel.

Role of saliva in the caries experience and calculus formation of young patients undergoing hemodialysis.

OBJECTIVES: The aims of this study were to investigate the caries experience, periodontal status, oral hygiene habits, and salivary parameters of children and adolescents undergoing hemodialysis (HD) and to compare them with their healthy counterparts. METHODS: Fifty-two HD patients were matched for age, sex, ethnicity, and social class with 52 healthy subjects for analysis of the number of decayed, missing and filled teeth, plaque and gingival index, dental calculus accumulation, measurements of pocket depth, clinical attachment level, gingival recession, and bleeding on probing. Stimulated saliva samples were collected to assess salivary flow rate, pH and buffer capacity, and salivary concentrations of calcium, phosphate, and urea by colorimetric method. RESULTS: HD patients had lower dental caries (p = 0.004), greater plaque and calculus accumulation (p = 0.001), and reported flossing less often than the controls (p = 0.013). Regarding salivary analysis, HD patients showed significantly higher values of pH, buffer capacity, and salivary urea concentration when compared to the controls (p = 0.001). CONCLUSION: HD patients had lower caries experience, higher accumulation of dental plaque, and calculus deposition than their healthy counterparts, probably due to the differences found in their salivary biochemical parameters. CLINICAL SIGNIFICANCE: A significant number of children and adolescents undergoing hemodialysis are candidates for kidney transplantation and should receive complete pre-transplant dental exams and dental treatment. Our results open the way for the development of an individualized dental protocol for these patients with preventive measures and treatment of the poor oral health in HD patients.

ABO blood group in primary antiphospholipid syndrome: influence in the site of thrombosis?

Antiphospholipid syndrome (APS) is characterized by vascular thrombosis and/or obstetric complications associated with presence of antiphospholipid antibodies (aPL) but additional factors would also induce thrombosis. ABO (H) blood groups are known to be closely related to thrombosis, especially non-O blood type with venous events. The aim of this study was to investigate possible role of ABO (H) blood types in the thrombotic events in primary APS (PAPS). Seventy PAPS patients were selected for the study and were divided according to ABO blood group in: O PAPS (n = 26) and non-O PAPS (n = 44). ABO blood group phenotyping was performed by indirect technique. aPL anticardiolipin (aCL) and anti-ßeta2 glycoprotein-1 (aß2GPI) and the concentrations and activities of von Willebrand factor (VWF) were measured with ELISA. Lupus anticoagulant (LA) was detected by coagulation assays. A significant higher frequency of venous events was observed in non-O PAPS group (72.7 vs. 46.2 %, p = 0.040). In contrast, the frequency of arterial events was significantly higher in the O PAPS compared to the non-O PAPS group (69.2 vs. 36.4 %, respectively; p = 0.013). Frequencies of aCL, LA, aß2GPI and triple aPL positivity were similar in both groups (p > 0.05). VWF antigen (75.54 ± 8.68 vs. 79.51 ± 7.07 IU/dl, p = 0.041) and activity (70.23 ± 11.96 vs. 77.92 ± 13.67 %, p = 0.020) were decreased in O PAPS compared to non-O blood group. VWF:CB/VWF:Ag ratio was similar among groups (p > 0.05). This is the first report that confirms the role of ABO blood system in thrombosis of PAPS and suggests that non-O blood group was related with venous events and O blood group with arterial thrombosis.

Resins-based denture soft lining materials modified by chlorhexidine salt incorporation: an in vitro analysis of antifungal activity, drug release and hardness.

OBJECTIVES: To evaluate the in vitro growth inhibition of Candida albicans, the rate of chlorhexidine release and shore A hardness from resins-based denture soft lining materials modified by chlorhexidine diacetate (CDA) or chlorhexidine hydrochloride (CHC) incorporation. METHODS: Resin discs were prepared from soft denture liners based on poly (methyl methacrylate) (PMMA) or poly (ethyl methacrylate) (PEMA) containing 0.5, 1.0 and 2.0 wt.% of CDA or CHC. For antifungal activity resin discs were placed on agar plates inoculated with C. albicans, after 48 h at 37°C the diameters of inhibition zones were measured. For the chlorhexidine release, discs were immersed into distilled water at 37°C, and spectral measurements were made after 48 h. Shore A hardness was evaluated at the baseline, 2 and 7 days, using 6mm thick rectangular specimens also immersed into distilled water at 37°C. Data were statistically processed by SigmaStat software using ANOVA and all pairwise multiple comparison procedures was done using the Holm-Sidak method, with α=0.05 (p<0.001). RESULTS: CDA added to PMMA soft liner and PEMA soft liner had a dose-related inhibitory effect on C. albicans and on chlorhexidine release rate (p<0.001). The PMMA and PEMA hardness increased statistically by time but not for the different CDA concentrations. CHC had no inhibitory effect on C. albicans. SIGNIFICANCE: Chlorhexidine diacetate released from resins-based soft lining materials can be convenient to reduce the biofilm development on the material surface and treat denture stomatitis, without depending on patient compliance.

Effect of the secretory leucocyte proteinase inhibitor (SLPI) on Candida albicans biological processes: a therapeutic alternative?

OBJECTIVES: The aim of this study was to evaluate the effect of SLPI on the growth and biological processes of Candida albicans. METHODS: Two C. albicans strains were used in this study, a clinical isolate resistant to fluconazole (PRI) and a reference strain ATCC 24433. The minimal inhibitory concentration (MIC) was determined according to the CLSI methodology. The influence of SLPI on secreted serine proteinase activities (SSP) was measured by the cleavage of specific substrate, and surface hydrophobicity was determined by the aqueous-hydrocarbon biphasic separation method. Flow cytometry was performed to investigate receptors for SLPI and variations in the cell wall mannoprotein expression. Interaction between yeast and epithelium was assessed using the MA-104 cells lineage. Ultrastructure was analyzed by transmission electron microscopy (TEM). RESULTS: MIC values were calculated as 18 and 18.9µM for the PRI and ATCC 24433, respectively. SSP activity was reduced by 48.8% by 18µM of SLPI and cell surface hydrophobicity increased by 11.1%. Flow cytometry suggest the existence of SLPI binding sites on the surface of the yeast. Results showed a reduction in the expression of mannoproteins in 20.8% by the cells treated with 80µM of SLPI, and 18µM reduced the adhesion of yeasts to mammalian cells in 60.1%. TEM revealed ultrastructural changes in cells treated with 80µM of SLPI, such as the presence of membrane-like structures within the cytoplasm. CONCLUSIONS: SLPI exerts a significant influence on C. albicans viability and biological processes. Considering its constitutive and physiologic features, SLPI may become a promising tool for the development of new methodologies for the treatment and control of candidiasis.

Salivary lactoferrin in HIV-infected children: correlation with Candida albicans carriage, oral manifestations, HIV infection and its antifungal activity.

OBJECTIVES: This study aimed to evaluate the concentration of lactoferrin in the saliva of HIV infected and healthy children and analyze the associations between lactoferrin levels, Candida sp. colonization, oral manifestations and medical data. Also, the antifungal ability of lactoferrin to inhibit the growth of Candida albicans isolated from saliva of these children was investigated in vitro. SUBJECTS AND METHODS: Saliva was collected from 70 HIV-infected and 50 healthy children, followed by oral manifestation assessments. The salivary lactoferrin was quantified using an ELISA Kit. The salivary samples were cultured and the Candida spp. colonies counted and then identified by sugar assimilation and fermentation. The antifungal activity of lactoferrin was analyzed in vitro with 10 isolates of C. albicans from each group. RESULTS: The HIV infected children (mean age 9.8 ± 2.8) had higher lactoferrin levels (median 6.13 µg/ml (3.58-7.89)) and were colonized three times more by Candida sp. than the control group (mean age 9.4 ± 2.4) (median 5.74 µg/ml (3.12-6.86)) (p=0.003). Statistical associations were found considering the salivary lactoferrin levels and Candida sp. and oral manifestations between the groups. No associations between lactoferrin concentrations and oral manifestations, immunosuppression, presence of AIDS and use of HAART were observed in the HIV group. The percentage of dead C. albicans due to lactoferrin was inversely proportional to C. albicans cell density for both groups (p<0.001). CONCLUSIONS: HIV-infected children have higher concentrations of lactoferrin and it was associated with Candida sp. colonization but no association with medical data was found. Also, both groups showed similar lactoferrin antifungal activity.