Atividade antifúngica de nanofibras de policaprolactona/nistatina: estudo in vitro e in vivo / Antifungal activity of polycaprolactone/nystatin nanofibers: in vitro and in vivo study

Este estudo avaliou a eficácia in vitro e in vivo de mantas de nanofibras (NF) de policaprolactona (PCL) incorporadas com nistatina (NIS) no tratamento da estomatite protética (EP) em modelos animais. NF foram sintetizadas com diferentes concentrações de NIS, totalizando quatro soluções: PCL puro, PCL/NIS 0,045 g, PCL/NIS 0,090 g e PCL/NIS 0,225 g. A liberação da NIS foi analisada por espectroscopia Ultravioleta-Visível. A capacidade das mantas de inibirem o biofilme de Candida albicans, principal fator etiológico da EP, dividindo-se cinco grupos (N=5) compostos por um grupo com controle de células de C. albicans e com PCL puro, além das três concentrações de NIS. A seguir, foi analisada a viabilidade celular em queratinócitos humanos (HaCat) por meio do teste colorimétrico de resazurina. Cinco grupos foram divididos (N=10): controle celular, PCL puro e as três concentrações de NIS. Em modelos animais de ratos Wistar albinos (N=18), dispositivos palatinos (DP) de resina acrílica foram confeccionados simulando próteses totais e utilizados para a indução da EP. Para isso, DP contaminados com C. albicans foram cimentados na região molar da cavidade bucal dos animais e permaneceram em boca por 48 h. Após esse período, os DP foram removidos e os animais foram divididos em três grupos: (C) controle; (B1) com tratamento por mantas de PCL/NIS 0,045 g e (B2) PCL/NIS 0,225 g, com N=6. Então novos DP, livres de contaminação, foram cimentados na cavidade oral dos animais e permaneceu por mais 48 h. Após esse período, os animais foram eutanasiados, a contagem de UFC/ mL foi realizada e os palatos foram coletados para a análise histológica. A curva padrão de NIS obtida apresentou R2 de 0,99. As três concentrações de NF apresentaram liberação de NIS, com pico no tempo de 6 h e valores de 66,26 µg/ mL para PCL/NIS 0,045 g, de 333,87 µg/ mL para PCL/NIS 0,090 g e 436,51 µg/ mL para PCL/NIS 0,225 g, constantes até o fim do experimento. Os grupos com NIS reduziram em 2,5 log10 de crescimento do biofilme fúngico em relação aos grupos sem tratamento, Controle e PCL, sem diferença estatística significativa. Não foi observada citotoxicidade nas células HaCat, com viabilidade celular de 93,7% para PCL/NIS 0,045 g, 72,6% para PCL/NIS 0,090 g e 72,4% para PCL/NIS 0,225 g. A indução da EP nos três grupos foi possível e, porém, sem redução significativa na contagem de UFC/ mL de C. albicans nos grupos B1 e B2. Na análise histológica do grupo C pôde-se observar infiltração de hifas de Candida na camada queratinizada, presença de células inflamatórias formando micro abscessos e um discreto infiltrado inflamatório no tecido conjuntivo subjacente ao epitélio infectado. Nos grupos B1 e B2 não foram encontradas alterações epiteliais, concluindo-se que as NF demonstraram atividade antifúngica in vitro e foram efetivas na prevenção da penetração de hifas no tecido palatino de animais com DP (AU)

This study evaluated the in vitro and in vivo efficacy of nanofiber (NF) mats of polycaprolactone (PCL) incorporated with nystatin (NIS) in the treatment of denture stomatitis (DS) in animal models. NFs were synthesized with different concentrations of NIS, totaling four solutions: pure PCL, PCL/NIS 0.045 g, PCL/NIS 0.090 g, and PCL/NIS 0.225 g. The release of NIS was analyzed by Ultraviolet-Visible spectroscopy. The ability of the mats to inhibit Candida albicans biofilm, the main etiological factor of DS, was assessed by dividing five groups (N=5) composed of a group with C. albicans cell control and with pure PCL, in addition to the three concentrations of NIS. Next, cell viability in human keratinocytes (HaCat) was analyzed using the resazurin colorimetric test. Five groups were divided (N=10): cell control, pure PCL, and the three concentrations of NIS. In albino Wistar rat animal models (N=18), palatal devices (PD) made of acrylic resin were fabricated to simulate total prostheses and used to induce DS. For this, PD contaminated with C. albicans were cemented in the molar region of the animals' oral cavity and remained in the mouth for 48 hours. After this period, the PDs were removed, and the animals were divided into three groups: (C) control; (B1) treated with PCL/NIS 0.045 g mats, and (B2) PCL/NIS 0.225 g, with N=6. Then new, uncontaminated PDs were cemented in the animals' oral cavity and remained for another 48 hours. After this period, the animals were euthanized, UFC/ mL counts were performed, and the palates were collected for histological analysis. The standard NIS curve obtained showed an R2 of 0.99. The three concentrations of NF showed NIS release, with a peak at 6 h and values of 66.26 µg/ mL for PCL/NIS 0.045 g, 333.87 µg/ mL for PCL/NIS 0.090 g, and 436.51 µg/ mL for PCL/NIS 0.225 g, remaining constant until the end of the experiment. The groups with NIS reduced fungal biofilm growth by 2.5 log10 compared to the untreated groups, Control and PCL, with no significant statistical difference. No cytotoxicity was observed in HaCat cells, with cell viability of 93.7% for PCL/NIS 0.045 g, 72.6% for PCL/NIS 0.090 g, and 72.4% for PCL/NIS 0.225 g. Induction of DS in the three groups was possible; however, there was no significant reduction in UFC/ mL counts of C. albicans in groups B1 and B2. Histological analysis of group C revealed infiltration of Candida hyphae in the keratinized layer, presence of inflammatory cells forming micro abscesses, and a discreet inflammatory infiltrate in the connective tissue underlying the infected epithelium. No epithelial alterations were found in groups B1 and B2, concluding that NFs demonstrated in vitro antifungal activity and were effective in preventing hyphal penetration into palatal tissue in animals with PD.(AU)

Comparison of tea tree oil 5%, tea tree oil 10%, and nystatin inhibition zones against vaginal Candida isolates in pregnancy.

INTRODUCTION: Vulvovaginal candidiasis (VVC) in pregnancy frequently develops into recurrent infections. Clinical study suggests that conventional topical treatments for VVC are not always enough to eradicate Candida spp. from the vaginal microenvironment. This study aimed to evaluate the antifungal activity of tea tree oil (TTO) 5% and TTO 10% against Candida species causing VVC in pregnancy. METHODOLOGY: In vitro experimental study was conducted in the Mycology Laboratory at Dermatovenereology Outpatient Clinic Dr. Soetomo General Hospital Surabaya. Eighteen isolates of Candida species were isolated from the vaginal thrush of 15 pregnant women diagnosed with VVC from March to May 2021. Antifungal susceptibility of TTO 5% and TTO 10% was evaluated by the disc diffusion method, with the inhibitory zone diameter as the main outcome. RESULTS: The mean inhibitory zone diameter of TTO 5%, TTO 10%, and nystatin against all Candida spp. was 7.26 mm, 8.64 mm, and 25.57 mm, respectively (p < 0.001). The mean inhibitory zone diameter of TTO 5%, TTO 10%, and nystatin tend to be larger in C. albicans compared to the non-albicans, but the difference is not significant. Nystatin displayed the largest mean inhibitory zone diameters compared to TTO 5% and TTO 10% (p < 0.001) in all Candida species. Increased concentration from TTO 5% to TTO 10% resulted in a slight increment in the mean inhibitory zone diameters in all-Candida species (p = 0.001). CONCLUSIONS: Tea Tree Oil displayed antifungal activity against Candida species causing VVC in pregnancy. Further studies are required to investigate optimal TTO concentrations as a VVC treatment in pregnancy.

CYP broth: a tool for Yersinia pestis isolation in ancient culture collections and field samples.

We developed a simple new selective LB-based medium, named CYP broth, suitable for recovering long-term stored Y. pestis subcultures and for isolation of Y. pestis strains from field-caught samples for the Plague surveillance. It aimed to inhibit the growth contaminating microorganisms and enrich Y. pestis growth through iron supplementation. The performance of CYP broth on microbial growth from different gram-negative and gram-positive strains from American Type Culture Collection (ATCC®) and other clinical isolates, field-caught rodent samples, and more importantly, on several vials of ancient Y. pestis subcultures was evaluated. Additionally, other pathogenic Yersinia species such as Y. pseudotuberculosis and Y. enterocolitica were also successfully isolated with CYP broth. Selectivity tests and bacterial growth performance on CYP broth (LB broth supplemented with Cefsulodine, Irgasan, Novobiocin, nystatin and ferrioxamine E) were evaluated in comparison with LB broth without additive; LB broth/CIN, LB broth/nystatin and with traditional agar media including LB agar without additive, and LB agar and Cefsulodin-Irgasan-Novobiocin Agar (CIN agar) supplemented with 50 µg/mL of nystatin. Of note, the CYP broth had a recovery twofold higher than those of the CIN supplemented media or other regular media. Additionally, selectivity tests and bacterial growth performance were also evaluated on CYP broth in the absence of ferrioxamine E. The cultures were incubated at 28 °C and visually inspected for microbiological growth analysis and O.D.625 nm measurement between 0 and 120 h. The presence and purity of Y. pestis growth were confirmed by bacteriophage and multiplex PCR tests. Altogether, CYP broth provides an enhanced growth of Y. pestis at 28 °C, while inhibiting contaminant microorganisms. The media is a simple, but powerful tool to improve the reactivation and decontamination of ancient Y. pestis culture collections and for the isolation of Y. pestis strains for the Plague surveillance from various backgrounds. KEY POINTS: • The newly described CYP broth improves the recuperation of ancient/contaminated Yersinia pestis culture collections • CYP broth was also efficient in reducing environmental contamination in field-capture samples, improving Y. pestis isolation • CYP broth can also be used for the isolation of Y. enterocolitica and Y. pseudotuberculosis.

Alginate oligosaccharides enhance the antifungal activity of nystatin against candidal biofilms.

Background: The increasing prevalence of invasive fungal infections in immuno-compromised patients is a considerable cause of morbidity and mortality. With the rapid emergence of antifungal resistance and an inadequate pipeline of new therapies, novel treatment strategies are now urgently required. Methods: The antifungal activity of the alginate oligosaccharide OligoG in conjunction with nystatin was tested against a range of Candida spp. (C. albicans, C. glabrata, C. parapsilosis, C. auris, C. tropicalis and C. dubliniensis), in both planktonic and biofilm assays, to determine its potential clinical utility to enhance the treatment of candidal infections. The effect of OligoG (0-6%) ± nystatin on Candida spp. was examined in minimum inhibitory concentration (MIC) and growth curve assays. Antifungal effects of OligoG and nystatin treatment on biofilm formation and disruption were characterized using confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM) and ATP cellular viability assays. Effects on the cell membrane were determined using permeability assays and transmission electron microscopy (TEM). Results: MIC and growth curve assays demonstrated the synergistic effects of OligoG (0-6%) with nystatin, resulting in an up to 32-fold reduction in MIC, and a significant reduction in the growth of C. parapsilosis and C. auris (minimum significant difference = 0.2 and 0.12 respectively). CLSM and SEM imaging demonstrated that the combination treatment of OligoG (4%) with nystatin (1 µg/ml) resulted in significant inhibition of candidal biofilm formation on glass and clinical grade silicone surfaces (p < 0.001), with increased cell death (p < 0.0001). The ATP biofilm disruption assay demonstrated a significant reduction in cell viability with OligoG (4%) alone and the combined OligoG/nystatin (MIC value) treatment (p < 0.04) for all Candida strains tested. TEM studies revealed the combined OligoG/nystatin treatment induced structural reorganization of the Candida cell membrane, with increased permeability when compared to the untreated control (p < 0.001). Conclusions: Antimicrobial synergy between OligoG and nystatin against Candida spp. highlights the potential utility of this combination therapy in the prevention and topical treatment of candidal biofilm infections, to overcome the inherent tolerance of biofilm structures to antifungal agents.

Clinical efficacy of 5-aminolevulinic acid-mediated photodynamic therapy versus topical antifungal agent and surgical excision for the treatment of hyperplastic candidiasis.

BACKGROUND: Hyperplastic candidiasis (HC) is a chronic infection of oral mucosa caused by Candida. Owing to its potential for malignant transformation, its intervention requires attention. Conventional surgical resection might lead to irreversible damage and impact the patient's quality of life. Hence, this study aimed to evaluate the clinical efficacy of 5-aminolevulinic acid (5-ALA)-mediated photodynamic therapy (PDT) alone and in combination with topical antifungal therapy (i.e., nystatin [combination therapy]) in comparison with nystatin and surgical resection for the treatment of HC. METHODS: Forty subjects with clinical and histopathological diagnoses of HC were included in the study. Four study groups, with 10 participants each, were formed as follows: Group-I - receiving antifungal agent [nystatin]; Group-II - receiving surgical resection; Group-III - receiving PDT; and Group-IV - receiving 5-ALA-mediated PDT and nystatin [combination therapy]. Salivary and mucosal samples were collected for the quantification of Candida albicans and the treatment responses to different interventions were recorded at week-4, week-6, and week-8 after finishing therapies. RESULTS: At the 3rd follow-up (i.e., at end of the 8th week after the interventions), complete improvement in 3 (30%), 2 (20%), 1 (10%), and 5 (50%) patients in group-I, group-II, group-III, and group-IV, respectively was observed. A statistically significant difference was obtained when the intervention responses were compared at week-4 (p<0.01), week-6 (p<0.01), and week-8 (p<0.0001) follow-ups in group-I, group-II, and group-IV subjects. At the 8-week follow-up regarding the salivary and mucosal samples, the lowest colony-forming units/milliliter score of C. albicans was observed in group-IV subjects. CONCLUSION: The application of 5-ALA-mediated PDT in combination with nystatin gel possesses the potential as a well-tolerated and safe therapeutic modality for the treatment of patients with HC.

Cyclic and Linear Peptides Containing Alternate WW and RR Residues as Molecular Cargo Delivery Tools.

Cell-impermeable and negatively charged compounds' cellular uptake across the cell membranes remains challenging. Herein, the synthesis of four linear [(WWRR)2, (WWRR)3, (WWRR)4, and (WWRR)5] and four cyclic ([WWRR]2, [WWRR]3, [WWRR]4, and [WWRR]5) peptides containing alternate two tryptophan (WW) and two arginine (RR) residues and their biological evaluation as molecular transporters are reported. The peptides did not show any significant cytotoxicity in different cell lines (MDA-MB-23, SK-OV-3, and HEK 293) at a concentration of 5 µM and after 3 h of incubation time. The uptake of fluorescence-labeled cargo molecules (F'-GpYEEI, F'-siRNA, and F'-3TC) in the presence of the peptides was monitored in different cell lines (SK-OV-3 and MDA-MB-231) with fluorescence-activated cell sorting. Among all the peptides, [WWRR]5 (C4) showed the highest cellular uptake of cargo molecules, indicating it can act as effective molecular transporter. Confocal microscopy in MDA-MB-231 cells showed the cellular uptake of F'-GpYEEI in the presence of C4 and the intracellular localization of fluorescence-labeled C4 (F'-C4) in the cytosol. The F'-C4 cellular uptake was found to be concentration- and time-dependent, as shown by flow cytometry in MDA-MB-231 cells. Confocal microscopy and flow cytometry of F'-C4 in MDA-MB-231 cells were examined alone and in the presence of different endocytosis inhibitors (chlorpromazine, methyl-ß-cyclodextrin, chloroquine, and nystatin). The data showed that the cellular uptake of F'-C4 in the presence of chlorpromazine, chloroquine, and methyl-ß-cyclodextrin was reduced but not completely eliminated, indicating that both energy-independent and energy-dependent pathways contributed to the cellular uptake of F'-C4. Similar results were obtained using the confocal microscopy of C4 and F'-GpYEEI in the presence of endocytosis inhibitors (chlorpromazine, methyl-ß-cyclodextrin, chloroquine, and nystatin). These data indicate that C4 has the potential to be used as a cell-penetrating peptide and cargo transporter.

A sensitive NIR mitochondria-targeting fluorescence probe for visualizing viscosity in living cells and mice.

Mitochondria are the powerhouses in cells, providing the energy needed for cellular activities. However, the abnormalities in the mitochondrial microenvironment (e.g., the increased viscosity) can lead to mitochondrial dysfunctions and diseases. Herein, we develop a series of near-infrared (NIR) fluorescence probes for the detection of viscosity. After screening, probe CQ-4 is selected since it shows a great fluorescence enhancement (89-fold) in the NIR window. Its specific response to viscosity is not influenced by pH, polarity and biological species. Under stimulation with monensin or nystatin, CQ-4 can measure the cellular viscosity changes with good biocompatibility. In addition, we can observe an increase of viscosity during starvation. CQ-4 is applied to distinguish cancer cells from normal cells based on the viscosity differences. Furthermore, the probe has been successfully applied to image viscosity in inflamed and tumor-bearing mice in vivo. Therefore, CQ-4 may contribute to the future study about viscosity in the physiological and pathological processes.

In vivo antifungal activity and biocompatibility of Cryptocarya moschata.

The objective of this study was evaluate, in vivo model, the antifungal activity of Cryptocarya moschata extract against Candida albicans and its biocompatibility. The animals (N = 50) were divided into groups (n = 5): CI/CG: candidiasis was induced and treated with C. moschata extract (0.045 g/mL); CI/NG: candidiasis was induced and treated with nystatin; CI/NT: candidiasis was induced and no treated; CI/CG-2: candidiasis was induced and treated with C. moschata extract (0.045 g/mL), reapplied after 24 h; CI/NG-2: candidiasis was induced and treated with nystatin, reapplied after 24 h; NCI/NT: candidiasis was not induced and no treated; NCI/CG: candidiasis was not induced and treated with C. moschata extract (0.045 g/mL); NCI/NG: candidiasis was not induced treated with nystatin; NCI/CG-2: candidiasis was not induced and treated with C. moschata extract (0.045 g/mL), reapplied after 24 h; NCI/NG-2: candidiasis was not induced and treated with nystatin, reapplied after 24 h. The fungi present in the lingual dorsum of mice were collected and analyzed by the count of colony-forming units. In addition, histological analysis was performed. Histologically, there was no cell damage in the mice's tongue, and there was a decrease in Candida biofilm, similar to the use of nystatin. It was concluded that the C. moschata extract was effective against C. albicans and was biocompatible.

FXYD3 facilitates Na+ and liquid absorption across human airway epithelia by increasing the transport capacity of the Na/K ATPase.

Na/K ATPase activity is essential for ion transport across epithelia. FXYD3, a γ subunit of the Na/K ATPase, is expressed in the airway, but its function remains undetermined. Single-cell RNA sequencing and immunohistochemistry revealed that FXYD3 localizes within the basolateral membrane of all airway epithelial cells. To study FXYD3 function, we reduced FXYD3 expression using siRNA. After permeabilizing the apical membrane with nystatin, epithelia pretreated with FXYD3-targeting siRNA had lower ouabain-sensitive short-circuit currents than control epithelia. FXYD3-targeting siRNA also reduced amiloride-sensitive short-circuit currents and liquid absorption across intact epithelia. These data are consistent with FXYD3 facilitating Na+ and liquid absorption. FXYD3 may be needed to maintain the high rates of Na+ and fluid absorption observed for airway and other FXYD3-expressing epithelia.

Anti-biofilm properties of eucalyptol in combination with antifungals against Candida albicans isolates in patients with hematological malignancy.

Oral candidiasis is a fungal infection caused mainly by Candida albicans and it is a major problem among hematologic malignancy patients. Biofilm formation is an attributable factor to both virulence and drug resistance of Candida species. The aim of the study was to evaluate the biofilm-producing ability of oral C. albicans isolates and to evaluate the inhibitory activity of eucalyptol on Candida biofilm, alone and in combination with antifungal agents. Samples were collected from the oral cavity of 106 patients with hematologic malignancy. The isolated yeasts were identified by PCR-sequencing. Then C. albicans isolates were analyzed for their biofilm-producing ability by crystal violet staining and MTT assay. The minimum biofilm inhibition concentrations (MBIC) of eucalyptol, amphotericin B, itraconazole, and nystatin and the in vitro interaction of eucalyptol with these drugs were tested according to CLSI-M-27-A3 protocol and checkerboard methods, respectively. From 106 patients, 50 (47.2%) were confirmed for oral candidiasis [mean ± SD age 39 ± 14 years; female 31 (62%) and male 19 (38%)]. C. albicans was isolated from 40 of 50 (80%) patients. From 40 C. albicans isolates, 24 (60%) and 16 (40%) were moderate and weak biofilm producer, respectively. The geometric mean MBIC of amphotericin B, itraconazole, nystatin and eucalyptol were 3.93 µg/mL, 12.55 µg/mL, 0.75 µg/mL and 798 µg/mL, respectively. Eucalyptol interacted synergistically with amphotericin B, itraconazole and nystatin against 12.5, 10, and 22.5% of isolates, respectively. Eucalyptol demonstrated promising activity against biofilm of C. albicans when tested alone or combined with antifungal drugs.

Association of the essential oil of Cymbopogon citratus (DC) Stapf with nystatin against oral cavity yeasts.

Oral thrush or candidiasis is a fairly common infectious fungal disease that is mainly caused by species of the genus Candida spp. In this work, we evaluated the antifungal activity of the essential oil of Cymbopogon citratus (DC) Stapf (lemongrass) and its combination with nystatin against oral cavity yeasts. The oil was extracted by the steam distillation method, and its constituents were quantified. The yeasts were identified using MALDI-TOF MS. The sensitivity to the essential oil and its association with nystatin was verified by the minimum inhibitory concentration (MIC) and checkerboard methods. In the essential oil samples, there were 84.53% of citral. A total of 64.77% of strains were identified as Candida albicans. On susceptibility tests, 83.55% of the yeast isolates were inhibited in concentrations of ≥16 µg/mL of nystatin. C. citratus's oil was capable of inhibiting and killing all the isolates tested with concentrations that varied from 0.137 to 2.2 mg/mL. The association of oil and nystatin had an additive effect on more than 78% of the strains. The association of this herbal drug with nystatin potentialized the antifungal effect on yeast samples isolated from the oral cavity of oncologic patients.

Origanum vulgare L. essential oil inhibits virulence patterns of Candida spp. and potentiates the effects of fluconazole and nystatin in vitro.

BACKGROUND: Recurrence and resistance of Candida spp. infections is associated with the ability of these microorganisms to present several virulence patterns such as morphogenesis, adhesion, and biofilm formation. In the search for agents with antivirulence activity, essential oils could represent a strategy to act against biofilms and to potentiate antifungal drugs. OBJECTIVE: To evaluate the antivirulence effect of Origanum vulgare L. essential oil (O-EO) against Candida spp. and to potentiate the effect of fluconazole and nystatin. METHODS: The effect of O-EO was evaluated on ATCC reference strains of C. albicans and non-albicans Candida species. Minimum inhibitory concentration (MIC) was determined through broth microdilution assay. Adhesion to microplates was determined by crystal violet (CV) assay. An adapted scratch assay in 24-well was used to determine the effect of essential oil on biofilms proliferation. Viability of biofilms was evaluated by MTT reduction assay and through a checkerboard assay we determined if O-EO could act synergistically with fluconazole and nystatin. RESULTS: MIC for C. albicans ATCC-90029 and ATCC-10231 was 0.01 mg/L and 0.97 mg/L, respectively. For non-albicans Candida strains MIC values were 2.6 mg/L for C. dubliniensis ATCC-CD36 and 5.3 mg/L for C. krusei ATCC-6258. By using these concentrations, O-EO inhibited morphogenesis, adhesion, and proliferation at least by 50% for the strains assayed. In formed biofilms O-EO decreased viability in ATCC 90029 and ATCC 10231 strains (IC50 7.4 and 2.8 mg/L respectively). Finally, we show that O-EO interacted synergistically with fluconazole and nystatin. CONCLUSIONS: This study demonstrate that O-EO could be considered to improve the antifungal treatment against Candida spp.

Prolonged ethanol exposure modulates constitutive internalization and recycling of 5-HT1A receptors.

Alcohol exposure alters the signaling of the serotoninergic system, which is involved in alcohol consumption, reward, and dependence. In particular, dysregulation of serotonin receptor type 1A (5-HT1AR) is associated with alcohol intake and withdrawal-induced anxiety-like behavior in rodents. However, how ethanol regulates 5-HT1AR activity and cell surface availability remains elusive. Using neuroblastoma 2a cells stably expressing human 5-HT1ARs tagged with hemagglutinin at the N-terminus, we found that prolonged ethanol exposure (18 h) reduced the basal surface levels of 5-HT1ARs in a concentration-dependent manner. This reduction is attributed to both enhanced receptor internalization and attenuated receptor recycling. Moreover, constitutive 5-HT1AR internalization in ethanol naïve cells was blocked by concanavalin A (ConA) but not nystatin, suggesting clathrin-dependent 5-HT1AR internalization. In contrast, constitutive 5-HT1AR internalization in ethanol-treated cells was blocked by nystatin but not by ConA, indicating that constitutive 5-HT1AR internalization switched from a clathrin- to a caveolin-dependent pathway. Dynasore, an inhibitor of dynamin, blocked 5-HT1AR internalization in both vehicle- and ethanol-treated cells. Furthermore, ethanol exposure enhanced the activity of dynamin I via dephosphorylation and reduced myosin Va levels, which may contribute to increased internalization and reduced recycling of 5-HT1ARs, respectively. Our findings suggest that prolonged ethanol exposure not only alters the endocytic trafficking of 5-HT1ARs but also the mechanism by which constitutive 5-HT1AR internalization occurs.

Clinical and morphopathological assay in vulvovaginal candidiasis.

Candida vulvovaginitis is characterized by the appearance of inflammatory changes in the vaginal and vulvar epithelium secondary to infection with Candida species. The purpose of this study was to analyze and compare the clinical, microbiological, and histopathological aspects of pregnant and non-pregnant patients, symptomatic or asymptomatic in the case of candida vaginitis and to correlate the microscopic aspects with the symptoms before applying the local treatment with Nystatin. The study presents a retrospective analysis of the management of vaginitis in 166 pregnant or non-pregnant patients during 2021-2022. We observed the structure of the Malpighian squamous epithelium without keratinization present on the vaginal mucosa and the structure of the subepithelial connective tissue, which shows increased numerical values of inflammatory and vascular cellularity in the case of candida vaginitis symptomatic compared to asymptomatic ones. We noticed also in the microscopic study that in cases of asymptomatic patients before treatment, the number of inflammatory cells and blood vessels situated immediately under the epithelium was significantly lower compared to their number in symptomatic patients before treatment. Analyzing the results obtained after the administration of the treatment proposed by us, we can say that local Nystatin treatment is beneficial and safe for pregnant and non-pregnant patients and is a good alternative for patients with recurrent vulvovaginal candidiasis.

Nystatin is commonly prescribed as prophylaxis in children beyond the neonatal age.

The indications for nystatin as prophylaxis or treatment are limited. In the PASOAP (Pediatric Antifungal Stewardship Optimizing Antifungal Prescription) study, high use of nystatin in hospitalized children beyond the neonatal age was observed. In this report, we present the data on nystatin use in infants and children ≥ 3 months who participated in the PASOAP study. Nystatin was prescribed mainly for prophylaxis. Congenital heart disease, cystic fibrosis, and chronic renal disease were the most commonly reported conditions in children receiving prophylactic nystatin. There is sparse evidence supporting the use of nystatin prophylaxis beyond neonates; trials in specific pediatric patient groups are required.

The topical antifungal nystatin has not many indications. Prophylaxis of invasive candidiasis in very low birth weight neonates is one of them. In our study, we found that nystatin prophylaxis was used frequently beyond this specific neonatal group. Stronger evidence justifying its use is required.

The newly synthesized thiazole derivatives as potential antifungal compounds against Candida albicans.

Recently, the occurrence of candidiasis has increased dramatically, especially in immunocompromised patients. Additionally, their treatment is often ineffective due to the resistance of yeasts to antimycotics. Therefore, there is a need to search for new antifungals. A series of nine newly synthesized thiazole derivatives containing the cyclopropane system, showing promising activity against Candida spp., has been further investigated. We decided to verify their antifungal activity towards clinical Candida albicans isolated from the oral cavity of patients with hematological malignancies and investigate the mode of action on fungal cell, the effect of combination with the selected antimycotics, toxicity to erythrocytes, and lipophilicity. These studies were performed by the broth microdilution method, test with sorbitol and ergosterol, checkerboard technique, erythrocyte lysis assay, and reversed phase thin-layer chromatography, respectively. All derivatives showed very strong activity (similar and even higher than nystatin) against all C. albicans isolates with minimal inhibitory concentration (MIC) = 0.008-7.81 µg/mL Their mechanism of action may be related to action within the fungal cell wall structure and/or within the cell membrane. The interactions between the derivatives and the selected antimycotics (nystatin, chlorhexidine, and thymol) showed additive effect only in the case of combination some of them and thymol. The erythrocyte lysis assay confirmed the low cytotoxicity of these compounds as compared to nystatin. The high lipophilicity of the derivatives was related with their high antifungal activity. The present studies confirm that the studied thiazole derivatives containing the cyclopropane system appear to be a very promising group of compounds in treatment of infections caused by C. albicans. However, this requires further studies in vivo. KEY POINTS: • The newly thiazoles showed high antifungal activity and some of them - additive effect in combination with thymol. • Their mode of action may be related with the influence on the structure of the fungal cell wall and/or the cell membrane. • The low cytotoxicity against erythrocytes and high lipophilicity of these derivatives are their additional good properties.

Facile Synthesis of Bio-Antimicrobials with "Smart" Triiodides.

Multi-drug resistant pathogens are a rising danger for the future of mankind. Iodine (I2) is a centuries-old microbicide, but leads to skin discoloration, irritation, and uncontrolled iodine release. Plants rich in phytochemicals have a long history in basic health care. Aloe Vera Barbadensis Miller (AV) and Salvia officinalis L. (Sage) are effectively utilized against different ailments. Previously, we investigated the antimicrobial activities of smart triiodides and iodinated AV hybrids. In this work, we combined iodine with Sage extracts and pure AV gel with polyvinylpyrrolidone (PVP) as an encapsulating and stabilizing agent. Fourier transform infrared spectroscopy (FT-IR), Ultraviolet-visible spectroscopy (UV-Vis), Surface-Enhanced Raman Spectroscopy (SERS), microstructural analysis by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and X-Ray-Diffraction (XRD) analysis verified the composition of AV-PVP-Sage-I2. Antimicrobial properties were investigated by disc diffusion method against 10 reference microbial strains in comparison to gentamicin and nystatin. We impregnated surgical sutures with our biohybrid and tested their inhibitory effects. AV-PVP-Sage-I2 showed excellent to intermediate antimicrobial activity in discs and sutures. The iodine within the polymeric biomaterial AV-PVP-Sage-I2 and the synergistic action of the two plant extracts enhanced the microbial inhibition. Our compound has potential for use as an antifungal agent, disinfectant and coating material on sutures to prevent surgical site infections.

Rosmarinus officinalis L. (rosemary) extract has antibiofilm effect similar to the antifungal nystatin on Candida samples.

Candida spp. are naturally opportunistic and can promote infections. These yeasts can form biofilm, after penetration and adhesion to the biotic or abiotic surfaces. Preexisting diseases, treatments with drugs and radiation therapy, medical procedures, and parafunctional habits favor the installation of a fungal infection. Increased resistance to the available antifungals has become a concern. Therefore, alternative methods to control them have been evaluated, including the use of plant substances. In this study, the antibiofilm effect of R. officinalis L. extract was analyzed on C. albicans, C. dubliniensis, C. glabrata, C. krusei, and C. tropicalis. A phytochemical analysis of the extract was performed. Biofilms were formed for 48 h and exposed to the different concentrations of the extract (50, 100, and 200 mg/mL) for 5 min or 24 h. The effect of the plant extract was compared to the antifungal nystatin. Rosmarinus officinalis L. extract was constituted of phenols and flavonoids, highlighting the presence of chlorogenic acid derivatives in its composition. Biofilm reductions were observed after exposure to the plant extract for both periods. The plant extract provided a reduction similar to the antifungal. Thus, R. officinalis L. extract showed antibiofilm effect on Candida spp. comparable to the nystatin.

Histoplasmosis: informe de un caso / Histoplasmosis: a case report

Objetivo: Describir el diagnóstico y el tratamiento interdisciplinario de un caso clínico de histoplasmosis. Caso clínico: Un paciente masculino de 39 años, con antecedentes de consumo de drogas, alcohol, tabaquismo crónico y VIH+ sin adherencia al tratamiento, acudió al Servicio de Odontología por una interconsulta del Servicio de Clínica Médica para la evaluación de lesiones erosivas en paladar duro y blando, reborde alveolar anterior, dorso lingual y lesión tumoral en encía anterosuperior. El diagnóstico definitivo se obtuvo por medio del análisis de muestras de biopsia transbronquial, lavado broncoalveolar y biopsia de lesión en piel. El paciente recibió tratamiento sistémico con antimicóticos (anfotericina B e itraconazol según esquema) y tratamiento local con colutorio de clorhexidina al 0,12% y 100.000 UI de nistatina en suspensión. Al momento del alta, presentaba una considerable mejoría de su estado general y de las lesiones orales, con disminución de sintomatología dolorosa. Se indicó turno para control a los 7 días de forma ambulatoria, al cual el paciente no asistió. El abordaje interdisciplinario y el análisis de los diferentes aspectos socioeconómicos, culturales, ambientales y sistémicos del paciente facilitaron el diagnóstico temprano de la enfermedad (AU)

Aim: To describe the diagnosis and interdisciplinary treatment of a clinical case of histoplasmosis. Clinical case: 39-year-old male patient with a history of drug abuse, alcohol, and chronic smoking, HIV+ without treatment compliance, attends the dental department referred by the medical department for the diagnosis of erosive lesions in the hard and soft palate, anterior alveolar ridge, lingual dorsum and tumor lesion in the anterosuperior gingiva. The definitive diagnosis was obtained by the analysis of transbronchial biopsy, bronchoalveolar lavage and skin lesion biopsy. The patient received systemic treatment with antifungals (amphotericin b, itraconazole according to protocol), and local treatment with 0.12% chlorhexidine mouthwash and 100,000 IU nystatin suspension. At the time of medical discharge, the patient presented a considerable improvement in his general condition and of the oral lesions with a reduced pain. A 7 days recall was prescribed, however the patient failed to attend. The interdisciplinary approach to the patient and the analysis of the different socio-economic, cultural, environmental and systemic aspects of the patient facilitates the early diagnosis of the disease (AU)