Improving the Antimicrobial Potency of Berberine for Endodontic Canal Irrigation Using Polymeric Nanoparticles.

To address the challenges posed by biofilm presence and achieve a substantial reduction in bacterial load within root canals during endodontic treatment, various irrigants, including nanoparticle suspensions, have been recommended. Berberine (BBR), a natural alkaloid derived from various plants, has demonstrated potential applications in dentistry treatments due to its prominent antimicrobial, anti-inflammatory, and antioxidant properties. This study aimed to produce and characterize a novel polymeric nanoparticle of poly (lactic-co-glycolic acid) (PLGA) loaded with berberine and evaluate its antimicrobial activity against relevant endodontic pathogens, Enterococcus faecalis, and Candida albicans. Additionally, its cytocompatibility using gingival fibroblasts was assessed. The polymeric nanoparticle was prepared by the nanoprecipitation method. Physicochemical characterization revealed spheric nanoparticles around 140 nm with ca, -6 mV of surface charge, which was unaffected by the presence of BBR. The alkaloid was successfully incorporated at an encapsulation efficiency of 77% and the designed nanoparticles were stable upon 20 weeks of storage at 4 °C and 25 °C. Free BBR reduced planktonic growth at ≥125 µg/mL. Upon incorporation into PLGA nanoparticles, 20 µg/mL of [BBR]-loaded nanoparticles lead to a significant reduction, after 1 h of contact, of both planktonic bacteria and yeast. Sessile cells within biofilms were also considered. At 30 and 40 µg/mL, [BBR]-loaded PLGA nanoparticles reduced the viability of the sessile endodontic bacteria, upon 24 h of exposure. The cytotoxicity of BBR-loaded nanoparticles to oral fibroblasts was negligible. The novel berberine-loaded polymeric nanoparticles hold potential as a promising supplementary approach in the treatment of endodontic infections.

Impact of secreted glucanases upon the cell surface and fitness of Candida albicans during colonisation and infection.

Host recognition of the pathogen-associated molecular pattern (PAMP), ß-1,3-glucan, plays a major role in antifungal immunity. ß-1,3-glucan is an essential component of the inner cell wall of the opportunistic pathogen Candida albicans. Most ß-1,3-glucan is shielded by the outer cell wall layer of mannan fibrils, but some can become exposed at the cell surface. In response to host signals such as lactate, C. albicans shaves the exposed ß-1,3-glucan from its cell surface, thereby reducing the ability of innate immune cells to recognise and kill the fungus. We have used sets of barcoded xog1 and eng1 mutants to compare the impacts of the secreted ß-glucanases Xog1 and Eng1 upon C. albicans in vitro and in vivo. Flow cytometry of Fc-dectin-1-stained strains revealed that Eng1 plays the greater role in lactate-induced ß-1,3-glucan masking. Transmission electron microscopy and stress assays showed that neither Eng1 nor Xog1 are essential for cell wall maintenance, but the inactivation of either enzyme compromised fungal adhesion to gut and vaginal epithelial cells. Competitive barcode sequencing suggested that neither Eng1 nor Xog1 strongly influence C. albicans fitness during systemic infection or vaginal colonisation in mice. However, the deletion of XOG1 enhanced C. albicans fitness during gut colonisation. We conclude that both Eng1 and Xog1 exert subtle effects on the C. albicans cell surface that influence fungal adhesion to host cells and that affect fungal colonisation in certain host niches.

Probing gene function in Candida albicans wild-type strains by Cas9-facilitated one-step integration of two dominant selection markers: a systematic analysis of recombination events at the target locus.

The adaptation of gene deletion methods based on the CRISPR-Cas9 system has facilitated the genetic manipulation of the pathogenic yeast Candida albicans, because homozygous mutants of this diploid fungus can now be generated in a single step, allowing the rapid screening of candidate genes for their involvement in a phenotype of interest. However, the Cas9-mediated double-strand breaks at the target site may result in an undesired loss of heterozygosity (LOH) on the affected chromosome and cause phenotypic alterations that are not related to the function of the investigated gene. In our present study, we harnessed Cas9-facilitated gene deletion to probe a set of genes that are constitutively overexpressed in strains containing hyperactive forms of the transcription factor Mrr1 for a possible contribution to the fluconazole resistance of such strains. To this aim, we used gene deletion cassettes containing two different dominant selection markers, caSAT1 and HygB, which confer resistance to nourseothricin and hygromycin, respectively, for simultaneous genomic integration in a single step, hypothesizing that this would minimize undesired LOH events at the target locus. We found that selection for resistance to both nourseothricin and hygromycin strongly increased the proportion of homozygous deletion mutants among the transformants compared with selection on media containing only one of the antibiotics, but it did not avoid undesired LOH events. Our results demonstrate that LOH on the target chromosome is a significant problem when using Cas9 for the generation of C. albicans gene deletion mutants, which demands a thorough examination of recombination events at the target site. IMPORTANCE: Candida albicans is one of the medically most important fungi and a model organism to study fungal pathogenicity. Investigating gene function in this diploid yeast has been facilitated by the adaptation of gene deletion methods based on the bacterial CRISPR-Cas9 system, because they enable the generation of homozygous mutants in a single step. We found that, in addition to increasing the efficiency of gene replacement by selection markers, the Cas9-mediated double-strand breaks also result in frequent loss of heterozygosity on the same chromosome, even when two different selection markers were independently integrated into the two alleles of the target gene. Since loss of heterozygosity for other genes can result in phenotypic alterations that are not caused by the absence of the target gene, these findings show that it is important to thoroughly analyze recombination events at the target locus when using Cas9 to generate gene deletion mutants in C. albicans.

Absence of farnesol salvage in Candida albicans and probably in other fungi.

Farnesol salvage, a two-step pathway converting farnesol to farnesyl pyrophosphate (FPP), occurs in bacteria, plants, and animals. This paper investigates the presence of this pathway in fungi. Through bioinformatics, biochemistry, and physiological analyses, we demonstrate its absence in the yeasts Saccharomyces cerevisiae and Candida albicans, suggesting a likely absence across fungi. We screened 1,053 fungal genomes, including 34 from C. albicans, for potential homologs to four genes (Arabidopsis thaliana AtFOLK, AtVTE5, AtVTE6, and Plasmodium falciparum PfPOLK) known to accomplish farnesol/prenol salvage in other organisms. Additionally, we showed that 3H-farnesol was not converted to FPP or any other phosphorylated prenol, and exogenous farnesol was not metabolized within 90 minutes at any phase of growth and did not rescue cells from the toxic effects of atorvastatin, but it did elevate the levels of intracellular farnesol (Fi). All these experiments were conducted with C. albicans. In sum, we found no evidence for farnesol salvage in fungi. IMPORTANCE: The absence of farnesol salvage constitutes a major difference in the metabolic capabilities of fungi. In terms of fungal physiology, the lack of farnesol salvage pathways relates to how farnesol acts as a quorum-sensing molecule in Candida albicans and why farnesol should be investigated for use in combination with other known antifungal antibiotics. Its absence is essential for a model (K. W. Nickerson et al., Microbiol Mol Biol Rev 88:e00081-22, 2024), wherein protein farnesylation, protein chaperones, and the unfolded protein response are combined under the unifying umbrella of a cell's intracellular farnesol (Fi). In terms of human health, farnesol should have at least two different modes of action depending on whether those cells have farnesol salvage. Because animals have farnesol salvage, we can now see the importance of dietary prenols as well as the potential importance of farnesol in treating neurodegenerative diseases such as Parkinson's disease, Alzheimer's disease, and multiple sclerosis.

Efficacy of denture cleansers on Candida albicans adhesion and their effects on the properties of conventional, milled CAD/CAM, and 3D-printed denture bases.

OBJECTIVES: Evaluate the efficacy of denture cleaners on the adhesion of Candida albicans and their effects on the surface, optical, and mechanical properties of resins for conventional, milled, and 3D-printed denture bases. MATERIALS AND METHODS: A total of 240 resin samples were made, 120 for testing Candida albicans adhesion, optical stabilities (ΔE00), roughness (Ra), hydrophilicity (°), surface free energy (Owens-Wendt) and 120 samples for testing Candida albicans adhesion, surface microhardness (Knoop), flexural strength and modulus of elasticity in a three-point test, in which they were divided into 3 groups of denture resin (n = 40) and subdivided into 5 cleaners of dentures (n = 8). Data were evaluated by two-way ANOVA and Tukey's test for multiple comparisons (α = 0.05). RESULTS: Denture cleaners with an alkaline solution and dilute acid composition were those that showed the greatest effectiveness in reducing Candida albicans (P < 0.001), however 1% NaOCl significantly affected the properties of the resins (P < 0.05). Denture 3D-printed showed that the surface microhardness was significantly lower for all cleansers (P < 0.05). CONCLUSIONS: Listerine demonstrated superior efficacy in reducing Candida albicans with minimal effect on denture properties, whereas 1% NaOCl had a significant negative impact on the properties. The mechanical properties were significantly lower in 3D-printed resin than in other resins for all denture cleansers. CLINICAL RELEVANCE: Denture base materials are being sold to adapt to the CAD/CAM system, increasing the number of users of dentures manufactured with this system. Despite this, there is little investigation into denture cleaners regarding the adhesion capacity of microorganisms and the optical, surface and mechanical properties of dentures, thus requiring further investigation.

Candida albicans-A systematic review to inform the World Health Organization Fungal Priority Pathogens List.

Candida albicans is a common fungal pathogen and amongst the leading causes of invasive candidiasis globally. This systematic review examines the characteristics and global impact of invasive infections caused by C. albicans. We searched on PubMed and Web of Science for studies reporting on criteria such as mortality, morbidity, drug resistance, preventability, yearly incidence, and distribution/emergence during the period from 2016 to 2021. Our findings indicate that C. albicans is the most common Candida species causing invasive disease and that standard infection control measures are the primary means of prevention. However, we found high rates of mortality associated with infections caused by C. albicans. Furthermore, there is a lack of data on complications and sequelae. Resistance to commonly used antifungals remains rare. Although, whilst generally susceptible to azoles, we found some evidence of increasing resistance, particularly in middle-income settings-notably, data from low-income settings were limited. Candida albicans remains susceptible to echinocandins, amphotericin B, and flucytosine. We observed evidence of a decreasing proportion of infections caused by C. albicans relative to other Candida species, although detailed epidemiological studies are needed to confirm this trend. More robust data on attributable mortality, complications, and sequelae are needed to understand the full extent of the impact of invasive C. albicans infections.

Homo and Hetero-Branched Lipopeptide Dendrimers: Synthesis and Antimicrobial Activity.

Di-branched and tetra-branched versions of a previously reported analogue of the lipopeptide battacin were successfully synthesised using thiol-maleimide click and 1, 2, 3-triazole click chemistry. Antimicrobial studies against drug resistant clinical isolates of Escherichia coli (ESBL E. coli Ctx-M14), Pseudomonas aeruginosa (P. aeruginosa Q502), and Methicillin resistant Staphylococcus aureus (MRSA ATCC 33593), as well as clinically isolated Acinetobacter baumannii (A. baumannii ATCC 19606), and P. aeruginosa (ATCC 27853), revealed that the dendrimeric peptides have antimicrobial activity in the low micromolar range (0.5 -- 4 µM) which was 10 times more potent than the monomer peptides. Under high salt concentrations (150 mM NaCl, 2 mM MgCl2, and 2.5 mM CaCl2) the di-branched lipopeptides retained their antimicrobial activity while the monomer peptides were not active (>100 µM). The di-branched triazole click lipopeptide, Peptide 12, was membrane lytic, showed faster killing kinetics, and exhibited antibiofilm activity against A. baumannii and MRSA and eradicated > 85 % preformed biofilms at low micromolar concentrations. The di-branched analogues were > 30-fold potent than the monomers against Candida albicans. Peptide 12 was not haemolytic (HC10 = 932.12 µM) and showed up to 40-fold higher selectivity against bacteria and fungi than the monomer peptide. Peptide 12 exhibited strong proteolytic stability (>80 % not degraded) in rat serum over 24 h whereas > 95 % of the thiol-maleimide analogue (Peptide 10) was degraded. The tetra-branched peptides showed comparable antibacterial potency to the di-branched analogues. These findings indicate that dual branching using triazole click chemistry is a promising strategy to improve the antimicrobial activity and proteolytic stability of battacin based lipopeptides. The information gathered can be used to build effective antimicrobial dendrimeric peptides as new peptide antibiotics.

In Silico and In Vitro Evaluation of the Antifungal Activity of a New Chromone Derivative against Candida spp.

Candida species are frequently implicated in the development of both superficial and invasive fungal infections, which can impact vital organs. In the quest for novel strategies to combat fungal infections, there has been growing interest in exploring synthetic and semi-synthetic products, particularly chromone derivatives, renowned for their antimicrobial properties. In the analysis of the antifungal activity of the compound (E)-benzylidene-chroman-4-one against Candida, in silico and laboratory tests were performed to predict possible mechanisms of action pathways, and in vitro tests were performed to determine antifungal activity (MIC and MFC), to verify potential modes of action on the fungal cell membrane and wall, and to assess cytotoxicity in human keratinocytes. The tested compound exhibited predicted affinity for all fungal targets, with the highest predicted affinity observed for thymidylate synthase (-102.589 kJ/mol). MIC and CFM values ranged from 264.52 µM (62.5 µg/mL) to 4232.44 µM (1000 µg/mL). The antifungal effect likely occurs due to the action of the compound on the plasma membrane. Therefore, (E)-benzylidene-chroman-4-one showed fungicidal-like activity against Candida spp., possibly targeting the plasma membrane.

DNA Damage Checkpoints Govern Global Gene Transcription and Exhibit Species-Specific Regulation on HOF1 in Candida albicans.

DNA damage checkpoints are essential for coordinating cell cycle arrest and gene transcription during DNA damage response. Exploring the targets of checkpoint kinases in Saccharomyces cerevisiae and other fungi has expanded our comprehension of the downstream pathways involved in DNA damage response. While the function of checkpoint kinases, specifically Rad53, is well documented in the fungal pathogen Candida albicans, their targets remain poorly understood. In this study, we explored the impact of deleting RAD53 on the global transcription profiles and observed alterations in genes associated with ribosome biogenesis, DNA replication, and cell cycle. However, the deletion of RAD53 only affected a limited number of known DNA damage-responsive genes, including MRV6 and HMX1. Unlike S. cerevisiae, the downregulation of HOF1 transcription in C. albicans under the influence of Methyl Methanesulfonate (MMS) did not depend on Dun1 but still relied on Rad53 and Rad9. In addition, the transcription factor Mcm1 was identified as a regulator of HOF1 transcription, with evidence of dynamic binding to its promoter region; however, this dynamic binding was interrupted following the deletion of RAD53. Furthermore, Rad53 was observed to directly interact with the promoter region of HOF1, thus suggesting a potential role in governing its transcription. Overall, checkpoints regulate global gene transcription in C. albicans and show species-specific regulation on HOF1; these discoveries improve our understanding of the signaling pathway related to checkpoints in this pathogen.

Mitochondrial Protease Oct1p Regulates Mitochondrial Homeostasis and Influences Pathogenicity through Affecting Hyphal Growth and Biofilm Formation Activities in Candida albicans.

Mitochondria, as the core metabolic organelles, play a crucial role in aerobic respiration/biosynthesis in fungi. Numerous studies have demonstrated a close relationship between mitochondria and Candida albicans virulence and drug resistance. Here, we report an octapeptide-aminopeptidase located in the mitochondrial matrix named Oct1p. Its homolog in the model fungus Saccharomyces cerevisiae is one of the key proteins in maintaining mitochondrial respiration and protein stability. In this study, we utilized evolutionary tree analysis, gene knockout experiments, mitochondrial function detection, and other methods to demonstrate the impact of Oct1p on the mitochondrial function of C. albicans. Furthermore, through transcriptome analysis, real-time quantitative PCR, and morphological observation, we discovered that the absence of Oct1p results in functional abnormalities in C. albicans, affecting hyphal growth, cell adhesion, and biofilm formation. Finally, the in vivo results of the infection of Galleria mellonella larvae and vulvovaginal candidiasis in mice indicate that the loss of Oct1p led to the decreased virulence of C. albicans. In conclusion, this study provides a solid theoretical foundation for treating Candida diseases, developing new targeted drugs, and serves as a valuable reference for investigating the connection between mitochondria and virulence in other pathogenic fungi.

Pitavastatin Calcium Confers Fungicidal Properties to Fluconazole by Inhibiting Ubiquinone Biosynthesis and Generating Reactive Oxygen Species.

Fluconazole (FLC) is extensively employed for the prophylaxis and treatment of invasive fungal infections (IFIs). However, the fungistatic nature of FLC renders pathogenic fungi capable of developing tolerance towards it. Consequently, converting FLC into a fungicidal agent using adjuvants assumes significance to circumvent FLC resistance and the perpetuation of fungal infections. This drug repurposing study has successfully identified pitavastatin calcium (PIT) as a promising adjuvant for enhancing the fungicidal activity of FLC from a comprehensive library of 2372 FDA-approved drugs. PIT could render FLC fungicidal even at concentrations as low as 1 µM. The median lethal dose (LD50) of PIT was determined to be 103.6 mg/kg. We have discovered that PIT achieves its synergistic effect by inhibiting the activity of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, thereby impeding ubiquinone biosynthesis, inducing reactive oxygen species (ROS) generation, triggering apoptosis, and disrupting Golgi function. We employed a Candida albicans strain that demonstrated a notable tolerance to FLC to infect mice and found that PIT effectively augmented the antifungal efficacy of FLC against IFIs. This study is an illustrative example of how FDA-approved drugs can effectively eliminate fungal tolerance to FLC.

Globospiramine Exhibits Inhibitory and Fungicidal Effects against Candida albicans via Apoptotic Mechanisms.

Candidiasis is considered an emerging public health concern because of the occurrence of drug-resistant Candida strains and the lack of an available structurally diverse antifungal drug armamentarium. The indole alkaloid globospiramine from the anticandidal Philippine medicinal plant Voacanga globosa exhibits a variety of biological activities; however, its antifungal properties remain to be explored. In this study, we report the in vitro anticandidal activities of globospiramine against two clinically relevant Candida species (C. albicans and C. tropicalis) and the exploration of its possible target proteins using in silico methods. Thus, the colony-forming unit (CFU) viability assay revealed time- and concentration-dependent anticandidal effects of the alkaloid along with a decrease in the number of viable CFUs by almost 50% at 60 min after treatment. The results of the MIC and MFC assays indicated inhibitory and fungicidal effects of globospiramine against C. albicans (MIC = 8 µg/mL; MFC = 8 µg/mL) and potential fungistatic effects against C. tropicalis at lower concentrations (MIC = 4 µg/mL; MFC > 64 µg/mL). The FAM-FLICA poly-caspase assay showed metacaspase activation in C. albicans cells at concentrations of 16 and 8 µg/mL, which agreed well with the MIC and MFC values. Molecular docking and molecular dynamics simulation experiments suggested globospiramine to bind strongly with 1,3-ß-glucan synthase and Als3 adhesin-enzymes indirectly involved in apoptosis-driven candidal inhibition.

Antifungal Activity, Synergism with Fluconazole or Amphotericin B and Potential Mechanism of Direct Current against Candida albicans Biofilms and Persisters.

Candida albicans, as a notorious fungal pathogen, is associated with high morbidity and mortality worldwide due to its ability to form biofilms and persisters that can withstand currently available antifungals. Direct current (DC) has demonstrated a promising antimicrobial effect and synergistic effect with antimicrobials against various infections. Here, we first found DC exerted a killing effect on C. albicans planktonic and biofilm cells. Moreover, DC showed a synergistic effect with fluconazole (FLC) and amphotericin B (AMB). Notably, near-to-complete eradication of AMB-tolerant C. albicans biofilm persisters was achieved upon DC treatment. Next, the mechanism of action of DC was explored through mapping the genes and proteomic profiles of DC-treated C. albicans. The multi-omics analysis, quantitative real-time PCR and assay of reactive oxygen species (ROS) demonstrated DC exerted an antifungal effect on C. albicans by increasing cellular oxidative stress. As revealed by multiple analyses (e.g., protein assay based on absorbance at 280 nm and rhodamine 6G assay), DC was able to enhance membrane permeability, inhibit drug efflux and increase cellular FLC/AMB concentration of C. albicans, thereby mediating its synergism with the antifungals. Furthermore, DC inhibited superoxide dismutase 2 (SOD2) expression and manganese-containing SOD (Mn SOD) activity, leading to ROS production and enhanced killing of C. albicans biofilm persisters. The current findings demonstrate that the adjunctive use of DC in combination with antifungals is a promising strategy for effective control of C. albicans infections and management of antifungal resistance/tolerance in Candida biofilms.

Padina Minor Extract Confers Resistance against Candida Albicans Infection: Evaluation in a Zebrafish Model.

Padina minor is a seaweed rich in polysaccharides often used in food, feed, fertilizers, and antibacterial drugs. This study is the first to evaluate the effect of feeding zebrafish with Padina minor extract on preventing and treating C. albicans infections. This study evaluated the growth, survival, and disease resistance effects of P. minor extract on zebrafish. The fish were divided into four groups: three groups treated with 1%, 5%, or 10% P. minor extract and one untreated group (c, control). Subsequently, we analyzed how the extract affected the immune function of zebrafish infected with C. albicans. Based on the lethal concentration (LC50) calculated in the first stage, 1% was used as the effective therapeutic concentration. The results showed that the growth rate of the 1% feed group was the best, and no significant difference in survival rates between the four groups was observed. Feeding with 1% P. minor extract downregulated the expression of key inflammatory genes like tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-1ß), and IL-10, effectively preventing and treating C. albicans infections in zebrafish. This study is a preliminary evaluation of the therapeutic efficacy of P. minor extracts against C. albicans.

Candida albicans and Candida glabrata: global priority pathogens.

SUMMARYA significant increase in the incidence of Candida-mediated infections has been observed in the last decade, mainly due to rising numbers of susceptible individuals. Recently, the World Health Organization published its first fungal pathogen priority list, with Candida species listed in medium, high, and critical priority categories. This review is a synthesis of information and recent advances in our understanding of two of these species-Candida albicans and Candida glabrata. Of these, C. albicans is the most common cause of candidemia around the world and is categorized as a critical priority pathogen. C. glabrata is considered a high-priority pathogen and has become an increasingly important cause of candidemia in recent years. It is now the second most common causative agent of candidemia in many geographical regions. Despite their differences and phylogenetic divergence, they are successful as pathogens and commensals of humans. Both species can cause a broad variety of infections, ranging from superficial to potentially lethal systemic infections. While they share similarities in certain infection strategies, including tissue adhesion and invasion, they differ significantly in key aspects of their biology, interaction with immune cells, host damage strategies, and metabolic adaptations. Here we provide insights on key aspects of their biology, epidemiology, commensal and pathogenic lifestyles, interactions with the immune system, and antifungal resistance.

H3K56 acetylation affects Candida albicans morphology and secreted soluble factors interacting with the host.

In recent years, epigenetics has been revealed as a mechanism able to modulate the expression of virulence traits in diverse pathogens, including Candida albicans. Indeed, epigenetic regulation can sense environmental changes, leading to the rapid and reversible modulation of gene expression with consequent adaptation to novel environments. How epigenetic changes can impact expression and signalling output, including events associated with mechanisms of morphological transition and virulence, is still poorly studied. Here, using nicotinamide as a sirtuin inhibitor, we explored how the accumulation of the H3K56 acetylation, the most prominent histone acetylation in C. albicans, might affect its interaction with the host. Our experiments demonstrate that H3K56 acetylation profoundly affects the production and/or secretion of soluble factors compromising actin remodelling and cytokine production. ChIP- and RNA-seq analyses highlighted a direct impact of H3K56 acetylation on genes related to phenotypic switching, biofilm formation and cell aggregation. Direct and indirect regulation also involves genes related to cell wall protein biosynthesis, ß-glucan and mannan exposure, and hydrolytic secreted enzymes, supporting the hypothesis that the fluctuations of H3K56 acetylation in C. albicans might impair the macrophage response to the yeast and thus promote the host-immune escaping.

Association between Salivary Hormones, Dental Caries, and Cariogenic Microorganisms during Pregnancy.

Objective: This study aimed to identify the salivary levels of six hormones (progesterone, estradiol, testosterone, cortisol, thyroxine T3, and triiodothyronine T4) in pregnant women, and to assess the association between salivary hormones, dental caries, and cariogenic microorganisms. Methods: This cross-sectional study included 181 low-income US pregnant women who were in their third trimester. Demographic details, oral hygiene practices, and medical backgrounds were obtained via questionnaires and medical records. Calibrated dentists obtained data on plaque index and caries status through comprehensive oral examinations. Unstimulated saliva was collected 2 h before eating and brushing. Salivary hormones were measured with a multiplex assay. Oral Streptococcus mutans (S. mutans) and Candida albicans (C. albicans) were quantified via colony-forming unit (CFU) counts. A latent model was used to generate clusters of pregnant women based on salivary hormone levels, followed by post-clustering analysis. Factors associated with salivary cariogenic microorganisms were further evaluated via multiple regression analyses. Results: Estradiol, progesterone, testosterone, cortisol, T3, and T4 in saliva were detectable at rates of 92%, 97%, 77%, 99%, 71%, and 50%, respectively. Three distinct participant clusters (high, intermediate, and low) were identified based on salivary hormone levels. Intermediate-level and high-level clusters had increased numbers of decayed teeth, decayed surfaces, ICDAS scores, and salivary S. mutans and C. albicans, compared to the low-level cluster (p < 0.05). Covariate analysis demonstrated that the high-level cluster was positively associated with salivary carriage of S. mutans (CFU/mL) (p < 0.05). Participants with higher levels of progesterone, estradiol, testosterone, and cortisol were associated with a high carriage status of S. mutans in saliva (>105 CFU/mL) (p < 0.05). Conclusions: This study demonstrated the feasibility of detecting salivary hormones during pregnancy and revealed the positive association between salivary steroid hormones and cariogenic pathogens.

Elucidating the structural basis for the enhanced antifungal activity of amide derivative against Candida albicans: a comprehensive computational investigation.

The continuous search for more effective options against well-known pathogens such as Candida albicans remains the rationale for the search for novel lead compounds from various sources. This study aims to investigate the chemical structure, chemical properties, of 5-(2-((5-(((1S,3R) -3-(5-acetamido-1,3,4-thiadiazolidin-2-yl) cyclopentyl) methyl)-1,3,4-thiadiazolidin-2-yl)amino)-2-oxoethyl)-2-methyl-2,3-dihydro-1H-pyrazol-3-ide designated ATCTP using DFT method ωB97XD/-311 + + g(2d, 2p) and the biological potential of compound ATCTP against Candida albicans using molecular docking and ADMET studies. Geometry optimization was carried out in DMSO, ethanol. gas and water revealing minute discrepancies in bond length and wider differences in bond angles. Frontier molecular orbital investigations reveal HOMO-LUMO energy gap magnitude in decreasing order of ATCTP_Gas > ATCTP_Water > ATCTP_ethanol > ATCTP_DMSO inferring that water influences chemical stability of the compound the most compared to ethanol and DMSO. Density of state investigations have revealed electron density contributions at corresponding energy peaks. In silico pharmacokinetic predicts ATCTP not to be cytotoxic, hepatotoxic, immunotoxic or mutagenic but probable mutagen. Molecular docking investigation of ATCTP against aspartic proteinase of Candida albicans (ID: 2QZX) in comparison with standard drug Fluconazole. Compound ATCTP had higher binding affinity (- 8.1 kcal/mol) compared to that of the standard drug fluconazole (- 5.6 kcal/mol) which records 4 conventional hydrogen interactions compared to 2 formed in the interaction of ATCTP + 2QZX. ATCTP also reports binding affinity of - 7.2 kcal/mol which reportedly surpassed that of 2QZX interaction with fluconazole (- 5.7 kcal/mol). ATCTP binds with lanosterol14-α-demethylase (5v5z) with binding affinity of - 9.7 kcal/mol binding to active site amino acid residues of the protein compared to fluconazole + 5v5z (- 8.0 kcal/mol). ATCTP is therefore recommended to be a lead compound for the possible design of a new and more effective anti-candida therapeutic compound.

Mechanism and bioinformatics analysis of the effect of berberine-enhanced fluconazole against drug-resistant Candida albicans.

Biofilms produced by Candida albicans present a challenge in treatment with antifungal drug. Enhancing the sensitivity to fluconazole (FLC) is a reasonable method for treating FLC-resistant species. Moreover, several lines of evidence have demonstrated that berberine (BBR) can have antimicrobial effects. The aim of this study was to clarify the underlying mechanism of these effects. We conducted a comparative study of the inhibition of FLC-resistant strain growth by FLC treatment alone, BBR treatment alone, and the synergistic effect of combined FLC and BBR treatment. Twenty-four isolated strains showed distinct biofilm formation capabilities. The antifungal effect of combined FLC and BBR treatment in terms of the growth and biofilm formation of Candida albicans species was determined via checkerboard, time-kill, and fluorescence microscopy assays. The synergistic effect of BBR and FLC downregulated the expression of the efflux pump genes CDR1 and MDR, the hyphal gene HWP1, and the adhesion gene ALS3; however, the gene expression of the transcriptional repressor TUP1 was upregulated following treatment with this drug combination. Furthermore, the addition of BBR led to a marked reduction in cell surface hydrophobicity. To identify resistance-related genes and virulence factors through genome-wide sequencing analysis, we investigated the inhibition of related resistance gene expression by the combination of BBR and FLC, as well as the associated signaling pathways and metabolic pathways. The KEGG metabolic map showed that the metabolic genes in this strain are mainly involved in amino acid and carbon metabolism. The metabolic pathway map showed that several ergosterol (ERG) genes were involved in the synthesis of cell membrane sterols, which may be related to drug resistance. In this study, BBR + FLC combination treatment upregulated the expression of the ERG1, ERG3, ERG4, ERG5, ERG24, and ERG25 genes and downregulated the expression of the ERG6 and ERG9 genes compared with fluconazole treatment alone (p < 0.05).