Flocculation phenomenon of Candida albicans by lysozyme

Young colonies of Sabouraud's glucose agar room temperature culture ofCandida species from human isolation were suspended in distilled water. The suspension was mixed with a solution of lysozyme and incubated in a 37° C water bath. Within 3–5 hours, various species ofCandida cells showed flocculation to varying degrees which occurred at varying periods of onset. Among sevenCandida species,Candida albicans andCandida stellatoidea showed the strongest flocculation, earliest onset and most solution clarity than did any other species.Candida stellatoidea was indistinguishable fromCandida albicans in its degree of flocculation, and in the clarity of solution.Candida species may be arranged in the following order according to their decreasing positivity in flocculation:

1. Candida albicans
2. Candida stellatoidea
3. Candida tropicalis
4. Candida krusei
5. Candida pseudotropicalis
6. Candida parapsilosis
7. Candida guilliermondii
8. Saccharomyces species may be placed afterCandida guilliermondii.

It seems possible to separate theCandida species into 3 groups by the rate of flocculation, and clarity of solution. Group I.Candida albicans andCandida stellatoidea. Group II.Candida tropicalis, C. krusei andCandida pseudotropicalis. Group III.Candida parapsilosis andCandida guilliermondii. Saccharomyces specimens (S. cerevisiae and others) were placed after group III.     

Antimicrobial agents

The effect of 2-aminotridecane and its addition salts with different substituted phenols on growth ofCandida albicans was studied and the fungistatic and fungicidal activity of these substances was compared. The addition 2-aminotridecane salts with 2,4,5-trichlorophenol and with 2-chloro-4-introphenol were found to be the most active in all three tests. The activity of the other addition salts tested differed little from that of actual 2-aminotridecane.     

Nosocomial Bloodstream Candida Infections in a Tertiary-Care Hospital in South Brazil: A 4-Year Survey

The aims of this study were to evaluate the epidemiology of nosocomial candidemia in a tertiary hospital in South Brazil and the in vitro antifungal susceptibility of isolates. Blood strains from 108 patients were identified by PCR-based method. Some 30.5 % of candidemia were caused by Candida tropicalis, 28.7 % were due to Candida albicans, 24.1 % with Candida parapsilosis sensu stricto, 8.3 % with Candida glabrata sensu lato, 1.8 % involved Candida krusei and 6.6 % with other species. Candidemia was more common in intensive care unit settings (66 %). In vitro susceptibility to antifungal drugs was determined by a microdilution method; and new species-specific clinical breakpoints for fluconazole and voriconazole were applied. Overall susceptibility rates were 100 % for itraconazole, 91 % for fluconazole, 98 % for voriconazole and 99 % for amphotericin B. Fluconazole resistance was mostly among C. parapsilosis sensu stricto isolates (26.9 %). Most of the findings reported here agreed with epidemiological features common to other tertiary hospitals in Brazil; but also revealed some peculiarities, such as a high frequency of C. tropicalis associated with candidemia. Besides, high rate of fluconazole resistance among C. parapsilosis stricto sensu isolates was obtained when applying the new species-specific clinical breakpoints.     

Killer behavior within the Candida parapsilosis complex

A group of 29 isolates of Candida parapsilosis sensu stricto, 29 of Candida orthopsilosis, and 4 of Candida metapsilosis were assayed for the presence of killer activity using Saccharomyces cerevisiae ATCC 26609 as a sensitive strain. All C. metapsilosis isolates showed killer activity at 25 °C while strains of C. parapsilosis sensu stricto or C. orthopsilosis did not exhibit this activity. Sensitivity to killer toxins was evaluated using a set of previously reported killer strains of clinical origin. Only 11 isolates of the C. parapsilosis complex were inhibited by at least one killer isolate without resulting in any clear pattern, except for C. parapsilosis sensu stricto ATCC 22019, which was inhibited by every killer strain with the exception of C. parapsilosis and Candida utilis. The lack of sensitivity to killer activity among isolates of the genus Candida suggests that their toxins belong to the same killer type. Differentiation of species within the C. parapsilosis complex using the killer system may be feasible if a more taxonomically diverse panel of killer strains is employed.     

Interaction between the antimicrobial peptide Aurein 1.2 dimer and mannans

We have previously described the structure and the ability of a dimeric analog of the antimicrobial peptide Aurein 1.2 to aggregate Candida albicans. In this study, circular dichroism and fluorescence spectroscopy data showed that this aggregation is related to the interaction between the peptide and mannans, the main component of yeast cell wall. In this context, we propose a model in which dimers interact with the polysaccharide leading to cells aggregation.     

Candidacidal mechanism of the arenicin-3-derived peptide NZ17074 from Arenicola marina

The candidacidal mechanisms of NZ17074, which is a variant of arenicin-3 from Arenicola marina, against human pathogenic fungus Candida albicans are reported in this work. The minimum inhibitory concentration (MIC) of NZ17074 toward C. albicans was 4 μg/ml, and this peptide exerted marked candidacidal activity in an energy-dependent and salt-sensitive manner. The flow cytometric analysis using propidium iodide (PI) showed that the plasma membrane of cells treated with NZ17074 was perturbed and that the cells were arrested in the G2/M phase. The dihydrorhodamine-123 (DHR-123) staining showed that the reactive oxygen species (ROS) production of C. albicans increased after exposure to NZ17074. Typical cellular disruption events, such as mitochondrial degradation, nuclear fragmentation, nuclear membrane disruption, and chromatin condensation, were further revealed through rhodamine 123 (RH123) staining, 4′,6-diamidino-2-phenylindole (DAPI) staining, and transmission electron microscopy. In addition, the intracellular localization of this peptide was concentration dependent: it was located in the membrane at low concentrations (4 to 8 μg/ml) and penetrated into the cytoplasm at high concentrations (16 to 32 μg/ml). Our results suggested that NZ17074 exerts its candidacidal effects by disrupting the cell membrane, inducing apoptosis, and interrupting the cell cycle. These findings showed the potential of NZ17074 as a new candidacidal peptide, in addition to its antibacterial activities.     

Genetic Diversity Among Candida albicans Isolates Associated with Vertical Transmission in Preterm Triplets

We report a case of congenital candidiasis in triplets, in the context of premature labor at 25 weeks gestation, without symptomatic vaginitis or chorioamnionitis. All three infants died as a result of prematurity, aggravated by systemic candidiasis. Multi-locus sequence typing confirmed vertical transmission of Candida albicans from the mother to the triplets and revealed a slight diversity among the strains isolated from the neonates.     

Counterimmunoelectrophoresis as a routine mycoserological procedure

Counterimmunoelectrophoresis (CIE) has been compared in a diagnostic laboratory with agar gel double diffusion (DD) as a routine procedure for detection of antibodies to pathogenic and allergenic fungi and actinomycetes. It was shown to be of particular value in detecting antibodies to Aspergillus fumigatus. Thus 72 of 106 sera in which precipitins were detected were positive by CIE alone. Some sera were positive only by CIE to antigens prepared from Histoplasma capsulatum, Allescheria boydii, Candida albicans and C. parapsilosis.     

Stability and zymogenic nature of chitin synthase fromCandida albicans

Chitin synthase preparations from both yeast and mycelialCandida albicans were chiefly in a zymogenic form, activatable by trypsin treatment. This was especially marked for preparations that had been solubilized with digitonin treatment. Endogenous activation of chitin synthase zymogen was observed over many days in preparations stored in glycerol (33%, wt/v) at?12°C and over many hours in preparations stored at 30°C. Gel chromatography of enzyme preparations suggested that zymogen was preferentially retarded on the columm matrices in comparison with active enzyme.     

Candidaemia Observed at a University Hospital in Milan (Northern Italy) and Review of Published Studies from 2010 to 2014

Background

Candida species represent the fourth leading cause of nosocomial bloodstream infections (BSI) worldwide. However, candidaemia rates and species involved vary geographically.

Objectives

To evaluate the epidemiological pattern, risk factors for mortality and antifungal therapy of Candida BSI over a 5-year period (2008–2012) in a university hospital in northern Italy together with a review of the recent literature concerning candidaemia.

Methods

A retrospective cohort study cross-linked with microbiology database was performed.

Results

A total of 89 Candida BSI were identified in 42 males (47 %) and 47 females (52.8 %). The median age was 69 years (interquartile range 55–78) with 61.8 % of patients being older than 65 years. Considering all hospitalized patients, the overall incidence rate of candidaemia increased significantly from 2008 to 2012 (from 0.4 to 1.68 episodes per 10,000 patient/days) (p = 0.0001) with a mean linear increase in 5 new cases per year. Candida albicans was the predominant species isolated (64 %) followed by C. glabrata (19.1 %). The latter species was observed with significantly higher frequency in Internal Medicine and Intensive Care Units (ICU). In-hospital crude mortality was 41.6 %.

Conclusions

Candidaemia is an increasing BSI in our university hospital, in accordance with that observed in northern Italy, and it is still associated with high in-hospital crude mortality.     

Aerobic endospore-forming bacteria isolated from Antarctic soils as producers of bioactive compounds of industrial interest

Spore-forming bacteria are known to produce various enzymes and bioproducts valuable to different industries and to bear the harsh conditions found in the Antarctic environment. However, aerobic or facultative spore-forming bacterial communities found in maritime Antarctic soils yet remain poorly studied. In this study, 80 spore-forming and cold-adapted bacterial strains were isolated from nine different soil samples of King George Island, in maritime Antarctica, and further clustered into amplified ribosomal DNA restriction analysis groups within each soil. Representative strains were then identified as belonging to Bacillus, Rummeliibacillus, Paenibacillus and Sporosarcina by 16S rRNA gene sequencing. The ability to produce extracellular enzymes, antimicrobial substances and biosurfactants was determined in all isolates. The enzymatic activities most frequently found among the isolates were as follows: esterase (45 %), caseinase (30 %), amylase (16.2 %) and gelatinase (15 %). Biosurfactant production was detected in 25 % of the isolates. The growth inhibition of methicillin-resistant Staphylococcus aureus was observed in 13.7 % of the strains tested, but only two strains inhibited the growth of Candida albicans. The isolated spore-forming bacterial species were also compared with the characteristics of the different Antarctic soils sampled based on their physicochemical properties, showing that pH, C and P were the main factors correlated with the distribution of this group of bacteria in the Antarctic soils studied. These Antarctic endospore-forming bacterial strains may have a potential for industrial processes occurring at low temperatures.     

A possible function of the chilamydospores of Candida albicans

Strains ofCandida albicans produce large numbers of chlamydospores in a liquid medium composed of 1% sodium taurocholate in distilled water. Studies made of these chlamydospores revealed that they do not contain endospores or ascospores but they do contain globules which are lipoid in character. These globules are extruded when chlamydospores split in the liquid medium or under slight pressure. Evidence was not obtained which could support van der Walt's (1967, 1969) statements to the effect that chlamydospores bud, germinate or change into other types of cells. Furthermore, no morphological or cytological evidence was found to substantiate the proposed life cycle ofC. albicans as outlined by van der Walt (1969). It is suggested that a possible function of the chlamydospores ofC. albicans is that of a storage cell.     

Antimicrobial activity and carbohydrate specificity of new mycelial lectins from Fusarium sp.

In the present study, ten Fusarium sp. were screened for the presence of lectins by hemagglutination assay using human and animal erythrocytes. Amongst them nine species, namely F. acuminatum, F. chlamydosporium, F. coeruleum, F. compactum, F. concolor, F. crookwellense, F. culmorum, F. decemcellulare and F. dimerum were found to possess lectin activity. Neuraminidase treatment to rabbit erythrocytes considerably augmented hemagglutination titre, but no such effect was observed with protease-treated erythrocytes. Lectins were tested for inhibition of hemagglutination activity against a panel of carbohydrates. Majority of the lectins were inhibited by L-fucose, D-galactose, bovine submaxillary mucin and dextran. γ-Globulin was inhibitory against lectins from F. acuminatum, F. chlamydosporium, F. compactum and F. culmorum at a concentration of >250 μg/mL, whereas bovine submaxillary mucin and porcine stomach mucin were observed to be strongest inhibitors of lectin from F. compactum with minimum inhibitory concentration of 7.18 μg/mL and 15.6 μg/mL, respectively. Most of the lectins displayed antimicrobial activity against Bacillus cereus, Escherichia coli, Staphylococcus aureus and Aspergillus niger. Lectins from F. chlamydosporium, F. culmorum and F. crookwellense have also exhibited antimicrobial activity against Candida albicans. These findings illustrate the significance of Fusarium sp. lectins in clinical applications.     

Epidemiology and Antifungal Susceptibilities of Yeasts Causing Vulvovaginitis in a Teaching Hospital

Vulvovaginal candidiasis is one of the most common mycosis. However, the information about antifungal susceptibilities of the yeasts causing this infection is scant. We studied 121 yeasts isolated from 118 patients with vulvovaginal candidiasis. The isolates were identified by phenotypic and molecular methods, including four phenotypic methods described to differentiate Candida albicans from C. dubliniensis. Antifungal susceptibility testing was performed according to CLSI documents M27A3 and M27S4 using the drugs available as treatment option in the hospital. Diabetes, any antibacterial and amoxicillin treatment were statistically linked with vulvovaginal candidiasis, while oral contraceptives were not considered a risk factor. Previous azole-based over-the-counter antifungal treatment was statistically associated with non-C.albicans yeasts infections. The most common isolated yeast species was C. albicans (85.2 %) followed by C. glabrata (5 %), Saccharomyces cerevisiae (3.3 %), and C. dubliniensis (2.5 %). Fluconazole- and itraconazole-reduced susceptibility was observed in ten and in only one C. albicans strains, respectively. All the C. glabrata isolates showed low fluconazole MICs. Clotrimazole showed excellent potency against all but seven isolates (three C. glabrata, two S. cerevisiae, one C. albicans and one Picchia anomala). Any of the strains showed nystatin reduced susceptibility. On the other hand, terbinafine was the less potent drug. Antifungal resistance is still a rare phenomenon supporting the use of azole antifungals as empirical treatment of vulvovaginal candidiasis.     

Initial attachment ofCandida albicans cells to buccal epithelial cells

The rapid-freezing technique was applied in association with scanning and transmission electron microscopy to observe the initial attachment (or contact) ofCandida albicans cells to exfoliated human buccal epithelial cells. Low temperature scanning electron microscopy provided detailed three-dimensional morphological features of the yeast-epithelial cell association; adhesion ofC. albicans cells to host cells was primarily owing to an interaction between fibrillar layer of the yeast cell wall and the membrane interdigitations of the epithelial cells. Such a particular interconnection between the two cells was confirmed by the freeze-substitution fixation for transmission electron microscopy. These results clearly demonstrate the outermost fibrillar cell wall layer ofC. albicans responsible for adhesion to host cells.     

Killing Rates for Caspofungin Against Candida albicans After Brief and Continuous Caspofungin Exposure in the Presence and Absence of Serum

It was previously demonstrated that brief (≤1 h) exposures to echinocandins are as effective to kill Candida albicans cells as continuous 24-h exposure. However, killing rates after continuous and short (1 h) echinocandin exposures to C. albicans have not yet been evaluated in RPMI-1640 with and without 50 % serum. We evaluated four echinocandin susceptible C. albicans bloodstream isolates, ATCC 10231 type strain and an echinocandin-resistant isolate (DPL20, FKS F645P). Caspofungin MICs, time-kill and postantifungal effect (PAFE) tests were performed in RPMI-1640 with and without 50 % serum. Killing rates (k values) in time-kill and PAFE experiments were determined for each strain and concentration. In time-kill experiments, colony count decreases were isolate- and concentration-dependent at 0.25, 1, 4, 8, 16 and 32 mg/L in RPMI-1640, but concentration-independent at 1, 4, 8, 16 and 32 mg/L in 50 % serum. One-hour caspofungin exposure at 4, 16 and 32 mg/L resulted in CFU decreases comparable with the results obtained in time-kill experiments in RPMI-1640, but 50 % serum at 4, 16 and 32 mg/L allowed growth of all isolates (k values were negative) (P < 0.05–0.001). PAFE in 50 % serum decreased markedly at 4, 16 and 32 mg/L. Killing rates remained high and concentration-independent in 50 % serum in case of continuous but not in case of brief caspofungin exposure. As only a short growth inhibition without killing was observed in 50 % serum, clinical relevance of caspofungin PAFE in vivo is questionable.     

The effect of parenteral alimentation fluid,undiluted or diluted with saline or frseh sera,on the growth of Candida albicans in vitro at 37 °C

Parenteral alimentation is often complicated by Candida albicans infection which may be fatal. This study investigated the effect of alimentation fluid (Aminosol) on C. albicans' growth in vitro. It was found that concentrated Aminosol (1400 milliosmoles) maintained C. albicans in a viable state but inhibited replication. Dilution of alimentation fluid to physiological concentrations (300 milliosmoles) with either saline or aged pooled normal sera promoted in vitro growth of C. albicans which was equivalent to that obtained in BHI broth and was slightly less than that obtained in Sabouraud's broth. The effects of fresh sera with full complement activity were also investigated. In fresh sera appropriately diluted with physiological saline, some clumping of the yeasts was observed and all formed germ tubes. Growth as defined by budding or the formation of hyphae was inhibited. When Aminosol was diluted to 300 milliosmoles with fresh sera, all yeasts were noted to be in clumps with germ tubes as well as continually growing hyphae. Growth was approximately equal to that seen in Aminosol similarly diluted with saline.     

Non-targeted metabolomic reveals the effect of salt stress on global metabolite of halotolerant yeast Candida versatilis and principal component analysis

As one of the major microbes in the soy sauce fermentation, Candida versatilis enriches the flavor and improves the quality of soy sauce. In this study, a combination of five different GC-MS and LC-MS-based metabolome analytical approaches was used to analyze the intracellular, extracellular and whole metabolites of C. versatilis. Our results found out that a total of 132, 244 and 267 different metabolites were detectable from the intracellular, extracellular and whole part, respectively. When exposed to 0. 9 and 18 % salt, respectively, 114, 123 and 129 different intracellular metabolites, 184, 200 and 178 extracellular metabolites and 177, 188 and 186 whole metabolites were detected, respectively. Our data showed that salt enhances the metabolic capacity of C. versatilis, especially its amino acid and enhances the synthesis and secretion of some metabolites of C. versatilis, especially the aldehydes and phenols, such as vanillin, guaiacol and 5-hydroxymethylfurfural. Our data also showed that special attention has to be paid to the generation of biogenic amines when C. versatilis was treated with salt.     

Intracellular aspartic proteinase Apr1p of Candida albicans is required for morphological transition under nitrogen-limited conditions but not for macrophage killing

Vacuolar hydrolases have been thoroughly characterized in Saccharomyces cerevisiae, but their homologues in the fungal pathogen Candida albicans have received less attention. The genes APR1 and CPY1 of C. albicans encode putative vacuolar aspartic proteinase and serine carboxypeptidase, respectively. We examined properties of apr1Δ and cpy1Δ mutants, showing that Cpy1p molecular species detected in cell lysates of apr1Δ and its parental strain did not differ in molar mass. Processing of Cpy1p precursor is apparently independent of Apr1p. This is in contrast to S. cerevisiae, where vacuolar aspartic proteinase Pep4p is known to participate in the activation of other vacuolar hydrolases including serine carboxypeptidase. We also found that both apr1Δ and cpy1Δ strains are able to form hyphae in nutrient-rich filamentation media. However, proline as a sole nitrogen source induced filamentation only in cpy1Δ and its parental strain, but not in apr1Δ. This indicates the importance of Apr1p for the morphological transition under nitrogen-limited conditions. Despite that, the ability of apr1Δ to kill murine macrophages was not reduced under the conditions tested.     

Effects of antimicrobial peptide L-K6, a temporin-1CEb analog on oral pathogen growth,Streptococcus mutans biofilm formation,and anti-inflammatory activity

Dental caries and periodontitis are common bacterial mouth infections. As a potentially attractive substitute for conventional antibiotics, antimicrobial peptides have been widely tested and used for controlling bacterial infections. In this study, we tested the efficacy of the peptides from the skin secretions of Rana chensinensis for killing several major cariogenic and periodontic pathogens as well as Candida albicans. L-K6, a temporin-1CEb analog, exhibited high antimicrobial activity against the tested oral pathogens and was able to inhibit Streptococcus mutans biofilm formation and reduce 1-day-old S. mutans biofilms with a minimum biofilm inhibitory concentration and reducing concentration of 3.13 and 6.25 μM, respectively. The results of confocal laser scanning microscopy demonstrated that the peptide significantly reduced cell viability within oral biofilms. Furthermore, as little as 5 μM L-K6 significantly inhibited lipopolysaccharide (LPS)- and interleukin-1β-induced productions of interleukin-8 and tumor necrosis factor-α from THP-1 monocytic cells. This anti-inflammatory activity is associated with the binding of L-K6 to LPS and neutralizing LPS-induced proinflammatory responses in THP-1 cells, as well as dissociating LPS aggregates. Our results suggest that L-K6 may have potential clinical applications in treating dental caries by killing S. mutans within dental plaque and acting as anti-inflammatory agents in infected tissues.