Identification of <Emphasis Type="Italic">Candida</Emphasis> Species Isolated from Bovine Mastitic Milk and Their In Vitro Hemolytic Activity in Western Turkey

In this study, identification of 207 Candida isolates, previously isolated from mastitic bovine quarter milk samples at the level of genus, was made using API 20 C AUX system. The most frequently isolated species were Candida krusei (34.8%), followed by Candida rugosa (16.4%), Candida kefyr (12.6%), Candida albicans (10.1%), and Candida tropicalis (9.2%). Less common isolates were Candida zeylanoides (5.8%), Candida parapsilosis (4.3%), Candida guilliermondii (3.4%), Candida famata (1.9%), and Candida glabrata (1.5%). Additionally, in vitro hemolytic activity of all Candida strains were also examined in the present study. C. krusei (72 isolates), C. kefyr (26), C. albicans (21), C. tropicalis (19), C. zeylanoides (12), and C. glabrata (3) demonstrated both alpha and beta hemolysis at 48-h postinoculation. Only alpha hemolysis was detected in C. rugosa (34), C. guilliermondii (7), and C. famata (4), while C. parapsilosis (9) did not show any hemolytic activity after incubation for 72 h. Statistically significant difference (P < 0.001) was determined between the beta-hemolytic activities of Candida strains. The hemolytic activities of C. zeylanoides, C. albicans and C. kefyr were higher than other strains. This is the first study to describe variable hemolysis types exhibited by different Candida strains isolated from bovine mastitic milk in Turkey.     

A study on Candida biofilm growth characteristics and its susceptibility to aureobasidin A

Background

Biofilm is known to contribute to the antifungal resistance of Candida yeasts. Aureobasidin A (AbA), a cyclic depsipeptide targeting fungal sphingolipid biosynthesis, has been shown to be effective against several Candida species.

Response to Oxidative Stress in Eight Pathogenic Yeast Species of the Genus <Emphasis Type="Italic">Candida</Emphasis>

In the course of an infection, the formation of reactive oxygen species by phagocytes and the antioxidant defense mechanisms of microorganisms play a crucial role in pathogenesis. In this study, isolates representing 8 pathogenic Candida species—Candida albicans, Candida dubliniensis, Candida famata, Candida glabrata, Candida guilliermondii, Candida krusei, Candida parapsilosis and Candida tropicalis—were compared with regard to their resistance to oxidative stress in vitro. We evaluated degree of resistance, induction of oxidative damage, capacity to adapt, and induction of antioxidant enzymes. The species showed variable sensitivity to oxidative attack. C. albicans, C. glabrata, and C. krusei were more resistant to oxidative stress under the conditions tested; C. parapsilosis and C. tropicalis presented medium resistance; and C. dubliniensis, C. famata, and C. guilliermondii were more sensitive. The overall greater resistance to oxidative stress of C. albicans and C. glabrata may provide an advantage to these species, which are the major causative agents of candidiasis.     

Oral Candida carriage of patients attending a dental clinic in Braga,Portugal

BackgroundThe ability of the Candida species to colonize surfaces can be considered as a risk factor for oral infection.AimsTo establish oral Candida carriage in patients attending a dental clinic in Braga, Portugal.MethodsA total of 97 patients were analysed. Swab samples were collected and directly cultured onto CHROMagar Candida. Representative yeasts were identified by polymerase chain reaction.ResultsFrom the samples analysed 54.6% (n=53) were Candida positive, and Candida albicans was the most frequently isolated species, accounting for 79% of all the species identified. Non-C. albicans Candida (NCAC) species recovered included Candida parapsilosis, Candida glabrata, Candida tropicalis, and Candida guilliermondii. There was a lack of association between the presence of C. albicans or NCAC species, and age, gender, or prostheses wearing in this population. In 17% of the cases (n=9), polymicrobial cultures, with two different Candida species, were identified.ConclusionsThis study shows a high Candida carriage rate among this population, thus pointing to the relevance of an accurate diagnostic approach in Candida species identification.     

Association between Candida biofilm-forming bloodstream isolates and the clinical evolution in patients with candidemia: An observational nine-year single center study in Mexico

Background

Candidemia is one of the most common nosocomial infections globally and it is associated with considerable excess mortality and costs. Abreast, biofilm-forming strains are associated with even higher mortality rates and poor prognosis for the patient.

Etiología de la candidiasis en pediatría: análisis comparativo con el paciente adulto

BackgroundCandida spp. represents a group of commensal yeasts that can act as pathogens and cause candidiasis in different anatomical locations.AimsThe aim of this study was to perform an epidemiological and comparative analysis between the isolates of Candida spp. in clinical specimens during a three year-period (2010-2012) from children (0-14 years) and adults (15-99 years) in the Valencian Community (RedMIVA).MethodsThe microbiological surveillance network of Valencian Community was used as the information source.Results and conclusionsCandida was isolated in 52,436 patients (1,604 [3.1%] children and 50,832 [96.9%] adults). Candida albicans was significantly (p < 0.05) the predominant species in both age groups, and in almost every type of clinical specimen. The distribution of other species varied depending on the sample type and age group. In blood specimens, Candida parapsilosis followed by C. albicans, Candida famata and Candida lusitaniae were the main species found in children, whereas C. albicans followed by C. parapsilosis, Candida glabrata and Candida tropicalis were the predominant species in adults. In sterile fluids, urine and lower respiratory tract samples, C. parapsilosis was the second most prevalent species in the children group, while C. glabrata and C. tropicalis were the main second species in adults.     

Candida albicans and Non-C. albicans Candida Species: Comparison of Biofilm Production and Metabolic Activity in Biofilms, and Putative Virulence Properties of Isolates from Hospital Environments and Infections

Candida albicans and, more recently, non-C. albicans Candida spp. are considered the most frequent fungi in hospitals. This study analyzed Candida spp. isolates and compared the frequency of different species, that is, C. albicans and non-C. albicans Candida spp., and the origins of isolates, that is, from hospital environments or infections. Yeast virulence factors were evaluated based on biofilm production and metabolic activity. Hemolysin production and the antifungal susceptibility profiles of isolates were also evaluated. Candida spp. were highly prevalent in samples collected from hospital environments, which may provide a reservoir for continuous infections with these yeasts. There were no differences in the biofilm productivity levels and metabolic activities of the environmental and clinical isolates, although the metabolic activities of non-C. albicans Candida spp. biofilms were greater than those of the C. albicans biofilms (p < 0.05). Clinical samples had higher hemolysin production (p < 0.05) and lower susceptibility to fluconazole (p < 0.05). Non-C. albicans Candida spp. predominated in samples collected from hospital environments and infections (p < 0.05). These species had a lower susceptibility to fluconazole and amphotericin B, and their biofilms had higher metabolic activities than those produced by C. albicans, which may explain the increased incidence of fungal infections with these yeasts during recent years.     

Sensibilidad a fluconazol y voriconazol de especies de Candida aisladas de pacientes provenientes de unidades de cuidados intensivos en Medellín,Colombia (2001–2007)

BackgroundDisseminated candidiasis is caused by different Candida species and mainly affects immunocompromised patients and those hospitalized in intensive care units (ICU).ObjectiveOur aim was to determine the frequency and susceptibility of Candida spp. isolates to fluconazole and voriconazole, obtained from patients hospitalized in ICU in the city of Medellin during the years 2001–2007.MethodsThe agar diffusion technique based on the protocols recommended by the CLSI from the United States (M44A) was used. The Chi² test and the Kruskal Wallis statistical methods were used to compare changes in the frequency of Candida spp. isolates and their susceptibility to azoles by year of isolation.ResultsA total of 337 isolates were analyzed, 147 (43.6%) of which corresponded to Candida albicans, followed by 79 (23.4%) Candida tropicalis, 47 (13.9%) Candida parapsilosis, 32 (9.5%) Candida glabrata, 12 (3.6%) Candida guilliermondii and 11 (3.3%) Candida krusei. The remaining isolates (2.7%) were distributed among other species (Candida famata, Candida lusitaniae, Candida lipolytica, Candida pelliculosa and Candida spp.) Most of these isolates (78.3%) were susceptible; 11.9% were dose-dependent susceptible (DDS) and 9.8% resistant to fluconazole. For voriconazole, we observed that 94.1% of the isolates were susceptible, 2.4% DDS and 3.6% resistant.ConclusionsThese data indicate a notable change in the species frequency, as well as a new susceptibility patterns that requires the precise identification of the causative organism and susceptibility testing in order to determine the characteristics of the isolates circulating in ICUs and then to treat them appropriately.     

Species Distribution and Virulence Factors of Candida spp. Isolated from the Oral Cavity of Kidney Transplant Recipients in Brazil

Although yeasts belonging to the genus Candida are frequently seen as commensals in the oral cavity, they possess virulence attributes that contribute for pathogenicity. The aims of the present study were to study the prevalence of Candida spp. isolated from the oral cavity of renal transplant recipients and to analyze strains virulence factors. We isolated a total of 70 Candida strains from 111 transplant recipients, and Candida albicans was the most prevalent species (82.86 %). Oral candidiasis was diagnosed in 14.4 % kidney transplant patients, while 11 isolates (15.7 %) corresponded to non-Candida albicans Candida (NCAC) species. C. albicans adhered to a higher extension than NCAC strains. Some isolates of Candida tropicalis were markedly adherent to human buccal epithelial cells and highly biofilm-forming strains. Regarding proteinase activity, Candida orthopsilosis was more proteolytic than Candida metapsilosis. Candida glabrata and Candida dubliniensis showed very low ability to form biofilm on polystyrene microtiter plates. We have demonstrated here diverse peculiarities of different Candida species regarding the ability to express virulence factors. This study will contribute for the understanding of the natural history and pathogenesis of yeasts belonging to the genus Candida in the oral cavity of patients who were submitted to kidney transplant and are under immunosuppressive therapies.     

The new mutation L321F in Candida albicans ERG11 gene may be associated with fluconazole resistance

BackgroundFor many years fluconazole has been commonly used to treat Candida infections. However, the indiscriminate use of this antimycotic therapy has favored the emergence of resistant isolates. Mutations in the ERG11 gene have been described as one of the primary mechanisms of resistance in Candida species.AimsIn this study we investigated missense mutations in ERG11 genes of Candida albicans, Candida glabrata and Candida tropicalis isolates previously evaluated by susceptibility testing to fluconazole.MethodsScreening for these mutations was performed on 19 Candida clinical isolates (eight C. albicans, five C. glabrata and six C. tropicalis) resistant and susceptible to fluconazole. The ERG11 gene was amplified by PCR with specific primers for each Candida species and analyzed by automated sequencing.ResultsWe identified 14 different missense mutations, five of which had not been described previously. Among them, a new mutation L321F was identified in a fluconazole resistant C. albicans isolate and it was analyzed by a theoretical three-dimensional structure of the ERG11p.ConclusionThe L321F mutation in C. albicans ERG11 gene may be associated with fluconazole resistance.     

Antifungal activity of amniotic fluid against different Candida species

BackgroundAlthough Candida is a commensal of the urogenital tract, intrauterine fungal infections are extremely uncommon in clinical practice.AimsIn the present work we evaluated whether amniotic fluid (AF) possesses direct antifungal activity against clinical isolates of Candida albicans and other Candida species.MethodsA total of 23 AF samples from pregnant women with gestational age of 38–41 weeks were obtained under aseptic conditions by the aspiration of the amniotic sac during cesarean section. Different Candida species were inoculated in amniotic fluid and Sabouraud broth, used as control, and were incubated at 37 °C for 48 h. Quantitative cultures of test samples and controls were performed at 0, 4, 8, 12, 24, and 48 h.ResultsAF collected from 23 pregnant women had consistent and significant inhibitory activity against all Candida isolates tested. Nonetheless, a complete inhibition of growth by all 23 AF samples tested was observed only against Candida glabrata.ConclusionsIt is likely that the antifungal activity of the AF against C. albicans, C. glabrata and Candida parapsilosis observed in vitro also exists in vivo, contributing to protect against intrauterine fungal infections.     

Heparin-benzyl alcohol enhancement of biofilms formation and antifungal susceptibility of vaginal Candida species isolated from pregnant and nonpregnant Saudi women

Biofilm formation by Candida species is a major contribute to their pathogenic potential.The aim of this study was to determine in vitro effects of EDTA, cycloheximide, and heparin-benzyl alcohol preservative on C. albicans (126) and non-albicans (31)vaginal yeast isolates biofilm formations and their susceptibility against three antifungal Etest strips. Results of the crystal violet-assay, indicated that biofilms formation were most commonly observed [100%] for C. kefyr, C. utilis, C. famata, and Rhodotorula mucilaginosa, followed by C. glabrata [70%], C. tropicalis [50%], C. albicans [29%], Saccharomyces cerevisiae [0.0%]. EDTA (0.3mg/ml) significantly inhibited biofilm formation in both C. albicans and non-albicans isolates (P=0.0001) presumably due to chelation of necessary metal cations for the process-completion. In contrast, heparin (-benzyl alcohol preservative) stimulated biofilm formation in all tested isolates, but not at significant level (P=0.567). Conversely, cycloheximide significantly (P=0.0001) inhibited biofilm formation in all C. albicans strains(126) and its effect was even 3 fold more pronounced than EDTA inhibition, probably due to its attenuation of proteins (enzymes) and/or complex molecules necessary for biofilm formation. Results also showed that all nonalbicans yeasts isolates were susceptible to 5-flucytosine (MIC₅₀, 0.016 µg/ml; MIC₉₀, 0.064 µg/ml), but 14% of C. albicans isolates were resistant (MIC₅₀, 0.064 µg/ml; MIC₉₀ >32 µg/ml). The MIC₅₀ value of amphotricin B for all C. albicans and non-albicans isolates was at a narrow range of 0.023 µg /ml, and the MIC₉₀ values were 0.047 µg/ml and 0.064 µg/ml respectively, thereby confirming its efficacy as a first line empiric- treatment of Candida spp infections.     

Phospholipase and Proteinase Activities of Clinical Isolates of Candida from Immunocompromised Patients

Sixty-one isolates of Candida recovered from HIV seropositive and cancer patients were studied for elaboration of putative virulence determinants – phospholipase (PL) and secreted aspartyl proteinase (Sap). Forty two (68.85%) isolates examined were PL producers and 51 (83.6%) were positive for Sap. 57.37% (35/61) isolates produced both enzymes. Enzymatic activity was more pronounced in Candida albicans with 100% PL and 94.1% Sap activity. In contrast, non-C. albicans species demonstrated only 29.6% PL and 70.3% Sap activity, indicating interplay of other virulence determinants in these yeasts in colonization and disease.     

Hemolytic Factor Production by Clinical Isolates of Candida Species

Most cases of fungal bloodstream infections (BIs) are attributed to Candida albicans; however, non-Candida albicans Candida species have recently been identified as common pathogens. Although hemolytic factor is known to be putative virulence factor contributing to pathogenicity in Candida species, its production is poorly evaluated. The present study was undertaken to analyze the production of hemolytic factor by C. albicans (10), C. tropicalis (13), and C. parapsilosis (8) isolates associated with BIs. Data of hemolysis zones on plate assay revealed that the majority of C. albicans isolates produced mild hemolytic activity whereas the majority of C. tropicalis produced strong activity. None of the tested C. parapsilosis isolates exhibited hemolysis on plate assay. We also evaluated the hemolytic activity in the cell-free broth. There were no significant differences (P > 0.05) in the secreted hemolytic activity among intra-species isolates. Different levels of secreted hemolytic factor were observed for Candida species, where C. tropicalis exhibited the highest production of hemolytic factor (P < 0.05) followed by C. albicans and C. parapsilosis. Inhibition of hemolysis (up to 89.12 %) from culture supernatant, following incubation with the lectin Concanavalin A (Con A), was observed for all three Candida species. This finding suggests that the secreted hemolytic factor of C. tropicalis and C. parapsilosis may be a mannoprotein, similar to that described for C. albicans.     

Identification of Candida isolated from the cutaneous candidiasis by the combined use of confirmatory medium and slide agglutination with monofactorial antibodies

Forty strains ofCandida and one ofTorulopsis were isolated from patients with cutaneous candidiasis. The isolates comprised 29 strains ofC. albicans, 7 strains ofC. tropicalis, 2 strains ofC. guilliermondii, and one each ofC. parakrusei, C. lipolytica, andT. famata were identified by the ordinary method. Besides the common pathogenC. albicans, a few other species ofCandida may be etiologic organisms of cutaneous candidiasis. These strains were re-examined by combined use of sucrose agar slants and slide agglutination tests with IgG monofactorial antibodies as a rapid identification method, especially for determining serotypes ofC. albicans. The new method was useful and reliable for rapid identification ofC. albicans and related species. All strains ofC. albicans isolated from skin lesions proved to be standard serotypes ofC. albicans.

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Zusammenfassung Vierzig Stämme vonCandida und eins vonTorulopsis wurden aus Kranken mit kutanen Candidamykosen isoliert. Neunundzwanzig Stämme vonC. albicans, 7 vonC. tropicalis, 2 vonC. guilliermondii, und je einer vonC. parakrusei, C. lipolytica undT. famata wurden mit dem ordinären Methode identifiziert. Außer dem gewohnlichen Erreger,C. albicans, konnten auch ein Paar andere Spezies vonCandida als den Erreger betrachtet werden. Sechsunddreißig Stämme vonC. albicans undC. tropicalis wurden mit der von uns verbesserten kombinierten serologischen und biologischen Methode untersucht, besonders um den Serotypus vonC. albicans festzusetzen. Die neue Methode war gut und zuverlässig als die rapide Identification vonC. albicans und verwandten Spezies. Alle aus der Hautläsion isoliertenC. albicans waren der in Japan allgemeine Serotypus vonC. albicans.

     

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.     

Cyclooxygenase inhibitors reduce biofilm formation and yeast-hypha conversion of fluconazole resistant Candida albicans

The incidence of fluconazole-resistant Candida albicans has been increasing worldwide. Both biofilm and fungal morphogenesis are main virulence factors of C. albicans cells. Extracellular fungal prostaglandins are synthesized during biofilm adhesion and development and through yeast-hypha conversion. Hence, we targeted prostaglandin synthesis with various cyclooxygenase (COX) inhibitors (aspirin, diclofenac, ketoprofen, tenoxicam, and ketorolac) and assessed their effect on fungal adhesion, biofilm formation, and yeast-hypha conversion in clinical isolates of Fluconazole resistant C. albicans. Significant reduction in fungal adhesion and detachment of mature biofilm was attained down to 1 mM concentrations of anti-inflammatory agents. Microscopical examination of fungal cells in the presence of the tested drugs showed significant reduction of germ tube formation. Therefore, COX inhibitors have a significant effect on reduction of Candida adhesion and biofilm development in correlation with fungal morphogenesis. Moreover, inhibition of C. albicans by COX inhibitors gave synergistic activity with fluconazole suggesting that combination therapeutic strategies may be fruitful for management of infection of Fluconazole resistant C. albicans.     

In vitro susceptibility to 5-fluorocytosine and nystatin of common clinical yeast isolates

The sensitivity to nystatin, 5-fluorocytosine (5-FC) or both was studied for 131 clinical isolates of Candida albicans, 47 of Candida parapsilosis, 34 of Candida tropicalis, 7 of Candida guilliermondii, 28 of Torulopsis glabrata and 1 of Torulopsis Candida.All strains were inhibited by concentrations of nystatin within the usual range of sensitivity except one strain of T. glabrata and another of T. Candida whose minima inhibitory concentrations (MICs) were respectively 250 U/ml and > 20000 U/ml.In respect to 5-FC it was found, after 7 days of incubation at 37 °C, the following frequencies of resistance: C. albicans 28/106 (26%), C. parapsilosis 11/47 (23%), C. tropicalis 24/34 (71%), C. guilliermondii 1/7, T. glabrata 1/28 (4%) and T. candida 0/1. It was particularly striking the activity of 5-FC against T. glabrata.     

Biofilm Production by Candida: Comparison of Bloodstream Isolates with Cervical Isolates

The present study was undertaken to investigate biofilm formation among the clinical Candida isolates from blood and cervical swabs. A total of 16 Candida blood isolates from neonates and 21 cervical isolates from pregnant women with vulvovaginitis were included in the study. Each isolate was identified to species level by various phenotypic tests. Biofilm formation was detected by colorimetric method. C. glabrata and C. albicans were the major isolates from blood and cervical swab respectively. The biofilm formation was found in 14 (87.5 %) blood isolates and only in 4 (19.1 %) cervical isolates.