Yeast Extract Peptone Dextrose Agar Research Articles

A New Species of Ganoderma (Ganodermataceae, Polyporales) from Southern China and Optimum Condition for Mycelia Production

The present study sought to propose Ganoderma guixiense sp. nov. as a new species based on phenotypic and genotypic evidence. Phylogenetic analyses were carried out based on the internal transcribed spacer (ITS), the large subunit of nuclear ribosomal RNA gene (nLSU), and the second subunit of RNA polymerase II (RPB2) sequence data. G. guixiense has been characterized by pileate basidiomata, long stipe, in addition to reddish-black zonate pileal surface. Basidiospores are broadly ellipsoid with one end tapering at maturity, and measuring 9–12.8 × 6.5–9.3 μm. Basidia are oval to subglobose. This study marks the first exploration of the biological characteristics of G. guixiense. The result indicated that the optimal medium of mycelial growth was observed on malt extract agar (MEA) and yeast extract peptone dextrose agar (YPD) while the optimal temperature was found to be 25–30 °C with pH range of 6–7.

Isolation and identification of non-saccharomyces ethanol and thermo-tolerant yeasts strains from fermented carbohydrate wastes

Ethanol is an important chemical with its potential as a biofuel to displace the environmentally unfriendly fossil fuels. Isolation and identification of ethanol and thermo-tolerant yeast capable of producing ethanol were carried out in this study. Seventy (70) samples from different fermented foods and agricultural wastes from millets, sorghum, maize, rice and guinea corn were obtained from local sellers in Panteka, Farin-Gida, Barkallahu and Barnawa in Kaduna State (North West), Nigeria were used in this research. The samples were collected using a sterile plastic container and were transported immediately to the Biotechnology laboratory of the Nigerian Defence Academy in iced chamber. They were serially diluted in distilled water and inoculated on yeast extract peptone dextrose agar (YEPD) media plates supplemented with 0.2% chloramphenicol. The plates were incubated at 30°C for 72 hours and developed colonies were sub-cultured to obtain pure culture which was observed for their morphological and microscopic characteristics. After examination, ten (10) yeast isolates were obtained from the seventy (70) samples collected. Flocculation test, Hydrogen sulphide test, preliminary alcohol production and sugar fermentation test were carried out on the isolates and their tolerance to ethanol at varying concentration 5%-20% (v/v) and temperature at 37oC-45oC were studied. Using the internal transcribed spacer (ITS) 1 and 2 regions primer, the DNA were sequenced and the isolates identified. Four isolates FG 17, FG 27, P1 and BN50 showed ethanol tolerance potentials at 20% ethanol and were identified as strains of Diutina rugosa, Candida sp, Candida ethanolica and Candida tropicalis while one isolate FG 35 showed good thermo-tolerance ability at 45oC and was identified as Trichosporon sp.

Begerowomyces aurantius sp. nov., a novel yeast isolated from koalas' habitat in a Japanese zoological park.

Koalas are iconic mammals indigenous to Australia. These rare animals and their habitats are occasionally associated with pathogenic fungi, including species of Cryptococcus, and consequently, monitoring the mycobiota of areas inhabited by koalas is of considerable importance. In this report, we describe a novel basidiomycetous yeast isolated from a site in Kanazawa Zoo, Japan, associated with captive koalas. Swab samples were collected from koala breeding environments, from which we isolated a novel unencapsulated yeast characterized by ovoid to ellipsoidal cells (3.2-4.9 × 3.5-5 μm). These cells were observed to undergo polar budding and grow as parent bud pairs, with an optimal growth temperature of 28°C. Colonies grown on yeast extract peptone dextrose agar at 28°C have a characteristic coral pink color. On the basis of physiological, morphological, and molecular characters, the new species was placed in the genus Begerowomyces, and the name Begerowomyces aurantius JCM33898T(LSEM1333T=CBS16241T) is proposed.

P015 A study to demonstrate heteroresistance and tolerance to azoles in Candida tropicalis

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM Objective Candida tropicalis rarely exhibits a trailing growth phenomenon in in vitro antifungal susceptibility testing (AFST) of azoles. This phenomenon is considered as ‘sensitive’ as per current CLSI guidelines. This phenomenon has been linked to treatment failure and such isolates could either be heteroresistant (HR) or tolerant, which is known with bacteria. HR subtype (<1%) of an isogenic susceptible cell population grows at drug concentrations at least 8 times higher than the minimum inhibitory concentration (MIC). Tolerant cells (5%-90% of the population) endure antimicrobial treatment several times above MIC. Characterizing these isolates could help relate their clinical susceptibility profiles to the treatment outcome. The aim of the study was to differentiate heteroresistance and tolerance from true azole resistance in C. tropicalis clinical isolates.MethodsA total of 247 clinical isolates of C. tropicalis collected over a period of 7 months (September 2021 through March 2022) from PGIMER, Chandigarh were included in the study. Identification was done by MALDI-TOF MS and antifungal susceptibility testing to fluconazole was performed by CLSI broth microdilution method (M27-A3). Isolates exhibiting trailing growth phenomena were considered for further characterization. Cells from the trailing growth of each isolate were sub-cultured onto yeast extract peptone dextrose (YPD) agar to observe for phenotypic variations with and without fluconazole. The area under the population analysis (PAP) curve (AUC) was performed to determine the degree of heteroresistance. Yeast cells within the range of 102-105 cells/ml were spot inoculated in six replicates each onto a gradient of fluconazole (0.125-256 μg/ml) in YPD agar and incubated at 35°C for 5 days to determine the viable colony forming units (CFU).ResultsOut of the 225 fluconazole susceptible C. tropicalis isolates, 10 (4.4%) were found to exhibit a trailing growth phenomenon. A dose-response relationship showed a multimodal population distribution in these isolates with varying degrees of heteroresistance as demonstrated by area under curve that ranged from 171-1331. The heterogeneous subpopulations sampled from the growth at the highest fluconazole concentration (64 μg/ml) exhibited a similar MIC (± one 2-fold difference) as that of the parent isolate when tested individually. The heterogeneity range that determines a fold difference breakpoint of the isolates varied from 16 to >256.ConclusionAn isogenic population of cells under the effect of fluconazole could give rise to phenotypically different subpopulations. With repeated exposure to the azole drugs, these seemingly susceptible isolates can emerge as fully resistant population. Clinically, the implications include relapse, treatment failure, and persistent chronic infection, owing to which this phenomenon needs attention. Current CLSI guidelines do not provide any criteria to separately classify these isolates from the susceptible and resistant varieties. Hence a definitive cut-off is warranted to identify the HR and tolerant subtypes. The AUC-PAP method could be refined further for discriminating heteroresistance from true resistance.

Potentials of Saccharomyces cerevisiae Y10 as a Starter Culture for Wine Production from Date Palm (Phoenix dactylifera) Fruits

A considerable quantity of agricultural produce in Nigeria gets spoilt due to lack of storage facilities and poor storage conditions. An example of such agricultural produce is date palm which is readily available in the northern parts of Nigeria. This study aimed at evaluating the potential of Saccharomyces cerevisiae Y10 isolated from decomposed sugarcane bagasse to produce wine using date palm fruits. The yeast was isolated using yeast extract peptone dextrose agar and was identified by molecular methods. Fermentation of date palm fruit juice was carried out anaerobically for 20 days during which physicochemical parameters and proximate composition were determined using standard methods. The yeast was identified as Saccharomyces cerevisiae Y10 with accession number MG321589. Analysis of physicochemical properties showed that pH dropped from 5.4 to 3.6, titratable acidity increased from 0.25 to 0.71% and specific gravity decreased from 1.1 to 1.01. Proximate composition of produced wine gave low total reducing sugar and low protein of 0.11 and 0.44% respectively and the final alcohol content was 10.4% at the 20th day of fermentation. This study has shown that Saccharomyces cerevisiae Y10 which was isolated from decaying sugarcane bagasse sample has the potential to be used as starter culture for fermentation of date palm for wine production.

Production, Partial Purification and Characterization of Two α-Amylase Isoforms from Saccharomyces cerevisiae strain YOP 1/2-2 Isolated from Tchapalo (Côte d’Ivoire)

Amylases play an important role in biotechnology and find applications in several industrial fields such as pharmaceutical, food, paper, cosmetics and detergents. Thus, it appears necessary to identify new sources of amylase, especially from microbial origin, due to the low energy consumption, cost-effective, high metabolic diversity, rapid cell growth, non-toxic and eco-friendly characteristics. In the present report, we carried out the production and partial purification of α-amylase by Saccharomyces cerevisiae strains isolated from Tchapalo, a traditional alcoholic beverage of Côte d'Ivoire. Five fungal isolates were screened initially for α-amylase production by using method of wells on Yeast Extract Peptone Dextrose Agar medium, a complete medium for yeast growth. One step DEAE-Sepharose Fast Flow was achieved for partial purification of α-amylase obtained. Among yeasts, isolate S. cerevisiae YOP 1/2-2 was able to provoke starch hydrolysis halo of 15.067±0.12 mm on starch agar plate after 48 h of incubation at 30°C. The partial purification of resulting enzyme showed two protein peaks, designated α-amylase 1 (AMY1) and α-amylase 2 (AMY2) with amylolytic activity and specific activities of 1.57-1.58 U/mg protein. Both isoforms (AMY1 and AMY2) were thermostable with optimal activity at 50 and 55°C, respectively, and at pH ranged from 4.5 to 5.3 in 0.1 M sodium acetate buffer. EDTA and Cd2+ strongly inhibited α-amylase activity by 72-75% and 64-65%, respectively, whereas cations Ca2+ and Mn2+ showed 85-99% and 71% increased amylolytic activity, respectively. All these properties show the potential uses of α-amylases from S. cerevisiae in the industrial transformation of starch.

Identification of Yeasts with Mass Spectrometry during Wine Production

The aim of the present study was to identify yeasts in grape, new wine “federweisser” and unfiltered wine samples. A total amount of 30 grapes, 30 new wine samples and 30 wine samples (15 white and 15 red) were collected from August until September, 2018, from a local Slovak winemaker, including Green Veltliner (3), Mūller Thurgau (3), Palava (3), Rhein Riesling (3), Sauvignon Blanc (3), Alibernet (3), André (3), Blue Frankish (3), Cabernet Sauvignon (3), and Dornfelder (3) grapes; federweisser and unfiltered wine samples were also used in our study. Wort agar (WA), yeast extract peptone dextrose agar (YPDA), malt extract agar (MEA) and Sabouraud dextrose agar (SDA) were used for microbiological testing of yeasts. MALDI-TOF Mass Spectrometry (Microflex LT/SH) (Bruker Daltonics, Germany) was used for the identification of yeasts. A total of 1668 isolates were identified with mass spectrometry. The most isolated species from the grapes was Hanseniaspora uvarum, and from federweisser and the wine—Saccharomyces cerevisiae.

Isolation, Identification, and Characterization of Pectinolytic Yeasts for Starter Culture in Coffee Fermentation.

This experiment was carried out to identify and select pectinolytic yeasts that have potential use as a starter culture for coffee fermentation during wet processing. The coffee fruit was fermented for 48 h at 28 °C and a sample was taken from the fermented solution and spread onto yeast extract-peptone-dextrose agar (YPDA) media and incubated at 28 °C. A total of 28 yeasts were isolated, eight of which had the ability to produce pectinase enzymes. The species of those eight yeasts were molecularly identified and confirmed. These yeasts are Wickerhamomyces anomalus (strain KNU18Y3), Saccharomycopsis fibuligera (strain KNU18Y4), Papiliotrema flavescens (strain KNU18Y5 and KNU18Y6), Pichia kudriavzevii (strain KNU18Y7 and KNU18Y8), and Saccharomyces cerevisiae (strain KNU18Y12 and KNU18Y13). The pectin degradation index of S. fibuligera (strain KNU18Y4), W. anomalus (strain KNU18Y3), and P. flavescens (strain KNU18Y6) were higher compared to the others, at 178%, 160%, and 152%, respectively. The pectinase enzyme assays were made on two growth media: coffee pulp media (CPM) and synthetic pectin media (SPM). S. fibuligera (strain KNU18Y4) and W. anomalus (strain KNU18Y3) had great potential in producing polygalacturonase (PG) and pectin lyase (PL) compared to others in both media. However, S. cerevisiae strains (KNU18Y12 and KNU18Y13) produced higher pectin methylesterase (PME). Using MEGA 6 software, the phylogenetic trees were constructed to determine the evolutionary relationship of newly identified yeasts from our experiment and previously published yeast species. The sequences of the yeasts were deposited in the National Center for Biotechnology Information (NCBI) database.

A survey to examine Candida albicans transmission through yolk sac and amniotic sac in chicken embryos

Candida albicans is the fungus responsible for the incidence of candidiasis. This fungus, in low volumes, is a natural inhabitant in various organs of birds which can act as a pathogen in some circumstances. Infection patterns are used to study microbial pathogenesis. The infection patterns can be used in the evaluation of host-pathogen interaction, pathogenicity, virulence agents, and remedy studies. In order to eliminate the diversity and differences of the infection patterns between invertebrates and mice, this study introduced an alternative, low-cost, and simple application of infection patterns for Candida albicans based on embryonated eggs. Similar to other infection patterns, embryonated eggs can be used as an acceptable pattern to study fungal pathogenesis. In this study, systemic candidiasis was achieved in chicken embryos by yolk sac and amniotic sac inoculations of Candida albicans and its histopathological demonstrations were examined in three organs including liver, kidney, and lung. Candida albicans was subcultured once on yeast extract-peptone-dextrose (YPD) agar. Plating serial dilutions on YPD agar plates was carried out to confirm the infectious doses. Subsequently, inoculation into yolk sac and amniotic sac was carried out with the intended infectious dose. After 18 days of incubation, histopathologic specimens were taken from embryonated chicken organs. Predominant lesions were observed in liver, kidney, and lung which included the accumulation of mononuclear cells. In conclusion, this study corroborated the embryo transmission of Candida albicans via yolk sack and amniotic routes by focusing on the importance of hygiene and biosecurity in breeder flocks and particularly in hatcheries.

Heavy metal tolerance in marine strains of Yarrowia lipolytica.

Heavy metal tolerance of two marine strains of Yarrowia lipolytica was tested on solid yeast extract peptone dextrose agar plates. Based on minimum inhibitory concentration esteems, it is inferred that the two strains of Y. lipolytica were tolerant to heavy metals such as Pb(II), Cr(III), Zn(II), Cu(II), As(V), and Ni(II) ions. The impact of various heavy metal concentrations on the growth kinetics of Y. lipolytica was likewise assessed. With increased heavy metal concentration, the specific growth rate was reduced with delayed doubling time. Furthermore, biofilm development of both yeasts on the glass surfaces and in microtitre plates was assessed in presence of different heavy metals. In microtitre plates, a short lag phase of biofilm formation was noticed without the addition of heavy metals in yeast nitrogen base liquid media. A lag phase was extended over increasing metal concentrations of media. Heavy metals like Cr(VI), Cd(II), and As(V) are contrastingly influenced on biofilms' formation of microtitre plates. Other heavy metals did not much influence on biofilms development. Thus, biofilm formation is a strategy of Y. lipolytica under stress of heavy metals has significance in bioremediation process for recovery of heavy metals from contaminated environment.

Evaluating the probiotic and therapeutic potentials of Saccharomyces cerevisiae strain (OBS2) isolated from fermented nectar of toddy palm

The purpose of this study is to evaluate the probiotic characteristics of 15 yeast strains isolated from nectar of toddy palm. Initially, the collected samples were inoculated on yeast extract peptone dextrose agar plates and the colonies so obtained were culturally and morphologically characterized. Commercial probiotic yeast, Saccharomyces boulardii served as the control in these experiments. Of the 15 yeast strains, the isolates that were resistant to antibiotics and worked synergistically with other cultures were considered for further evaluation. Selected isolates were evaluated in vitro for tolerance to simulated gastrointestinal conditions such as temperature, pH, bile and gastric juice. Further the yeast isolates were evaluated for their pathogenicity and adherence to intestinal epithelial cells. The 2 yeast isolates with efficient probiotic properties were finally characterized by sequencing their 5.8 S rRNA and partial sequences of internal transcribed spacer 1 and 2. The sequences were BLAST searched in the National Center for Biotechnology Information, nucleic acid database for sequence similarity of organisms and phylogenetic evolutionary analysis was carried out. Based on maximum similarity of basic local alignment search tool results, organisms were characterized as Pichia kudriavzevii OBS1 (100%) and Saccharomyces cerevisiae OBS2 (96%) and sequences were finally deposited in the GenBank data library. Among these two isolates, S. cerevisiae OBS2 displayed slight/moderate antioxidant and anticancer property. Hence, strain OBS2 can be utilized and explored as a potential probiotic for therapeutic applications.

Polymer-immobilized ready-to-use recombinant yeast assays for the detection of endocrine disruptive compounds

Recombinant yeast assays (RYAs) constitute a suitable tool for the environmental monitoring of compounds with endocrine disrupting activities, notably estrogenicity and androgenicity. Conventional procedures require yeast reconstitution from frozen stock, which usually takes several days and demands additional equipment. With the aim of applying such assays to field studies and making them more accessible to less well-equipped laboratories, we have optimized RYA by the immobilization of Saccharomyces cerevisiae cells in three different polymer matrices – gelatin, Bacto agar, and Yeast Extract Peptone Dextrose agar – to obtain a ready-to-use version for the fast assessment of estrogenic and androgenic potencies of compounds and environmental samples. Among the three matrices, gelatin showed the best results for both testosterone (androgen receptor yeast strain; AR-RYA) and 17β-estradiol (estrogen receptor yeast strain; ER-RYA). AR-RYA was characterized by a lowest observed effect concentration (LOEC), EC50 and induction factor (IF) of 1nM, 2.2nM and 51, respectively. The values characterizing ER-RYA were 0.4nM, 1.8nM, and 63, respectively. Gelatin immobilization retained yeast viability and sensitivity for more than 90d of storage at 4°C. The use of the immobilized yeast reduced the assay duration to only 3h without necessity of sterile conditions. Because immobilized RYA can be performed either in multiwell microplates or glass tubes, it allows multiple samples to be tested at once, and easy adaptation to existing portable devices for direct in-field applications.

Investigation of the expression levels of efflux pumps in fluconazole-resistant Candida albicans isolates

Widespread and repeated use of fluconazole in the prophylaxis and therapy resulted in resistance among Candida strains. Investigation of the expression of efflux pump encoding genes was aimed in fluconazole-resistant and -susceptible C.albicans isolates in order to determine the role of this mechanism in fluconazole resistance. Five fluconazole-resistant, six -susceptible and four trailing effect showing susceptible C.albicans isolates were included in the study. The MIC values of fluconazole and other antifungal agents were determined by the microdilution method. The fluconazole MIC values of the fluconazole-resistant strains were also studied by E-test performed on yeast extract peptone dextrose agar with and without cyclosporin A. The expression levels of CDR1, CDR2 and MDR1 transcripts were determined by real-time PCR method. The expression of these genes was normalized with their ACT1 levels and compared with the fluconazole-susceptible C.albicans ATCC 14053 strain. It was detected that all strains were susceptible to amphotericin B and all except one strain were also susceptible to clotrimazole. Three out of five fluconazole-resistant strains and three out of four trailing effect showing susceptible strains were resistant to 5-flucytosine, and all except one susceptible strains were found as intermediate to 5-flucytosine. All except one fluconazole-resistant strains were determined as resistant to itraconazole and ketoconazole, and had miconazole MIC values of ≥ 64 µg/ml. All fluconazole-susceptible isolates were detected to be susceptible to ketoconazole and dose dependent susceptible to itraconazole. Fluconazole-resistant and -susceptible strains were determined as susceptible to voriconazole. Out of five fluconazole-resistant isolates, two strains overexpressed high levels and three strains overexpressed mild levels of CDR1/2; one strain overexpressed high levels and three strains overexpressed low levels of MDR1 in comparison to C.albicans ATCC 14053 control strain. It was observed that CDR1, CDR2 and MDR1 gene expression levels were mild in strains showing trailing effect except one which highly expressed MDR1, while susceptible isolates except three expressed efflux pump genes at low levels. It was determined that the expression levels of CDR1 and CDR2 genes for the same strain were in parallel for all isolates. It can be concluded that overexpression of efflux pump genes is an important mechanism of resistance in fluconazole-resistant C.albicans isolates.

Evaluation of Sporidiobolus pararoseus strain YCXT3 as biocontrol agent of Botrytis cinerea on post-harvest strawberry fruits

This study evaluated Sporidiobolus pararoseus (Sp) strain YCXT3 as biocontrol agent of Botrytis cinerea (Bc), the causal agent of strawberry gray mold disease. Efficacy of live yeast cells and volatile organic compounds (VOCs) of Sp in suppression of Bc on strawberry fruits was determined. Results showed that in dual cultures of Sp and Bc on potato dextrose agar at 20°C, Sp did not inhibit mycelial growth of Bc. However, inoculation of the yeast cell suspensions of Sp (1×105 or 1×106 yeast cells ml−1) on strawberry fruits resulted in reducing the disease incidence from 96–100% in the control treatment to 39–50% in the Sp treatment and the disease severity index from 5.1–7.0 in the control treatment to 1.1–1.9 in the Sp treatment. We found that the VOCs from the Sp cultures on yeast extract peptone dextrose agar were highly effective in inhibiting both the conidial germination and the mycelial growth of Bc. A total of 39 VOCs, including 2-ethyl-1-hexanol, were identified in cultures of Sp using GC–MS. Authentic 2-ethyl-1-hexanol was found to have strong anti-fungal activity against Bc with the IC50 values of 1.5 and 5.4μll−1 for conidial germination and mycelial growth, respectively. The VOCs from the Sp cultures were effective in suppression of gray mold disease under the air-tight conditions. This study suggests that the strain YCXT3 of Sp is a promising agent for control of Bc and production of VOCs is a valid biocontrol mechanism for this yeast strain.

Phospholipase Cγ2 (PLCγ2) Is Key Component in Dectin-2 Signaling Pathway, Mediating Anti-fungal Innate Immune Responses

C-type lectin receptors (CLRs) such as Dectin-2 function as pattern recognition receptors to sense fungal infection. However, the signaling pathways induced by these receptors remain largely unknown. Previous studies suggest that the CLR-induced signaling pathway may utilize similar signaling components as the B cell receptor-induced signaling pathway. Phospholipase Cγ2 (PLCγ2) is a key component in B cell receptor signaling, but its role in other signaling pathways has not been fully characterized. Here, we show that PLCγ2 functions downstream of Dectin-2 in response to the stimulation by the hyphal form of Candida albicans, an opportunistic pathogenic fungus. Using PLCγ2- and PLCγ1-deficient macrophages, we found that the lack of PLCγ2, but not PLCγ1, impairs cytokine production in response to infection with C. albicans. PLCγ2 deficiency results in the defective activation of NF-κB and MAPK and a significantly reduced production of reactive oxygen species following fungal challenge. In addition, PLCγ2-deficient mice are defective in clearing C. albicans infection in vivo. Together, these findings demonstrate that PLCγ2 plays a critical role in CLR-induced signaling pathways, governing antifungal innate immune responses.

Control of Postharvest Botrytis Fruit Rot of Strawberry by Volatile Organic Compounds of Candida intermedia

A study was conducted to identify volatile organic compounds or volatiles produced by Candida intermedia strain C410 using gas chromatography-mass spectrometry, and to determine efficacy of the volatiles of C. intermedia in suppression of conidial germination and mycelial growth of Botrytis cinerea and control of Botrytis fruit rot of strawberry. Results showed that, among 49 volatiles (esters, alcohols, alkenes, alkanes, alkynes, organic acids, ketones, and aldehydes) identified from C. intermedia cultures on yeast extract peptone dextrose agar, two compounds, 1,3,5,7-cyclooctatetraene and 3-methyl-1-butanol, were the most abundant. Synthetic chemicals of 1,3,5,7-cyclooctatetraene; 3-methyl-1-butanol; 2-nonanone; pentanoic acid, 4-methyl-, ethyl ester; 3-methyl-1-butanol, acetate; acetic acid, pentyl ester; and hexanoic acid, ethyl ester were highly inhibitory to conidial germination and mycelial growth of B. cinerea. Inhibition of conidial germination and mycelial growth of B. cinerea by volatiles of C. intermedia was also observed. Meanwhile, results showed that incidence and severity of Botrytis fruit rot of strawberry was significantly (P < 0.01) reduced by exposure of the strawberry fruit to the volatiles from C. intermedia cultures or C. intermedia-infested strawberry fruit. These results suggest that the volatiles of C. intermedia C410 are promising biofumigants for control of Botrytis fruit rot of strawberry.

WdStuAp, an APSES Transcription Factor, Is a Regulator of Yeast-Hyphal Transitions in Wangiella ( Exophiala ) dermatitidis

APSES transcription factors are well-known regulators of fungal cellular development and differentiation. To study the function of an APSES protein in the fungus Wangiella dermatitidis, a conidiogenous and polymorphic agent of human phaeohyphomycosis with yeast predominance, the APSES transcription factor gene WdSTUA was cloned, sequenced, disrupted, and overexpressed. Analysis showed that its derived protein was most similar to the APSES proteins of other conidiogenous molds and had its APSES DNA-binding domain located in the amino-terminal half. Deletion of WdSTUA in W. dermatitidis induced convoluted instead of normal smooth colony surface growth on the rich yeast maintenance agar medium yeast extract-peptone-dextrose agar (YPDA) at 37 degrees C. Additionally, deletion of WdSTUA repressed aerial hyphal growth, conidiation, and invasive hyphal growth on the nitrogen-poor, hypha-inducing agar medium potato dextrose agar (PDA) at 25 degrees C. Ectopic overexpression of WdSTUA repressed the convoluted colony surface growth on YPDA at 37 degrees C, and also strongly repressed hyphal growth on PDA at 25 degrees C and 37 degrees C. These new results provide additional insights into the diverse roles played by APSES factors in fungi. They also suggest that the transcription factor encoded by WdSTUA is both a positive and negative morphotype regulator in W. dermatitidis and possibly other of the numerous human pathogenic, conidiogenous fungi capable of yeast growth.

Conidium production by insect pathogenic fungi on commercially available agars.

Conidium production by three species of insect pathogenic fungi, Metarhizium anisopliae, Beauveria bassiana and Verticillium lecanii, was assessed on various depths and types of commercially available agars. Conidium production was assessed after 14 d of growth on commercially available media as well as at three different agar depths. Metarhizium anisopliae and B. bassiana isolates showed greatest conidium production on potato dextrose agar (PDA) at a depth of 2 mm, whereas V. lecanii showed greatest conidium production on yeast extract-peptone-dextrose agar (YPDA) regardless of agar depth. Optimum conidium production for M. anisopliae and B. bassiana was not only dependent upon the isolate used but also on the medium type and agar depth. Conidia are the infective structures for insect pathogenic fungi and this study suggests a rationale basis for consistent conidium production for laboratory and commercial practices.

In vitro Candidacidal effect of polymorphonuclear neutrophils of Behçet's patients with or without ocular involvement.

In this study our objective was to evaluate the in vitro candidacidal effect of polymorphonuclear neutrophils of Behçet's patients with or without ocular involvement (inactive phase). Fifteen patients with ocular involvement and 15 patients without ocular involvement were studied. Candidacidal assay was performed to assess the in vitro killing of Candida species by the polymorphonuclear neutrophils of both the study and control groups. In brief, peripheral venous polymorphonuclear neutrophils of healthy volunteers and patients with Behçet's disease were obtained by density gradient centrifugation. These cells were incubated with Candida spp. (Candida albicans, Candida tropicalis, Candida glabrata) for 3 h. Samples were seeded onto yeast extract peptone dextrose agar, incubated at 25 degrees C in a 5% humidified incubator for another 48 h, and formed colonies were counted. There was no significant difference between the candidacidal activity of polymorphonuclear neutrophils of healthy volunteers and of Behçet's patients with or without ocular involvement. Although various immunological abnormalities are reported in Behçet's disease, in vitro killing of Candida spp., which is one of the parameters for the phagocytic function of polymorphonuclear neutrophils, was not affected.

Comparison of static and step-pulsed ultra-high pressure on the microbial stability of fresh cut pineapple

Fresh cut pineapple cubes inoculated with 104–5 CFU g−1 Saccharomyces cerevisiae were packed in heat-sealed polyethylene pouches and subjected at ambient temperature to static and step-pulsed ultra-high pressure (UHP) treatments. Static treatments included 100 and 9000 s at 270 MPa and 9000 s at 340 MPa. Step-pulsed pressure treatments included 100, 300 and 600 s at 0–270 MPa using 0·5-s and 10-s pulses. Inoculated treated and untreated samples were held at 4°C for 60 days. Bacteria and yeast counts were determined using plate count agar and yeast extract peptone dextrose agar, respectively. Static treatment at 270 and 340 MPa for 9000 s resulted in <240 CFU g−1 yeast and bacteria counts for up to 60 days. Step-pulsed pressure treatments for 100 s at 0–270 MPa using 0·5-s (200 pulses) and 10-s pulses (10 pulses) were more effective than a 100-s static 270-MPa treatment. Step-pulsed pressure treatments (300 and 600 s) using 0·5-s pulses (600 and 1200 pulses) and 10-s pulses (30 and 60 pulses) were as effective as 9000-s static pressure treatments at 270 and 340 MPa. This storage study confirmed the superiority of step-pulsed over static pressure treatments. © 1998 SCI.