Identification Of Yeasts Research Articles

Case Report: Successful removal of Candida parapsilosis from human embryos with the application of laser and micromanipulation techniques

Objective To describe a micromanipulation method for removal of yeast from contaminated embryos to rescue their development Design Review of 2 concurring IVF/ICSI cases resulting in blastocysts colonized by Candida parapsilosis. Method of yeast removal, isolation and identification of yeast species, and cycle outcome is reviewed. Materials and methods Mature oocytes retrieved from patients were injected with their partner's sperm and cultured under standard conditions. At fertilization, zygotes from two patients showed a contaminating mass in the culture droplets. Embryos from contaminated drops were rinsed multiple times and cultured in freshly equilibrated culture media. The following day culture media had no visual contamination. However, one patient's embryos had hyphae-like masses attached to their zonae pellucidae. Under an inverted microscope attached to a micromanipulator, embryos were held under suction. Hyphae-like masses were removed by applying suction through an assisted hatching pipette and firing a laser to facilitate the detachment of contaminating masses. Cleaned embryos were rinsed and cultured through day 6. All freezable quality blastocysts were vitrified on either day 5/6. Samples of visually contaminated culture drops (Day 1), post-rinse culture drops at the time of embryo vitrification (day 5/6), and stock solutions were sent to a reference lab for microbiological analysis. Patient whose embryos had hyphae mechanically removed underwent subsequent frozen embryo transfer cycle. Results Growth media stock solutions tested positive for Candida parapsilosis. Conclusion Candida parapsilosis hyphae were successfully removed from growing embryos using laser and micromanipulation techniques. After yeast contamination removal, normally developed blastocyst(s) may be transferred only after patient's consent. Disclosures None Funding None To describe a micromanipulation method for removal of yeast from contaminated embryos to rescue their development Review of 2 concurring IVF/ICSI cases resulting in blastocysts colonized by Candida parapsilosis. Method of yeast removal, isolation and identification of yeast species, and cycle outcome is reviewed. Mature oocytes retrieved from patients were injected with their partner's sperm and cultured under standard conditions. At fertilization, zygotes from two patients showed a contaminating mass in the culture droplets. Embryos from contaminated drops were rinsed multiple times and cultured in freshly equilibrated culture media. The following day culture media had no visual contamination. However, one patient's embryos had hyphae-like masses attached to their zonae pellucidae. Under an inverted microscope attached to a micromanipulator, embryos were held under suction. Hyphae-like masses were removed by applying suction through an assisted hatching pipette and firing a laser to facilitate the detachment of contaminating masses. Cleaned embryos were rinsed and cultured through day 6. All freezable quality blastocysts were vitrified on either day 5/6. Samples of visually contaminated culture drops (Day 1), post-rinse culture drops at the time of embryo vitrification (day 5/6), and stock solutions were sent to a reference lab for microbiological analysis. Patient whose embryos had hyphae mechanically removed underwent subsequent frozen embryo transfer cycle. Growth media stock solutions tested positive for Candida parapsilosis. Candida parapsilosis hyphae were successfully removed from growing embryos using laser and micromanipulation techniques. After yeast contamination removal, normally developed blastocyst(s) may be transferred only after patient's consent.

Identification and broth-microdilution antifungal susceptibility testing of yeast directly from automated blood cultures.

Aim: To standardize MALDI-TOF-MS based identification and antifungal susceptibility (AFST) for yeasts directly from automated blood cultures to reduce turnaround time. Materials & methods: Direct-ID after lysis-centrifugation (0.5% SDS) standardized in 40 and validated in 250 yeast positive samples. Direct-AFST was standardized with fluconazole (28 samples) and evaluated (70 samples) for seven antifungals. Results: Direct-ID had a high sensitivity (97.2%) and specificity (94.3%). Correct species-level identification showed 100% in C. tropicalis, C. krusei, C. parapsilosis. Direct-AFST had a 100% categorical agreementwith culture-AFST for posaconazole, anidulafungin and >90% categorical agreement for amphotericin B, voriconazole and fluconazole. Conclusion: Direct-ID and subsequent direct-AFST is a rapid and robust method to reduce the turnaround time for the diagnosis of invasive candidiasis.

Identification of oleaginous yeasts that metabolize aromatic compounds.

The valorization of lignin is critical for the economic viability of the bioeconomy. Microbial metabolism is advantageous for handling the myriad of aromatic compounds resulting from lignin chemical or enzymatic depolymerization. Coupling aromatic metabolism to fatty acid biosynthesis makes possible the production of biofuels, oleochemicals, and other fine/bulk chemicals derived from lignin. Our previous work identified Cutaneotrichosporon oleaginosus as a yeast that could accumulate nearly 70% of its dry cell weight as lipids using aromatics as a sole carbon source. Expanding on this, other oleaginous yeast species were investigated for the metabolism of lignin-relevant monoaromatics. Thirty-six oleaginous yeast species from the Phaff yeast collection were screened for growth on several aromatic compounds representing S-, G-, and H- type lignin. The analysis reported in this study suggests that aromatic metabolism is largely segregated to the Cutaenotrichosporon, Trichosporon, and Rhodotorula clades. Each species tested within each clade has different properties with respect to the aromatics metabolized and the concentrations of aromatics tolerated. The combined analysis suggests that Cutaneotrichosporon yeast are the best suited to broad spectrum aromatic metabolism and support its development as a model system for aromatic metabolism in yeast.

English

Yeasts are very important in food production by affecting the quality and safety of different food products. Yeasts are commonly used in the production of beer, wine and bread; but many yeast species have been isolated from different African fermented foods. The present study aims to determine the biodiversity of yeasts isolated from selected Nigerian fermented foods. A combination of phenotypic and molecular tests were employed in the identification of yeasts. A total of 145 yeasts were isolated from six different food products. The yeasts belong to six genera namely Saccharomyces, Candida, Cyberlindnera, Meyerozyma, Trichosporon and Galactomyces. The most frequently encounter was Saccharomyces cerevisiae followed by Candida glabrata. The present study has confirmed the biodiversity of yeasts isolated from fermented food of Nigerian origin. Key words: Phenotypic characterization, molecular, indigenous yeasts, Nigeria.

Isolation and Identification Of Ascomycetous Yeast Producing Phytase

The objective of the study was to isolate, identify and characterize phytase producing yeast. The yeast was isolated from various resources, and the identification of yeast was conducted by the sequencing of the ITS region. Of 41 isolates, 24 isolated produced phytase. The highest phytse producing yeast was identified as Candida natalensis YEG 132, Saturnispora quitensis YEg 135, and Candida albicans YEg 226. These isolates were able to use various carbon and nitrogen sources. The maximum phytase production depends on isolates, media composition, pH and incubation temperature. Temperature affected phytase production. All isolates produce the highest phytase was at 30°C. Increased temperature to 35° C and 40°C inhibits phytase production by all strains. S. quitensis was the highest isolate inhibited by an increase of incubation temperature. The pH of cultures affects phytase production and varies depending on isolate. S. quitensis and Candida natalesis produced the highest phytase activities at pH 5.0, while C. albicans was at pH 6.0. All isolates were able to assimilate glucose, dextrose, sucrose, xylose and lactose. In general, glucose was the best carbon source for phytase activity. But each isolate has its preferential on carbon sources. S. quitensis produced the highest phytase on glucose, but not on dextrose. Each isolate has preferential on nitrogen sources. Yeast extract was the best N-sources for C. albicans, while ammonium sulfate was best for C. natalensis, S. quitensis. Ammonium nitrate was not favorable for phytase production by C. natalensis. Candida natalensis YEG 132, Saturnispora quitensis YEg 135, and Candida albicans YEg 226 was a good candidate for phytase production.

Separation, Purification and Identification of Excellent Yeasts from the Natural Fermented Beverage of Boza

In this study, 40 yeasts were separated and purified from five Boza samples collected in five different areas of Xinjiang Uyghur Autonomous Region. By adopting traditional morphology observation, preliminary identification, physiological and biochemical experiment, 16 excellent strains with stable fermentation characteristics were obtained, and they were confirmed with the molecular biology identification. Final results show that 12 excellent yeast strains belong to Saccharomyces cerevisiae, one excellent yeast strain belongs to Pichiamembranifaciens, and one excellent yeast strain belongs to Pichifermentans.

Phenotypic and genotypic characterization of Candida albicans isolated from chicken

Candidiasis is an occasional opportunistic yeast disease of importance in poultry and it has also been reported to be a disease or an intestinal infection. The present study was directed mainly to throw lights firstly on the isolation of C. albicans species of chicken and secondly to the identification of the mentioned isolates by phenotypic and genotypic methods. A total 100 of swab samples collected from a crop of chicken from different localities in Cairo and Giza governorate. The samples were investigated for yeast species contamination and mycological examination, which the isolation of four yeast genera. The most frequent yeast species isolated from such samples followed by Candida spp., Saccharomyces spp., Torulopsis spp. and Rhodotorula spp. The observe incidence of Candida species isolated from all examined samples it seems that Candida species were (n=36/60, 60%) from chicken crop swab.The incidence of Candida albicans isolated from examining samples it seems (n=8/36, 22.2%) from crop swab. Early identification of yeast to the genus and species level is necessary for effective antifungal therapy, and can also facilitate control of infections. Explained that Cross-contamination is a particular problem and several recommendations have been published to control pathogens.

Comparison of the ASTA MicroIDSys and VITEK MS matrix-assisted laser desorption/ionization time-of-flight mass spectrometry systems for identification of clinical bacteria and yeasts

The ASTA MicroIDSys system (ASTA, Suwon, Korea) is a newly developed Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) system for identification of microorganisms. We compared the performance of the ASTA MicroIDSys system with that of the VITEK MS system (bioMérieux, Marcy l’Etoile, France) for identifying clinical microorganisms. A total 2055 isolates including 1910 bacteria and 145 yeasts were tested. Among them, the VITEK MS correctly identified 1999 (97.3%) isolates to species level and 26 (1.3%) to the genus level. The ASTA MicroIDSys correctly identified 1988 (96.7%) isolates to species level and 28 (1.4%) to the genus level. The VITEK MS and ASTA MicroIDSys misidentified one isolate and four (0.2%) isolates, respectively, and provided no identification for 29 (1.4%) and 35 (1.7%) isolates, respectively. The performance of the ASTA MicroIDSys was comparable to that of the VITEK MS for identification of clinically relevant bacterial and yeast isolates.

New insights into HcPTR2A and HcPTR2B, two high-affinity peptide transporters from the ectomycorrhizal model fungus Hebeloma cylindrosporum.

While plants mainly rely on the use of inorganic nitrogen sources like ammonium and nitrate, soil-borne microorganisms like the ectomycorrhizal fungus Hebeloma cylindrosporum can also take up soil organic N in the form of amino acids and peptides that they use as nitrogen and carbon sources. Following the previous identification and functional expression in yeast of two PTR-like peptide transporters, the present study details the functions and substrates of HcPTR2A and HcPTR2B by analysing their transport kinetics in Xenopus laevis oocytes. While both transporters mediated high-affinity di- and tripeptide transport, HcPTR2A also showed low-affinity transport of several amino acids-mostly hydrophobic ones with large side chains.

Isolation and Identification of Indigenous Wine Yeasts and Their Use in Alcoholic Fermentation.

SUMMARYResearch backgroundIn our study, spontaneous alcoholic fermentations were carried out to isolate non-Saccharomyces and Saccharomyces yeasts from grape must from different vine-growing regions in Slovenia. Additionally, the diversity of native Saccharomyces cerevisiae strains was evaluated during the process.Experimental approachDuring spontaneous alcoholic fermentations the yeast population of non-Saccharomyces and Saccharomyces yeasts was sampled. We used eleven microsatellite markers to determine the genetic diversity of S. cerevisiae strains. In addition, different ratios of the indigenous strains of S. cerevisiae, Hanseniaspora uvarum and Starmerella bacillaris were tested for their possible use in alcoholic fermentation with inoculated yeasts by monitoring its course and measuring the concentration of aroma compounds in wine.Results and conclusionsSequencing of the internal transcribed spacer (ITS) regions of ribosomal DNA showed that of 64 isolates, 46 strains represent S. cerevisiae and 18 strains belong to non-Saccharomyces yeasts. The identified non-Saccharomyces yeast species were H. uvarum, Pichia kudriavzevii, Saturnispora diversa and S. bacillaris. The dendrogram grouped S. cerevisiae strains into 14 groups. The number of S. cervisiae strains isolated from the musts was 10 (Posavje), 11 (Podravje) and 25 (Primorska vine-growing region). On the other hand, the alcoholic fermentation with inoculated yeasts, in which the native S. cerevisiae strain predominated over H. uvarum and S. bacillaris, gave the most promising result due to the highest alcoholvolume fraction, the lowest acetic acid concentration and significantly higher concentrations of volatile thiols 3-mercaptohexyl acetate (3MHA) and 3-mercaptohexan-1-ol (3MH), 2-methylpropanol, 2-methylbutanol, 3-methylbutanol and 2-phenylethanol) in the produced wine.Novelty and scientific contributionWe confirmed the potential use of indigenous S. cerevisiae and non-Saccharomyces yeasts in alcoholic fermentation with inoculated yeasts, which allows the positive properties of the yeast strains to be expressed and good quality wines to be produced. Thus, the results are encouraging for winemakers to create different wine styles associated with a particular terroir using indigenous yeasts.

Isolation, screening, identification and tolerance of yeast in cherry wine lees

Abstract In this study, yeast was isolated from cherry wine lees by rose Bengal medium, and its species was identified through three-stage screening, morphological observation and molecular biological identification. Moreover, the tolerance of screened strains was studied. The results showed that 30 strains of yeast were isolated from cherry wine lees, and five strains of yeast were selected, which were named YJN10, YJN16, YJN18, YJN19 and YJN28. After preliminary appraisal, strain YJN10 was Saccharomyces kudriavzevii, strain YJN16 was Saccharomyces paradoxus, and strains YJN18, YJN19, YJN28 were Saccharomyces cerevisiae. In the tolerance study, the tolerable sugar concentrations of the five strains were 650, 650, 550, 600 and 600 g/L. The tolerable alcohol volume fractions were 20, 20, 16, 18 and 18%. The tolerable molar concentration of potassium chloride was 1.8, 1.8, 1.5, 1.5 and 1.5 mol/L. Finally, strains YJN10, YJN16, YJN19 and YJN28 showed good tolerance, which laid a foundation for subsequent application in cherry wine fermentation.

Translation Initiation Site Profiling Reveals Widespread Synthesis of Non-AUG-Initiated Protein Isoforms in Yeast.

Genomic analyses in budding yeast have helped define the foundational principles of eukaryotic gene expression. However, in the absence of empirical methods for defining coding regions, these analyses have historically excluded specific classes of possible coding regions, such as those initiating at non-AUG start codons. Here, we applied an experimental approach to globally annotate translation initiation sites in yeast and identified 149 genes with alternative N-terminally extended protein isoforms initiating from near-cognate codons upstream of annotated AUG start codons. These isoforms are produced in concert with canonical isoforms and translated with high specificity, resulting from initiation at only a small subset of possible start codons. The non-AUG initiation driving their production is enriched during meiosis and induced by low eIF5A, which is seen in this context. These findings reveal widespread production of non-canonical protein isoforms and unexpected complexity to the rules by which even a simple eukaryotic genome is decoded.

Molecular identification and characterization of filamentous fungi and yeasts isolated in a pharmaceutical industry environment

Spectroscopic identification of brominated, non-brominated alkaloids and evaluation of antimicrobial activity of Eudistomin-I, Eudistomin H, from green ascidian Eudistoma virideRajaian Pushpabai Rajesh, Annappan Murugan

Effect of fungicides on the yeast population during spontaneous fermentation in the vinification of monastrell grapes

The influence of six fungicides on indigenous yeasts of grape var. Monastrell, after performing two treatments (Good Agricultural Practices-GAP and Critical Agricultural Practices-CAP), was studied. Fungicide residues have been determined using a method of multi-residual extraction that uses QuEChERS and liquid chromatography in tandem with triple quadrupole mass spectrometry. Identification of yeast was carried out by PCR and subsequent sequencing. The fungicide residues are below the EU maximum residue limits (MRLs) established in wine grapes. At the beginning and during the fermentation, most of the treated samples show counts (CFU/mL) higher than the control test (20–30% more), even in the most unfavorable conditions (treated the same day of harvest). It is noteworthy the absence of Hanseniaspora uvarum and the presence of Clavispora lusitaniae, Debaryomyces hansenii and Metschnikowia pulcherrima. Nevertheless, no fungicide either under treatment inhibits fermentation or delays it. Moreover, the evolution of yeast populations found during fermentation follows the normal sequence of species. This research might be of interest for scientists and companies working on either basic or production aspects respectively of the winemaking process.

A Novel Report on Killer Yeast Strains Identification Methods

This study was conducted to isolate and identify killer yeasts from soil samples that collected from different locations in Basrah and Dhi-Qar provinces. Seventy-five soil samples were collected from different areas, including sandy, arable, surface sediment and uncultivated soil, using dilution methods to cultivate a serial dilution of each soil sample. The results showed that a 112 isolates were identified biochemically using VITEK system and molecularly using internal transcribed spacer (ITS1- 5.8S-ITS2) marker. The molecular identification provided fast and precise identification results for the 112 isolates, whereas the VITEK test resulted low identification efficiency (8.2% were accurate and 91.8 % were not). The Diazonium blue B salts produced a good colour reaction in distinguishing between ascomycetes and basidiomycetes. The PCR was more accurate in identification of killer yeasts compared to the VITEK system.

Pathogenicity and drug-resistance analysis of Candida glabrata in patients with oral candidiasis

Objective: To investigate the detection rate, population features and in vitro antifungal drug susceptibility of Candida glabrata in patients with oral candidiasis. Methods: The saliva fungi culture cases and the ratio of Candida glabrata in patients with oral candidiasis were analyzed during 2014-2018. Fungal species were identified by CHROMagar Candida chromogenic medium and API 20C AUX yeast identification kit. Sixty strains with cfu≥200/ml were collected and performed the in vitro antifungal susceptibility tests by using ATB FUNGUS 3 susceptibility kit. Results: The number of patients with oral candidiasis was 12 401 and the ratio of Candida glabrata was 3.44% (427/12 401) from 2014 to 2018. The positive ratio of infection increased with age increasing.The positive rate of female aged 41 to 50 years old began to increased. Female aged more than 50 years old subgroups demonstrated the tendency towards increased positive rates with increasing age. It was found out through clinical data analyzing that the patients with oral candidiasis were the most and oral lichen planus, Sjögren's syndrome and xerostomia carrying high quantity ratios of infection. Candida glabrata had highest resistance(8.3%) to the fluconazole, 6.7% of the isolates were of non-WT phenotype to itraconazole and voriconazole, all of the isolates were of WT phenotype to amphotericin B. Conclusions: The ratio of infection in the patients with oral candidiasis increased with age increasing and the olders are susceptible to infection. Candida glabrata has a certain resistance to azoles and clinically reasonable use of antifungal drugs according to the results of drug susceptibility is very important.

Development and Validation of an in-House Library of Colombian Candida auris Strains with MALDI-TOF MS to Improve Yeast Identification.

Background: Candida auris is characterized for having a high genetic variability among species. MALDI-TOF MS library contains spectra from only three strains of C. auris, which makes difficult the identification process and gives low scores at the species level. Our aim was to construct and validate an internal library to improve C. auris identification with Colombian clinical strains. Methods: From 30 clinical strains, 770 mass spectra were obtained for the construction of the database. The validation was performed with 300 strains to compare the identification results in the BDAL and C. auris Colombia libraries. Results: Our library allowed a complete, 100% identification of the evaluated strains and a significant improvement in the scores obtained, showing a better performance compared to the Bruker BDAL library. Conclusions: The strengthening of the database is a great opportunity to improve the scoring and C. auris identification. Library data are available via ProteomeXchange with identifier PXD016387.

Epidemiology of yeast species causing bloodstream infection in Tehran, Iran (2015-2017); superiority of 21-plex PCR over the Vitek 2 system for yeast identification.

Introduction. Given the limited number of candidaemia studies in Iran, the profile of yeast species causing bloodstream infections (BSIs), especially in adults, remains limited. Although biochemical assays are widely used in developing countries, they produce erroneous results, especially for rare yeast species.Aim. We aimed to assess the profile of yeast species causing BSIs and to compare the accuracy of the Vitek 2 system and 21-plex PCR.Methodology. Yeast blood isolates were retrospectively collected from patients recruited from two tertiary care training hospitals in Tehran from 2015 to 2017. Relevant clinical data were mined. Identification was performed by automated Vitek 2, 21-plex PCR and sequencing of the internal transcribed spacer region (ITS1-5.8S-ITS2).Results. In total, 137 yeast isolates were recovered from 107 patients. The overall all-cause 30-day mortality rate was 47.7 %. Fluconazole was the most widely used systemic antifungal. Candida albicans (58/137, 42.3 %), Candida glabrata (30/137, 21.9 %), Candida parapsilosis sensu stricto (23/137, 16.8 %), Candida tropicalis (10/137, 7.3 %) and Pichia kudriavzevii (Candida krusei) (4/137, 2.9 %) constituted almost 90 % of the isolates and 10 % of the species detected were rare yeast species (12/137; 8.7 %). The 21-plex PCR method correctly identified 97.1 % of the isolates, a higher percentage than the Vitek 2 showed (87.6 %).Conclusion. C. albicans was the main cause of yeast-derived fungaemia in this study. Future prospective studies are warranted to closely monitor the epidemiological landscape of yeast species causing BSIs in Iran. The superiority of 21-plex PCR over automated Vitek 2 indicates its potential clinical utility as an alternative identification tool use in developing countries.

Use of MALDI-TOF MS for fungal species distribution of interdigital intertrigo in seafarers, Dakar, Senegal

To determine fungal species distribution of interdigital intertrigo among seafarers in Dakar, Senegal, a cross-sectional study was carried out from May to August 2017 among seafarers clinically diagnosed with interdigital intertrigo. A questionnaire was filled to each patient before sampling the affected folds and transporting to Aristide Le Dantec University Hospital where mycological analyses were realized. Species identification by MALDI-TOF MS was performed in Marseille, France. In total, 169 men (21–66 years) were included. Few of them (3%) had a high level of education and the duration of the mycosis exceed 10 years for 88% of patients. Direct microscopic examination (ME) was positive in 34.3%. Among samples with positive ME, 58.6% had positive culture. An overall incidence of 30.2% was found. Patients with confirmed cases aged between 28 and 66 years. Among them, those between 36–50 years were predominant (52.9%). Those with a high level of education were less representative (2%). For 52.1% of patients, the duration of the mycosis was superior to 10 years. Furthermore, 57% of cases were significantly associated with other types of tinea pedis and/or onychomycosis (P=0.03). Culture was positive in 23.7% isolating 43 strains successfully identified at the species level by MALDI-TOF MS for 31 isolates: 20 Candida and 11 dermatophytes. The rest was identified only at the genus level belonged to Fusarium. In definitive, MALDI-TOF MS could be a useful tool for routine and fast identification of dermatophytes, yeasts and NDFF in clinical mycology laboratories.

Molecular identification and antifungal susceptibility profile of yeast from vulvovaginal candidiasis

BackgroundAccurate identification Candida is important for successful therapy and epidemiology study. The aim of research is to study API 20C yeast identification system identification rate by using molecular identification as gold standard and tested the antifungal susceptibility of Candida from patients with vulvovaginal candidiasis (VVC).MethodsIn total, 3574 yeast isolates were obtained from patients with VVC. API 20C yeast identification, molecular identification and in vitro antifungal susceptibility were performed.ResultsC. albicans was the predominant Candida species [2748 isolates, 76.9%] in VVC. The isolates from vaginal samples represented 22 species based on molecular identification. The API 20C system identifies only 11 of the species encountered during the study period. Based on the API 20C system, 3273 (91.58%) isolates were correctly identified to the species level. The correct identification rate of the API 20C system for rare yeast was 15.29% (26/170 isolates). Antifungal susceptibility was tested in a total of 1844 isolates of Candida from patients with VVC. C. albicans was susceptible to most of the tested antifungals. The MICs of azoles for C. glabrata were higher than those for C. albicans. The MICs of echinocandins for C. parapsilosis were higher than those for C. albicans.ConclusionsThe API 20C yeast identification system can be used to reliably identify the most common Candida species while molecular methods are necessary for the identification of closely related, emerging, and rare yeast species. The results from this study suggest that much of the previous studies on the epidemiology of VVC should be re-thought. C. albicans was susceptible to most of the tested antifungals.