Biodiversity of Zygosaccharomyces species in food systems

Abstracts Zygosaccharomyces species are among the most problematic food spoilage yeasts. The two most infamous species are Zygosaccharomyces balii and Zygosaccharomyces rouxii, although they may also take a positive role during the production of some fermented foods. DNA sequence based yeast identification aided by freely available reference databases of barcoding DNA sequences has boosted the description rate of novel yeast species in the last two decades. The genus Zygosaccharomyces has been considerably expanded as well. Especially the number of the extremely osmotolerant Zygosaccharomyces species, related to Z. rouxii and regularly found in high-sugar foods, has enlarged. A brief account of recent developments in the taxonomy and biodiversity of this important food associated genus is given in this review.

Yeasts Associated with the Small-Intestinal Contents and Epithelium of Pon Yang Kham (Charolais Crossbred) Fattening Beef Cattle

Yeast diversity in the pia and small-intestinal epithelium of Pon Yang Kham fattening cattle in Thailand was studied using a culture-dependent method. A total of 701 yeasts were isolated from the pia of the duodenum, jejunum, and ileum of the small intestine, while 425 isolates were obtained from the epithelium of all three parts of the small intestine. Yeast identification was performed and ascomycetous yeasts were found at levels of 96.9% and 86.8% in the pia and small intestine, respectively, whereas basidiomycetous yeasts were found at levels of 2.3% and 12.7%. Candida parapsilosis was the species with the highest occurrence in the duodenal and jejunal pia, with an 83.3% and 77.8% frequency of occurrence (FO), respectively. Both C. parapsilosis and C. tropicalis were species with the highest occurrence in the ileum, with a 61.1% FO. Moreover, C. parapsilosis was the species with the highest occurrence in the epithelium of the duodenum, jejunum, and ileum, with FOs of 88.2%, 87.5%, and 87.2%, respectively. Principal coordinate analysis revealed no marked differences in yeast communities from either the pia or epithelium of all three parts of the small intestine. An estimation of the expected richness of the species showed that the observed species richness was lower than the predicted richness.

Population Dynamics and Yeast Diversity in Early Winemaking Stages without Sulfites Revealed by Three Complementary Approaches

Nowadays, the use of sulfur dioxide (SO2) during the winemaking process is a controversial societal issue. In order to reduce its use, various alternatives are emerging, in particular bioprotection by adding yeasts, with different impacts on yeast microbiota in early winemaking stages. In this study, quantitative-PCR and metabarcoding high-throughput sequencing (HTS) were combined with MALDI-TOF-MS to monitor yeast population dynamic and diversity in the early stages of red winemaking process without sulfites and with bioprotection by Torulaspora delbrueckii and Metschnikowia pulcherrima addition. By using standard procedures for yeast protein extraction and a laboratory-specific database of wine yeasts, identification at species level of 95% of the isolates was successfully achieved by MALDI-TOF-MS, thus confirming that it is a promising method for wine yeast identification. The different approaches confirmed the implantation and the niche occupation of bioprotection leading to the decrease of fungal communities (HTS) and Hanseniaspora uvarum cultivable population (MALDI-TOF MS). Yeast and fungi diversity was impacted by stage of maceration and, to a lesser extent, by bioprotection and SO2, resulting in a modification of the nature and abundance of the operational taxonomic units (OTUs) diversity.

Salt concentration effect on yeast diversity of the Ivorian traditional fermented fish adjuevan

This study aimed to investigate the yeast diversity in adjuevan fermented at the laboratory scale according to salt concentration following both traditional fermentation methods. Thus, the fish species Galeoides decadactylus was fermented with salt added at 10 %, 15 %, 20 %, 25 % and 30 % (w/w) for five days. Yeast identification using PCR-DDGE method reveled seven species which were Pichia fermentans, Candida zeylanoides, Candida sp, Hanseniaspora osmophila, Kluyveromyces sp; Torulaspora delbrueckii and Kluyveromyces marxianus. These species varied according to fermentation method used and salt concentration added with Pichia fermentans and Hanseniaspora osmophila as dominant strains. These results showed also that Kluyveromyces marxianus and Torulaspora delbrueckii were more tolerant to sodium chloride than the others. This work confirmed that yeasts were involved and participated in adjuevan production. Therefore, these yeasts should be tested for their functionality during the fermentation, and some might be useful as starter culture to produce better quality fermented fish adjuevan.

Viable Yeast Identification using Bag of Visual Words in Colored images

In this research it is reported a system to automate the process of identification of viable yeasts whose population control is a crucial task in the ethanol production process. The identification and counting of yeasts made by human vision under a light microscope, is repetitive and susceptible to errors. We used computer vision techniques such as BoVW, Color Coherence Vectors (CCV), Color Moments (CM), Bag-of-Color (BoC) and Opponent Color (OpC) were applied for extracting characteristics that were classified by the Naive Bayes, KNN, SVM and J48 algorithms in 2614 images of yeasts separated into three classes: viable, non-viable and background. The results were analyzed using software R, which in the ANOVA test resulted in a p value equal to 2e􀀀16 indicating a significant difference between the techniques. The OPC with SVM classifier showed the highest performance using the PCC Percent Correct Classification metric, about 95% compared to other techniques.

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.

Optimization of MALDI-ToF mass spectrometry for yeast identification: a multicenter study

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS) is routinely used in mycology laboratories to rapidly identify pathogenic yeasts. Various methods have been proposed to perform routine MS-based identification of clinically relevant species. In this study, we focused on Bruker technology and assessed the identification performance of three protocols: two pretreatment methods (rapid formic acid extraction directly performed on targets and full extraction using formic acid/acetonitrile in tubes) and a direct deposit protocol that omits the extraction step. We also examined identification performance using three target types (ground-steel, polished-steel, and biotargets) and two databases (Bruker and online MSI [biological-mass-spectrometry-identification application]) in a multicenter manner. Ten European centers participated in the study, in which a total of 1511 yeast isolates were analyzed. The 10 centers prospectively performed the three protocols on approximately 150 yeast isolates each, and the corresponding spectra were then assessed against two reference spectra databases (MSI and Bruker), with appropriate thresholds. Three centers evaluated the impact of the targets. Scores were compared between the various combinations, and identification accuracy was assessed. The protocol omitting the extraction step was inappropriate for yeast identification, while the full extraction method yielded far better results. Rapid formic acid extraction yielded variable results depending on the target, database and threshold. Selecting the optimal extraction method in combination with the appropriate target, database and threshold may enable simple and accurate identification of clinically relevant yeast samples. Concerning the widely used polished-steel targets, the full extraction method still ensured better scores and better identification rates.

Identification of Candida auris by Use of the Updated Vitek 2 Yeast Identification System, Version 8.01: a Multilaboratory Evaluation Study

ABSTRACT Candida auris is an emerging multidrug-resistant yeast that has been systematically incorrectly identified by phenotypic methods in clinical microbiology laboratories. The Vitek 2 automated identification system (bioMérieux) recently included C. auris in its database (version 8.01). We evaluated the performance of the Vitek 2 YST ID card to identify C. auris and related species. A panel of 44 isolates of Candida species (C. auris, n = 35; Candida haemulonii, n = 5; Candida duobushaemulonii, n = 4) were tested by three different hospital-based microbiology laboratories. Among 35 isolates of C. auris, Vitek 2 yielded correct identification in an average of 52% of tested samples. Low-discrimination (LD) results with an inability to distinguish between C. auris, C. duobushaemulonii, and Candida famata were obtained in an average of 27% of samples. Incorrect identification results were obtained in an average of 21% of samples, the majority (91%) of which were reported as C. duobushaemulonii and the remaining 9% of which were reported as Candida lusitaniae/C. duobushaemulonii. The proportion of correct identification was not statistically different across different centers (P = 0.78). Stratification by genetic clades demonstrated that 100% (n = 8) of the strains of the South American clade were correctly identified compared to 7% (n = 10) and 0% (n = 4) from the African and East Asian clades, respectively. None of the non-auris Candida strains (n = 9) were incorrectly identified as C. auris. Our results show that the Vitek 2 (version 8.01) yeast identification system has a limited ability to correctly identify C. auris. These data suggest that an identification result for C. duobushaemulonii should warrant further testing to rule out C. auris. The overall performance of the Vitek 2 seems to differ according to C. auris genetic clade, with the South American isolates yielding the most accurate results.

RareKodamaea ohmerikeratitis following a trivial vegetative trauma

Kodamaea ohmerikeratitis is an opportunistic pathogen seen in patients who have undergone invasive procedures and immunocompromised state. It has been identified in septicemia patients, resulting in mortality. To the best of our knowledge, we identified the first case ofK. ohmerikeratitis following an injury with vegetative material. A 57-year-old woman with underlying, poorly controlled diabetes mellitus was gardening when a tree leaf accidentally poked her in the eye. Two weeks later, the patient presented with right eye pain, redness and progressive blurring of vision due to a traumatised right cornea. Slit-lamp examination showed a small inferior paracentral corneal stromal infiltrate with overlying epithelial defect. A corneal scraping sample yieldedK. ohmerifrom Analytical Profile Index (API) 20C yeast identification system. She was treated with intensive topical amphotericin B and fluconazole. After 6 weeks of treatment, the keratitis resolved with faint scar tissue, and her visual acuity improved.