Kondoa tianchiensis f.a., sp. nov., an anamorphic yeast species isolated from plant leaves.

Two yeast strains, NYNU 236122 and NYNU 236180, were isolated from plant leaves collected in Tianchi Mountain, Henan Province, central China. Molecular phylogenetic analyses revealed the closest relatives of the strains are three described Kondoa species, Kondoa chamaenerii, Kondoa miscanthi, and Kondoa subrosea. Genetically, the isolated strains differed from the type strains of their three related species by 2-11(0.2-1.8%) base substitutions in the D1/D2 domain, 16-40 (2.6-5.6%) base mismatches in the internal transcribed spacer region, and more than 10.1% base substitutions in the partial RPB2 gene. Furthermore, the two strains differ physiologically from their closest related species, K. chamaenerii, in their ability to assimilate dl-lactate, nitrite, and l-lysine and their inability to assimilate nitrate. Additionally, they differ from K. miscanthi and K. subrosea in their ability to assimilate inulin, d-gluconate, and l-lysine. The species name of Kondoa tianchiensis f.a., sp. nov. is proposed with holotype CICC 33616T (Mycobank MB 853544).

Spencermartinsiella nicolii sp. nov., a potential opportunistic pathogenic yeast species isolated from rotting wood in Brazil.

Nineteen isolates representing a candidate for a novel yeast species belonging to the genus Spencermartinsiella were recovered from rotting wood samples collected at different sites in Atlantic Rainforest and Amazonian Forest ecosystems in Brazil. Similarity search of the nucleotide sequence of the intergenic spacer (ITS)-5.8S and large subunit D1/D2 regions of the ribosomal gene cluster showed that this novel yeast is closely related to Spencermartinsiella cellulosicola. The isolates differ by four nucleotide substitutions in the D1/D2 domain and six substitutions and 31 indels in the ITS region from the holotype of S. cellulosicola. Phylogenomic analysis based on 1474 single-copy orthologues for a set of Spencermartinsiella species whose whole genome sequences are available confirmed that the novel species is phylogenetically close to S. cellulosicola. The low average nucleotide identity value of 83% observed between S. cellulosicola and the candidate species confirms that they are distinct. The novel species produced asci with hemispherical ascospores. The name Spencermartinsiella nicolii sp. nov. is proposed. The holotype is CBS 14238T. The MycoBank number is MB855027. Interestingly, the D1/D2 sequence of the S. nicolii was identical to that of an uncultured strain of Spencermartinsiella causing systemic infection in a male adult crocodile (Crocodylus niloticus). The characterization of some virulence factors and antifungal susceptibility of S. nicolii isolates suggest that this yeast may be an opportunistic pathogen for animals, including humans; the isolates grow at 37 °C.

A Cluster of Diutina catenulata Funguria in Patients with Coronavirus Disease 2019 (COVID-19) Hospitalized in a Tertiary Reference Hospital from Rio de Janeiro, Brazil.

During the COVID-19 pandemic, fungal infections, especially pulmonary aspergillosis, mucormycosis, and invasive candidiasis, have emerged as a significant health concern. Beyond Candida albicans, the most common cause of invasive candidiasis, other rare ascomycetous yeast species have been described in tertiary care units, potentially posing a broader health threat. We have isolated, from September 2020 to June 2021, nine Diutina catenulata strains from urine samples of six patients. This was intriguing as this fungus had not been previously identified in our institution, nor after June 2021. Therefore, we decided to outline the clinical features of the patients with this rare pathogen, to describe phenotypic characteristics, including antifungal susceptibility profiles, of this yeast species and to identify the genetic makeup through whole-genome sequencing analysis to evaluate if this was a cluster of genetically similar D. catenulata isolates in our institution. The strains were identified through MALDI-TOF MS analyses and Sanger sequencing of two rDNA regions. All patients yielding D. catenulata from urine samples needed ventilator support and used urinary catheters during hospitalization for treatment of COVID-19. None of them had received COVID-19 vaccines. Morphological and biochemical profiles of the nine strains were largely consistent, although fluconazole susceptibility varied, ranging from 4 to 32 µg/mL. Phylogenomic analysis revealed minimal genetic variation among the isolates, with low intrapopulation variation, supported by the identification of only 84 SNPs across all strains. Therefore, we propose that the yeast strains isolated were part of a cluster of D. catenulata funguria in the context of COVID-19.

Reduction of Ribosomal Expansion Segments in Yeast Species of the Magnusiomyces/Saprochaete Clade.

Ribosomes are ribonucleoprotein complexes highly conserved across all domains of life. The size differences of ribosomal RNAs (rRNAs) can be mainly attributed to variable regions termed expansion segments (ESs) protruding out from the ribosomal surface. The ESs were found to be involved in a range of processes including ribosome biogenesis and maturation, translation, and co-translational protein modification. Here, we analyze the rRNAs of the yeasts from the Magnusiomyces/Saprochaete clade belonging to the basal lineages of the subphylum Saccharomycotina. We find that these yeasts are missing more than 400 nt from the 25S rRNA and 150 nt from the 18S rRNAs when compared to their canonical counterparts in Saccharomyces cerevisiae. The missing regions mostly map to ESs, thus representing a shift toward a minimal rRNA structure. Despite the structural changes in rRNAs, we did not identify dramatic alterations in the ribosomal protein inventories. We also show that the size-reduced rRNAs are not limited to the species of the Magnusiomyces/Saprochaete clade, indicating that the shortening of ESs happened independently in several other lineages of the subphylum Saccharomycotina.

Cyberlindnera qingyuanensis f.a., sp. nov., a yeast species isolated from rotting wood.

Two yeast strains were isolated from rotting wood samples collected on Qingyuan Mountain, Fujian Province, PR China. Phylogenetic analysis, based on the concatenated sequences of the internal transcribed spacer (ITS) region and the D1/D2 domain of the large subunit rRNA gene, revealed that these two strains represent a novel species of the genus Cyberlindnera. The proposed name for this new species is Cyberlindnera qingyuanensis f.a., sp. nov. (holotype: GDMCC 2.300; ex-type: PYCC 9925) although the formation of ascospores was not observed. The novel species differs from its close relative Cyberlindnera galapagoensis by 7.7% sequence divergence (37 substitutions and seven indels) in the D1/D2 domain and 9.7% sequence divergence (42 substitutions and 34 indels) in the ITS region, respectively. Additionally, Cyb. qingyuanensis differs from its close relative Cyb. galapagoensis by its ability to grow in cellobiose, l-rhamnose, ribitol, galactitol, and dl-lactate, its growth at 37 °C, and its inability to ferment raffinose. The Mycobank number is MB 854693.

Spathaspora marinasilvae sp. nov., a xylose-fermenting yeast isolated from galleries of passalid beetles and rotting wood in the Amazonian rainforest biome.

Four yeast isolates were obtained from rotting wood and galleries of passalid beetles collected in different sites of the Brazilian Amazonian Rainforest in Brazil. This yeast produces unconjugated allantoid asci each with a single elongated ascospore with curved ends. Sequence analysis of the internal transcribed spacer-5.8 S region and the D1/D2 domains of the large subunit ribosomal RNA (rRNA) gene showed that the isolates represent a novel species of the genus Spathaspora. The novel species is phylogenetically related to a subclade containing Spathaspora arborariae and Spathaspora suhii. Phylogenomic analysis based on 1884 single-copy orthologs for a set of Spathaspora species whose whole genome sequences are available confirmed that the novel species represented by strain UFMG-CM-Y285 is phylogenetically close to Sp. arborariae. The name Spathaspora marinasilvae sp. nov. is proposed to accommodate the novel species. The holotype of Sp. marinasilvae is CBS 13467 T (MycoBank 852799). The novel species was able to accumulate xylitol and produce ethanol from d-xylose, a trait of biotechnological interest common to several species of the genus Spathaspora.

Mrakia polaris sp. nov. and Mrakia amundsenii sp. nov. (Mrakiaceae, Cystofilobasidiales), two novel psychrophilic yeast species isolated from Arctic habitats.

Four yeast strains belonging to the basidiomycetous yeast genus Mrakia were isolated from diverse habitats in the Ny-Ålesund region (Svalbard, High Arctic): two from vascular plants, one from seawater and one from freshwater. Phylogenetic analysis, based on the ITS region and the D1/D2 domain of the 28S rRNA gene, identified these four strains as representing two novel species within the genus Mrakia. The names Mrakia polaris sp. nov. (MycoBank number: MB 852063) and Mrakia amundsenii sp. nov. (MycoBank number: MB 852064) are proposed. These two new species show distinct psychrophilic adaptations, as they exhibit optimal growth at temperatures between 10 and 15°C, while being unable to grow at 25°C. The holotype of M. polaris sp. nov. is CPCC 300345T, and the holotype of M. amundsenii sp. nov. is CPCC 300572T.

Wickerhamiella lachancei f.a. sp. nov., a novel ascomycetous yeast species isolated from flowers of Lantana camara in India.

Two isolates representing a novel species of the genus Wickerhamiella were obtained in India from nectar of flowers of Lantana camara, an ornamental exotic species native to Central and South America. Phylogenetic analyses of the D1/D2 domain of the 26S large subunit (LSU) rRNA gene, internal transcribed spacer (ITS) region, and physiological characteristics, supported the recognition of the novel species, that we designate Wickerhamiella lachancei sp. nov (MycoBank no. MB851709), with MCC 9929T as the holotype and PYCC 10003T as the isotype. Considering pairwise sequence similarity, the type strain of the novel species differs from the type strain of the most closely related species, Wickerhamiella drosophilae CBS 8459T, by 16 nucleotide substitutions and two gaps (3.9 % sequence variation) in the D1/D2 region (560 bp compared) and 28 nucleotide substitutions and five gaps (7.22 % sequence variation) in the ITS region (444 bp compared).

Wickerhamiella nakhonpathomensis f.a. sp. nov., a novel ascomycetous yeast species isolated from a mushroom and a flower in Thailand.

Strains SU22T (TBRC 14875T) and FLA11.5, representing a novel anamorphic yeast species, were respectively isolated from a fruiting body of a Coprinus species and an inflorescence of a Coffea species collected in Thailand. Analysis of the sequences of the D1/D2 domains of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) regions showed that the two strains differed by two nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and were identical in the ITS regions. Wickerhamiella drosophilae CBS 8459T was the most closely related species, but with 24-26 nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and 24 nucleotide substitutions in the ITS regions. A phylogenetic analysis, based on the sequences of the D1/D2 domains, indicated that the two strains represented a species in the genus Wickerhamiella which was distinct from other recognized species of the genus. Therefore, the two strains were assigned as a novel species, for which we propose the name Wickerhamiella nakhonpathomensis f.a. sp. nov. The holotype is TBRC 14875T (isotype PYCC 8914T). The MycoBank number of the novel species is MB 840833.

Analysis of glucose and xylose metabolism in new indigenous Meyerozyma caribbica strains isolated from corn residues.

Aiming to broaden the base of knowledge about wild yeasts, four new indigenous strains were isolated from corn residues, and phylogenetic-tree assemblings on ITS and LSU regions indicated they belong to Meyerozyma caribbica. Yeasts were cultivated under full- and micro-aerobiosis, starting with low or high cell-density inoculum, in synthetic medium or corn hydrolysate containing glucose and/or xylose. Cells were able to assimilate both monosaccharides, albeit by different metabolic routes (fermentative or respiratory). They grew faster in glucose, with lag phases ~ 10 h shorter than in xylose. The hexose exhaustion occurred between 24 and 34 h, while xylose was entirely consumed in the last few hours of cultivation (44-48 h). In batch fermentation in synthetic medium with high cell density, under full-aerobiosis, 18-20 g glucose l-1 were exhausted in 4-6 h, with a production of 6.5-7.0 g ethanol l-1. In the xylose medium, cells needed > 12 h to consume the carbohydrate, and instead of ethanol, cells released 4.4-6.4 g l-1 xylitol. Under micro-aerobiosis, yeasts were unable to assimilate xylose, and glucose was more slowly consumed, although the ethanol yield was the theoretical maximum. When inoculated into the hydrolysate, cells needed 4-6 h to deplete glucose, and xylose had a maximum consumption of 57%. Considering that the hydrolysate contained ~ 3 g l-1 acetic acid, it probably has impaired sugar metabolism. Thus, this study increases the fund of knowledge regarding indigenous yeasts and reveals the biotechnological potential of these strains.

Metahyphopichia suwanaadthiae sp. nov., an anamorphic yeast species in the order Saccharomycetales and reassignment of Candida silvanorum to the genus Metahyphopichia.

Seven yeast strains, representing a single novel anamorphic species, were isolated in Thailand. They consisted of five strains (DMKU-MRY16T, DMKU-SK18, DMKU-SK25, DMKU-SK30 and DMKU-SK32) obtained from five different mushrooms, and two strains (ST-224 and 11-14.2) derived from insect frass and soil, respectively. The pairwise sequence analysis indicated that all seven strains had identical sequences in the D1/D2 domains of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) region. Metahyphopichia silvanorum was the most closely related species, but with 11.9-12.4% nucleotide substitutions in the D1/D2 domains of the LSU rRNA gene and 13.1-13.3% nucleotide substitutions in the ITS region. The phylogenetic analyses based on the concatenated sequences of the ITS region and the D1/D2 domains of the LSU rRNA gene showed that the seven strains form a well-separated subclade in a clade containing M. silvanorum and Metahyphopichia laotica with high bootstrap support. A phylogenetic analysis of a multilocus dataset including the small subunit (SSU) rRNA gene, the ITS region, the D1/D2 domains of the LSU rRNA gene, translation elongation factor 1-alpha gene, actin gene and the RNA polymerase II subunit 2 gene, confirmed the presence of the monophyletic clade that also includes M. silvanorum and M. laotica, and strongly supported the phylogenetic isolation of the seven strains from its neighbouring species. Therefore, the seven strains were assigned as a single novel species of the genus Metahyphopichia, according to their phylogenetic relationships. The name Metahyphopichia suwanaadthiae sp. nov. is proposed to accommodate the seven strains. The holotype is DMKU-MRY16T (TBRC 11775T=NBRC 114386T=PYCC 8655T). The MycoBank number of the novel species is MB 841280. In addition, Candida silvanorum is reassigned to the genus Metahyphopichia. The MycoBank number of M. silvanorum comb. nov. is MB 841279.

Wickerhamiella martinezcruziae f. a., sp. nov., a yeast species isolated from tropical habitats.

Four yeast isolates with an affinity to the genus Wickerhamiella were obtained from beach sand, a marine zoanthid and a tree exudate at different localities in Brazil. Two other isolates with almost identical ITS and D1/D2 sequences of the large subunit rRNA gene were isolated from the small intestine of cattle and a grease trap in Thailand. These isolates represent a novel species phylogenetically related to Wickerhamiella verensis, Wickerhamiella osmotolerans, Wickerhamiella tropicalis, Wickerhamiella sorbophila and Wickerhamiella infanticola. The novel species differs by 15-30 nucleotide differences from these species in the D1/D2 sequences. The name Wickerhamiella martinezcruziae f.a., sp. nov. is proposed. The holotype of Wickerhamiella martinezcruziae sp. nov. is CBS 16104T. The MycoBank number is MB 839328.

Diversity of non-Saccharomyces yeasts of grape berry surfaces from representative Cabernet Sauvignon vineyards in Henan Province, China.

Non-Saccharomyces yeasts are important players during winemaking and may come from grapes grown in vineyards. To study the diversity of non-Saccharomyces yeasts on grape berry surfaces, 433 strains were isolated from different Cabernet Sauvignon vineyards grown in Henan Province. Our results demonstrated that these strains were classified into 16 morphotypes according to their growth morphology on Wallerstein Laboratory agar medium, and were identified as seven species from four genera-Hanseniaspora opuntiae, Hanseniaspora vineae, Hanseniaspora uvarum, Pichia occidentalis, Pichia kluyveri, Issatchenkia terricola and Saturnispora diversa-based on a series of molecular biological experiments. Hanseniaspora opuntiae was obtained from all sampling sites except Changyuan County, while Pichia kluyveri and Saturnispora diversa were only found in sites of Zhengzhou Grape Resource Garden and Minquan County, respectively. The site Minquan was home of the greatest species richness, while only one single species (Hanseniaspora opuntiae) was detected at NAPA winery from Zhengzhou or at Anyang County. Finally, this study suggested that the geographic distribution and diversity of non-Saccharomyces yeast populations on Cabernet Sauvignon grape berries were likely to be determined by a combination of grape varieties and environmental factors.

Cyberlindnera dasilvae sp. nov., a xylitol-producing yeast species isolated from rotting wood and frass of cerambycid larva.

Six yeast isolates were obtained from rotting wood samples in Brazil and frass of a cerambycid beetle larva in French Guiana. Sequence analysis of the ITS-5.8S region and the D1/D2 domains of the large subunit rRNA gene showed that the isolates represent a novel species of Cyberlindnera. This novel species is related to Cyberlindnera japonica, Cyberlindnera xylosilytica, Candida easanensis and Candida maesa. It is heterothallic and produces asci with two or four hat-shaped ascospores. The name Cyberlindnera dasilvae sp. nov. is proposed to accommodate the novel species. The holotype of Cy. dasilvae is CBS 16129T and the designated paratype is CBS 16584. The MycoBank number is 838252. All isolates of Cy. dasilvae were able to convert xylose into xylitol with maximum xylitol production within 60 and 72 h. The isolates produced xylitol with values ranging from 12.61 to 31.79 g l-1 in yeast extract-peptone-xylose medium with 5% xylose. When the isolates were tested in sugarcane bagasse hydrolysate containing around 35-38 g l-1d-xylose, isolate UFMG-CM-Y519 showed maximum xylitol production.

The Genome of the CTG(Ser1) Yeast Scheffersomyces stipitis Is Plastic.

Microorganisms need to adapt to environmental changes, and genome plasticity can lead to rapid adaptation to hostile environments by increasing genetic diversity. Here, we investigate genome plasticity in the CTG(Ser1) yeast Scheffersomyces stipitis, an organism with an enormous potential for second-generation biofuel production. We demonstrate that S. stipitis has an intrinsically plastic genome and that different S. stipitis isolates have genomes with distinct chromosome organizations. Real-time evolution experiments show that S. stipitis genome plasticity is common and rapid since extensive genomic changes with fitness benefits are detected following in vitro evolution experiments. Hybrid MinION Nanopore and Illumina genome sequencing identify retrotransposons as major drivers of genome diversity. Indeed, the number and position of retrotransposons are different in different S. stipitis isolates, and retrotransposon-rich regions of the genome are sites of chromosome rearrangements. Our findings provide important insights into the adaptation strategies of the CTG(Ser1) yeast clade and have critical implications in the development of second-generation biofuels. These data highlight that genome plasticity is an essential factor for developing sustainable S. stipitis platforms for second-generation biofuels production. IMPORTANCE Genomes contain genes encoding the information needed to build the organism and allow it to grow and develop. Genomes are described as stable structures where genes have specific positions within a chromosome. Changes in gene dosage and position are viewed as harmful. However, it is becoming increasingly clear that genome plasticity can benefit microbial organisms that need to adapt rapidly to environmental changes. Mechanisms of genome plasticity are still poorly understood. This study focuses on Scheffersomyces stipitis, a yeast that holds great potential for second-generation biofuel production generated from forestry and agriculture waste. We demonstrate that S. stipitis chromosomes are easily reshuffled and that chromosome reshuffling is linked to adaptation to hostile environments. Genome sequencing demonstrates that mobile genetic elements, called transposons, mediate S. stipitis genome reshuffling. These data highlight that understanding genome plasticity is important for developing sustainable S. stipitis platforms for second-generation biofuels production.

Microbial community diversity associated with Tibetan kefir grains and its detoxification of Ochratoxin A during fermentation.

Tibetan kefir grains (TKG) are multi-functional starter cultures used in foods and have been applied in various fermentation systems. This study aimed to investigate the microbial community composition of TKG, the detoxification abilities of TKG and their isolates towards common mycotoxins, and the potential for applying TKG and their associated microbial populations to avoid mycotoxin contamination in dairy products. Cultivation-independent high-throughput sequencing of bacterial and fungal rDNA genes indicated that Lactobacillus kefiranofaciens and Kazachstania turicensis were the most abundant bacterial and fungal taxa, respectively. In addition, 27 total isolates were obtained using cultivation methods. TKG removed more than 90% of the Ochratoxin A (OTA) after 24 h, while the isolate Kazachstania unisporus AC-2 exhibited the highest removal capacity (~46.1%). Further, the isolate exhibited good resistance to acid and bile salts environment. Analysis of the OTA detoxification mechanism revealed that both adsorption and degradation activities were exhibited by TKG, with adsorption playing a major detoxification role. Furthermore, the addition of OTA did not affect the microbial community structure of TKG. These results indicate that TKG-fermented products can naturally remove mycotoxin contamination of milk and could potentially be practically applied as probiotics in fermentation products.

An evolutionary model identifies the main evolutionary biases for the evolution of genome-replication profiles.

Recent results comparing the temporal program of genome replication of yeast species belonging to the Lachancea clade support the scenario that the evolution of the replication timing program could be mainly driven by correlated acquisition and loss events of active replication origins. Using these results as a benchmark, we develop an evolutionary model defined as birth-death process for replication origins and use it to identify the evolutionary biases that shape the replication timing profiles. Comparing different evolutionary models with data, we find that replication origin birth and death events are mainly driven by two evolutionary pressures, the first imposes that events leading to higher double-stall probability of replication forks are penalized, while the second makes less efficient origins more prone to evolutionary loss. This analysis provides an empirically grounded predictive framework for quantitative evolutionary studies of the replication timing program.

Signatures of optimal codon usage in metabolic genes inform budding yeast ecology.

Reverse ecology is the inference of ecological information from patterns of genomic variation. One rich, heretofore underutilized, source of ecologically relevant genomic information is codon optimality or adaptation. Bias toward codons that match the tRNA pool is robustly associated with high gene expression in diverse organisms, suggesting that codon optimization could be used in a reverse ecology framework to identify highly expressed, ecologically relevant genes. To test this hypothesis, we examined the relationship between optimal codon usage in the classic galactose metabolism (GAL) pathway and known ecological niches for 329 species of budding yeasts, a diverse subphylum of fungi. We find that optimal codon usage in the GAL pathway is positively correlated with quantitative growth on galactose, suggesting that GAL codon optimization reflects increased capacity to grow on galactose. Optimal codon usage in the GAL pathway is also positively correlated with human-associated ecological niches in yeasts of the CUG-Ser1 clade and with dairy-associated ecological niches in the family Saccharomycetaceae. For example, optimal codon usage of GAL genes is greater than 85% of all genes in the genome of the major human pathogen Candida albicans (CUG-Ser1 clade) and greater than 75% of genes in the genome of the dairy yeast Kluyveromyces lactis (family Saccharomycetaceae). We further find a correlation between optimization in the GALactose pathway genes and several genes associated with nutrient sensing and metabolism. This work suggests that codon optimization harbors information about the metabolic ecology of microbial eukaryotes. This information may be particularly useful for studying fungal dark matter-species that have yet to be cultured in the lab or have only been identified by genomic material.

Zygotorulaspora dagestanica sp. nov., a novel ascomycetous yeast species associated with the Georgian honeysuckle (Lonicera iberica M. Bieb.).

During an investigation of the yeast communities associated with wild fruit shrubs in Dagestan (Caucasus, Russia), four fermenting ascospore-producing yeast strains were isolated from leaves of the Georgian honeysuckle (Lonicera iberica M. Bieb.) and from soil underneath this plant. Phylogenetic analyses based on concatenated sequences of the ITS region and D1/D2 domains of the large subunit rRNA gene and concatenated sequences of the ribosomal DNA cystron, RPB2 and TEF1 genes showed that the isolated strains represented a new species of the genus Zygotorulaspora. The new species was placed in the basal position to other species of the clade and close to Zygotorulaspora mrakii. Based on the results of phylogenetic analyses and the phenotypic characteristics of the four studied strains, a novel species is described, for which the name Zygotorulaspora dagestanica sp. nov. is proposed. The holotype is KBP Y-4591T, three metabolically inactive cryopreserved isotype cultures are DSM 100088, VKM Y-3060 and VKPM Y-4318. The MycoBank number is MB 838285.

Metabarcoding analysis and fermentation performance of the dominant fungal microbiota associated with the Algerian traditional date product "Btana".

The study highlights the fungal diversity of the traditional Algerian date's product "Btana" prepared with direct Btana method (DBM) and undirect Btana method (UBM). Btana fungal populations were analyzed through 28S metabarcoding. Data treatment resulted in 122,997 reads representing three Phyla in which 76% reads (46 OTUs) belong to Ascomycota phylum. Zygosaccharomyces rouxii was the most prevailed species accounting for 35.40% of the total population. Similarity percentage analysis revealed a low level of resemblance in species in each of the two Btana types (DBM: 17.26%, UBM: 16.87). According to HPLC analysis, lactate was detected in nine samples within a range of 0.87-23.06 g/100g. Culture plating and subsequent D1/D2 domain of 28s DNA analysis showed the prevalence of Z. rouxii. Fermentation of non-renewed date medium revealed a high ethanol production (21.31 ± 2.89 g/100g) by Lachancea thermotolerans and 5.87 g/100g of lactates by Kluyveromyces delphensis. Enzymatic assay revealed a high esterase (C4) and naphtol-AS-BI-phosphohydrolase activity by L. thermotolerans, K. delphensis, and Pichia subpelliculosa, while a high level of α-fucosidase was recorded for L. thermotolerans and P. subpelliculosa. The current results demonstrated that the traditional date product Btana is a promising source for yeasts useful in production of value-added products like bioethanol and lactic acid using low-income date cultivars.