Characterization of yeast mutant strains for starter culture in Arabica coffee fermentation.

Arabica coffee is the most popular and best-selling type of coffee. During coffee fermentation, microorganisms are essential for the production of metabolites and volatile compounds that affect coffee flavor quality. This work aimed to study the mutation, selection, and characterization of the Wickerhamomyces anomalus strain YWP1-3 as a starter culture to enhance the flavor quality of Arabica coffee. The results revealed that six mutants could produce relatively high levels of the pectinase enzyme on pectin agar media and exhibited high activity levels, ranging from 332.35 to 415.88 U/ml in mucilage broth. Strains UV22-2, UV22-3, UV41-1 and UV32-1 displayed higher levels of amylase activity than did the wild type. The UV22-2 and UV22-3 mutants exhibited the highest pectin degradation indices of 49.22% and 45.97%, respectively, and displayed significantly enhanced growth rates in nitrogen yeast base media supplemented with various sugars; thus, these mutants were evaluated for their ability to serve as a starter for fermentation of Arabica coffee. The cupping scores of coffees derived from UV22-2 and UV22-3 were 83.5 ± 1.5 and 82.0 ± 2.14, respectively. The volatile compounds in the roasted coffee fermented by UV22-2 were analyzed by GC‒MS, which revealed higher levels of furfuryl alcohol and furfuryl acetate than did the other samples. These findings suggested that UV22-2 could be an influential starter culture for Arabica coffee fermentation.

Influence of Different Extraction Methods on the Changes in Bioactive Compound Composition and Antioxidant Properties of Solid-State Fermented Coffee Husk Extracts.

In brewing coffee, a huge amount of food waste is generated; that waste, coffee husks in particular, should be comprehensively exploited. They offer a rich source of bioactive compounds such as caffeine, chlorogenic acid, and trigonelline. The aim of this study was to investigate the effects of extraction methods on the bioactive compounds and antioxidant activity of such waste. Coffee husks in this study were fermented with S. cerevisiae based on a solid-state fermentation technique. The study method included ethanolic or water extraction with varied controllable factors, i.e., temperature (60, 100°C) and extraction technique. Bioactive contents were investigated with the Folin-Ciocalteu assay and 1H-NMR spectroscopy. The antioxidant activity was investigated with DPPH and FRAP assays. Results show that yields were the highest in the extract of fermented coffee husks at 100°C. The highest levels of bioactive contents (total trigonelline content at 3.59% and antioxidant activity at 23.35% (DPPH) and 25.9% (FRAP)) were found in the ethanolic extract of fermented coffee husks at 60°C. The bioactive content and bioactivity, including antioxidant activity, depended on different raw materials, preparation methods, and extraction conditions. This study illustrates the potential for using food waste such as coffee husks as a sustainable source of bioactive compounds or bioactive extracts.

North-to-South diversity of lipomycetaceous yeasts in soils evaluated with a cultivation-based approach from 11 locations in Japan.

To understand the species distribution, diversity, and density of lipomycetaceous yeasts in soil based on their north-to-south location in Japan, 1146 strains were isolated from soil samples at 11 locations from Hokkaido to Okinawa Prefecture and taxonomically characterized. Lipomycetaceous yeast strains were isolated efficiently from soil by selecting watery mucoid-like colonies on agar plates with nitrogen-depleted medium. Twenty-four (80%) of the 30 known species of the genus Lipomyces were isolated from the soil samples collected in Japan, including species recently proposed. Among the species isolated, L. starkeyi was the most predominant in Japan, except on Iriomote Island, Okinawa, and accounted for 60-98% of the isolated strains. Lipomyces yarrowii was the dominant species on Iriomote Island (64%). The second most dominant species were L. chichibuensis in Saitama Prefecture and L. doorenjongii from Yamaguchi to Okinawa Prefecture. The species diversity of lipomycetaceous yeasts was in Japan and the significant correlation with the latitude of the sampling sites was revealed.

Discovery of Oleaginous Yeast from Mountain Forest Soil in Thailand.

As an interesting alternative microbial platform for the sustainable synthesis of oleochemical building blocks and biofuels, oleaginous yeasts are increasing in both quantity and diversity. In this study, oleaginous yeast species from northern Thailand were discovered to add to the topology. A total of 127 yeast strains were isolated from 22 forest soil samples collected from mountainous areas. They were identified by an analysis of the D1/D2 domain of the large subunit rRNA (LSU rRNA) gene sequences to be 13 species. The most frequently isolated species were Lipomyces tetrasporus and Lipomyces starkeyi. Based on the cellular lipid content determination, 78 strains of ten yeast species, and two potential new yeast that which accumulated over 20% of dry biomass, were found to be oleaginous yeast strains. Among the oleaginous species detected, Papiliotrema terrestris and Papiliotrema flavescens have never been reported as oleaginous yeast before. In addition, none of the species in the genera Piskurozyma and Hannaella were found to be oleaginous yeast. L. tetrasporus SWU-NGP 2-5 accumulated the highest lipid content of 74.26% dry biomass, whereas Lipomyces mesembrius SWU-NGP 14-6 revealed the highest lipid quantity at 5.20 ± 0.03 g L-1. The fatty acid profiles of the selected oleaginous yeasts varied depending on the strain and suitability for biodiesel production.

New approach for screening of microorganisms from Arabica coffee processing for their ability to improve Arabica coffee flavor.

The purpose of this research was to isolate microorganisms from coffee fermentation processes and screen them for their potential to improve the flavor of Arabica coffee using a new approach that included pectin degradation ability and growth in mucilage broth. All of the studied microorganisms were isolated from 38 different samples of fresh coffee cherries, coffee mucilage and coffee pulp. A total of 262 microbial isolates were obtained and subjected to screening using pectinase screening agar medium for pectinolytic organisms. The results of the pectinase production test showed that 18 yeast isolates were found to produce pectinase that could degrade the pectin present in solid media. The sugar assimilation profiles and growth of selected strains in mucilage broth were studied. Therefore, 18 isolates from the selected yeasts were subjected to molecular identification by the use of 18S rRNA gene sequencing. The diversity of the yeast isolates was studied, and they were identified as Wickerhamomyces anomalus, Naganishia liquefaciens, Pichia kudriavzevii, Kazachstania naganishii and Kazachstania sp. Moreover, isolates SWU3YWP1-3, SWU3YSK9 and INFCY1-4 were used as a seed culture for Arabica coffee fermentation. The cupping sensory scores of the control (without yeast inoculation) and those inoculated with three isolated yeast strains that were determined by Q-Arabica Graders were 73.75, 84.75, 80.25 and 75.00, respectively. Unique flavors and aromas were detected. This is the first report of screening microorganisms from the Arabica coffee fermentation process by the combination of various properties with success in improving the quality of coffee beverage.

Fourteen novel lipomycetaceous yeast species isolated from soil in Japan and transfer of Dipodascopsis anomala to the genus Babjevia based on ascospore production phenotype.

Fourteen novel lipomycetaceous yeasts species were isolated from soil samples collected from the Hokkaido, Chiba and Okinawa prefectures of Japan. Phylogenetic analyses of the D1/D2 domains of the large subunit rRNAs and translation elongation factor 1 alpha genes (TEF1-α) revealed that five strains of two species from the soil in Furano-shi, Hokkaido were related to Dipodascopsis anomala and 29 strains representing 12 species from soils in Kamogawa-shi, Chiba and Iriomote Island, Okinawa were in the Myxozyma clade. The two species of Dipodascopsis form globose or ellipsoid ascospores in their sac-like ascus and pseudohyphae. Furthermore, these species produce ascospores in their pseudohyphae and do not produce an acicular ascus, which is common among the three species including D. anomala. Therefore, we propose transferring D. anomala to the genus Babjevia and amending Babjevia. Two novel species were described and included in the genus Babjevia: Babjevia hyphoforaminiformans sp. nov. (holotype NBRC 111233; MycoBank no. MB 829051) and Babjevia hyphasca sp. nov. (holotype NBRC 112965; MycoBank no. MB 829053). The 12 species in the Myxozyma clade produce neither ascospores nor pseudohyphae and have different characteristics in assimilating several carbon sources from each other. Thus, we propose that the novel species of Lipomyces be classified as forma asexualis (f.a.). From Kamogawa-shi, Chiba (19 strains representing five species): Lipomyces melibiosiraffinosiphilus f.a., sp. nov. (holotype NBRC 111411; MycoBank no. MB 829034), Lipomyces kiyosumicus f.a., sp. nov. (holotype NBRC 111424; MycoBank no. MB 829035), Lipomyces chibensis f.a., sp. nov. (holotype NBRC 111413; MycoBank no. MB 829036), Lipomyces kamogawensis f.a., sp. nov. (holotype NBRC 112967; MycoBank no. MB 829037), Lipomyces amatsuensis f.a., sp. nov. (holotype NBRC 111420; MycoBank no. MB 829041). From Iriomote island, Okinawa (10 strains representing seven species): Lipomyces taketomicus f.a., sp. nov. (holotype NBRC 112966; MycoBank no. MB 829042), Lipomyces yaeyamensis f.a., sp. nov. (holotype NBRC 110433; MycoBank no. MB 829050), Lipomyces iriomotensis f.a., sp. nov. (holotype NBRC 110436; MycoBank no. MB 829045), Lipomyces haiminakanus f.a., sp. nov. (holotype NBRC 110435; MycoBank no. MB 829046), Lipomyces komiensis f.a., sp. nov. (holotype NBRC 110440; MycoBank no. MB 829047), Lipomyces nakamensis f.a., sp. nov. (holotype NBRC 110434; MycoBank no. MB 829048), Lipomyces sakishimensis f.a., sp. nov. (holotype NBRC 110439; MycoBank no. MB 829049).

Development of biodegradation process for Poly(DL-lactic acid) degradation by crude enzyme produced by Actinomadura keratinilytica strain T16-1

Background: Plastic waste is a serious problem because it is difficult to degrade, thereby leading to global environment problems. Poly(lactic acid) (PLA) is a biodegradable aliphatic polyester derived from renewable resources, and it can be degraded by various enzymes produced by microorganisms. This study focused on the scale-up and evaluated the bioprocess of PLA degradation by a crude microbial enzyme produced by Actinomadura keratinilytica strain T16-1 in a 5 L stirred tank bioreactor. Results: PLA degradation after 72 h in a 5 L bioreactor by using the enzyme of the strain T16-1 under controlled pH conditions resulted in lactic acid titers (mg/L) of 16,651 mg/L and a conversion efficiency of 89% at a controlled pH of 8.0. However, the PLA degradation process inadvertently produced lactic acid as a potential inhibitor, as shown in our experiments at various concentrations of lactic acid. Therefore, the dialysis method was performed to reduce the concentration of lactic acid. The experiment with a dialysis bag achieved PLA degradation by weight loss of 99.93%, whereas the one without dialysis achieved a degradation of less than approximately 14.75%. Therefore, the dialysis method was applied to degrade a commercial PLA material (tray) with a conversion efficiency of 32%, which was 6-fold more than that without dialysis. Conclusions: This is the first report demonstrating the scale-up of PLA degradation in a 5 L bioreactor and evaluating a potential method for enhancing PLA degradation efficiency.

Diversity and fermentation products of xylose-utilizing yeasts isolated from buffalo feces in Thailand.

Twenty-eight xylose-utilizing yeast strains were isolated by enrichment culture from 11 samples of feces from the rectum of Murrah buffalo and Swamp buffalo in Thailand. On the basis of their morphological and biochemical characteristics, including sequence analysis of the D1/D2 region of the large-subunit ribosomal RNA gene (LSU rDNA), they were identified as Candida tropicalis (designated as Group I, 11 isolates), Candida parasilosis (Group II, 2 isolates), Candida mengyuniae (Group III, 2 isolates), Sporopachydermia lactativora (Group IV, 2 isolates), Geotrichum sp. (Group V, 5 isolates) and Trichosporon asahii (Group VI, 6 isolates). All isolates utilized xylose as the sole carbon source but 27 isolates could ferment xylose to ethanol (0.006-0.602 g L(-1)) and 21 isolates could ferment xylose to xylitol (0.19-22.84 g L(-1)). Candida tropicalis isolates produced the highest yield of xylitol (74.80%). Their ability to convert xylose to xylitol and ethanol ranged from 15.06 g L(-1) to 22.84 g L(-1) xylitol and 0.110 g L(-1) to 0.602 g L(-1) ethanol, respectively.

Paenibacillus cellulositrophicus sp. nov., a cellulolytic bacterium from Thai soil.

A cellulolytic bacterium, strain P2-1(T), isolated from soil in Thailand, was characterized using a taxonomic approach based on phenotypic and chemotaxonomic characteristics and the 16S rRNA gene sequence. The novel strain was Gram-positive, facultatively anaerobic, spore-forming and rod-shaped. It contained meso-diaminopimelic as the diagnostic diamino acid in the cell-wall peptidoglycan. The DNA G+C content was 52.7 mol%. The major isoprenoid quinone was MK-7. Anteiso-C(15 : 0) and iso-C(16 : 0) were the dominant cellular fatty acids. Phylogenetic analyses using the 16S rRNA gene sequence showed that the novel strain was affiliated to the genus Paenibacillus. Strain P2-1(T) was closely related to Paenibacillus cineris KCTC 3998(T), P. favisporus KCTC 3910(T) and P. rhizosphaerae KCTC 13015(T) with 96.3-96.5 % gene sequence similarity. DNA-DNA relatedness, physiological characteristics and some biochemical characteristics clearly distinguished strain P2-1(T) from related species of the genus Paenibacillus. Therefore, strain P2-1(T) represents a novel species of the genus Paenibacillus, for which the name Paenibacillus cellulositrophicus sp. nov. is proposed. The type strain is P2-1(T) (=KCTC 13135(T)=PCU 305(T)=TISTR 1888(T)).