Surveying of acid-tolerant thermophilic lignocellulolytic fungi in Vietnam reveals surprisingly high genetic diversity.

Thermophilic fungi can represent a rich source of industrially relevant enzymes. Here, 105 fungal strains capable of growing at 50 °C and pH 2.0 were isolated from compost and decaying plant matter. Maximum growth temperatures of the strains were in the range 50 °C to 60 °C. Sequencing of the internal transcribed spacer (ITS) regions indicated that 78 fungi belonged to 12 species of Ascomycota and 3 species of Zygomycota, while no fungus of Basidiomycota was detected. The remaining 27 strains could not be reliably assigned to any known species. Phylogenetically, they belonged to the genus Thielavia, but they represented 23 highly divergent genetic groups different from each other and from the closest known species by 12 to 152 nucleotides in the ITS region. Fungal secretomes of all 105 strains produced during growth on untreated rice straw were studied for lignocellulolytic activity at different pH and temperatures. The endoglucanase and xylanase activities differed substantially between the different species and strains, but in general, the enzymes produced by the novel Thielavia spp. strains exhibited both higher thermal stability and tolerance to acidic conditions. The study highlights the vast potential of an untapped diversity of thermophilic fungi in the tropics.

Moniliella floricola sp. nov., a species of black yeast isolated from flowers.

Moniliella yeasts were isolated from flower samples collected in Vietnam using an enrichment medium containing 50 % (w/w) glucose. The yeasts were identified as M. byzovii, M. dehoogii, M. megachiliensis, M. mellis, M. nigrescens and M. spathulata. A group of 20 strains representing a hitherto undescribed species of Moniliella was detected. ITS sequences indicated the presence of four genetic variants differing from each other by 4-14 nt. The strains, however, were identical in the TEF1 sequences and shared 1-2 nt differences in the D1/D2 regions. In the ITS-D1/D2 phylogenetic tree, the strains grouped together and formed a well-supported clade with insect-associated Moniliella species, including M. pollinis, M. megachiliensis and M. oedocephalis. The new group was most closely related to M. pollinis but differed from the latter by 95 nt (58 substitutions, 37 indels) in the ITS, 36 nt (31 substitutions, five indels) in the D1/D2, and 30 nt (30 substitutions) in the TEF1 sequences. Moniliella floricola sp. nov. is proposed to accommodate this group of isolates. The type strain and MycoBank number of M. floricola sp. nov. are TBY 30.1T (=CBS 12758T=NRRL Y-63660T) and MB 825274, respectively.

New insight into microbial diversity and functions in traditional Vietnamese alcoholic fermentation.

The roles of microorganisms in traditional alcoholic fermentation are often assumed based on abundance in the starter and activity in pure culture. There is a serious lack of hard evidence on the behavior and activity of individual microbial species during the actual fermentation process. In this study, microbial succession and metabolite changes during 7days of traditional Vietnamese alcoholic fermentation were monitored. Special attention was devoted to starch degradation. In total, 22 microbial species, including 6 species of filamentous fungi (Rhizopus microsporus, Rhizopus arrhizus, Mucor indicus, Mucor circinelloides, Cunninghamella elegans, Aspergillus niger), 1 yeast-like fungus (Saccharomycopsis fibuligera), 7 yeasts (Saccharomyces cerevisiae, Clavispora lusitaniae, Wickerhamomyces anomalus, Lindnera fabianii, Pichia kudriavzevii, Candida rugosa, Candida tropicalis), and 8 bacteria (Stenotrophomonas maltophilia, Lactobacillus brevis, Lactobacillus helveticus, Acinetobacter baumannii, Staphylococcus hominis, Bacillus megaterium, Enterobacter asburiae, Pediococcus pentosaceus) were identified. Despite the presence of a complex microbiota in the starter, the fermentation process is consistent and involves a limited number of functional species. Rapid change in microbial composition of fermentation mash was observed and it was correlated with ethanol content. Microbial biomass reached maximum during first 2days of solid state fermentation. Acidification of the medium took place in day 1, starch degradation in days 2, 3, 4, and alcohol accumulation from day 3. Although Sm. fibuligera dominated by cell count amongst potential starch degraders, zymography indicated that it did not produce amylase in the fermentation mash. In mixed culture with Rhizopus, amylase production by Sm. fibuligera is regulated by the moisture content of the substrate. Rhizopus was identified as the main starch degrader and S. cerevisiae as the main ethanol producer. Bacterial load was high but unstable in species composition and dominated by acid producers. M. indicus, Sm. fibuligera, W. anomalus and bacteria were regarded as satellite microorganisms. Their possible influence on organoleptic quality of fermentation product was discussed.

Moniliella byzovii sp. nov., a chlamydospore-forming black yeast isolated from flowers.

Yeasts of the genus Moniliella were isolated from 651 flower samples collected in Vietnam, using an enrichment medium containing 50 % glucose. Species of the genus Moniliella were found in 5 % of the samples and 54 strains were isolated. The strains were identified based on D1/D2 LSU rRNA gene sequences as M. megachiliensis (15 strains), M. dehoogii (14 strains), and M. mellis (2 strains). The remaining 23 strains could not be reliably placed under any known species. Among them, 12 strains isolated from flowers of Ipomoea pes-caprae and Calotropis gigantea were peculiar for the intensive formation of chlamydospores. These strains could be subdivided into pigmented and non-pigmented groups. Both groups were identical in PCR fingerprints generated with primer (GAC)5 and in D1/D2 and ITS sequences. The yeast was closely related to M. fonsecae but differed from the latter by 52 nt (or 10.3 % of divergence) in the D1/D2 sequence and 71 nt (or 16.9 % of divergence) in the ITS sequence. The name Moniliella byzovii sp. nov. is proposed for this novel species. The type strain is TBY 2041.7(T) = CBS 12757(T) = NRRL Y-63661(T). The MycoBank number is MB 803186.

Moniliella carnis sp. nov. and Moniliella dehoogii sp. nov., two novel species of black yeasts isolated from meat processing environments.

Thirteen strains of yeasts typical of the genus Moniliella were isolated from fermenting meat and meat processing tools in Vietnam. PCR fingerprints generated by primer (GAC)(5) subdivided the strains into two distinctive genetic groups. In a phylogenetic tree based on D1/D2 large subunit rRNA gene sequences, the strains formed a well-supported clade with Moniliella spathulata and Moniliella suaveolens but represented two new lineages. The names Moniliella carnis sp. nov. and Moniliella dehoogii sp. nov. are proposed. The two novel species can be distinguished from each other and from known species of Moniliella based on phenotypic characteristics. It is assumed that the yeasts were associated with fatty substances that contaminated the meat processing tools. The type strain of Moniliella carnis is KFP 246(T) ( = CBS 126447(T) = NRRL Y-48681(T)) and the type strain of Moniliella dehoogii is KFP 211(T) ( = CBS 126564(T) = NRRL Y-48682(T)).