Journal of Microbiology

Volume 56(1); January 2018

Review

[MINIREVIEW] Taxonomic hierarchy of the phylum Firmicutes and novel Firmicutes species originated from various environments in Korea: Chi Nam Seong , Joo Won Kang , Ji Hee Lee , So Yeon Seo , Jung Jae Woo , Chul Park , Kyung Sook Bae , Mi Sun Kim; J. Microbiol. 2018;56(1):1-10. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7318-x

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Abstract: This study assessed the taxonomic hierarchy of the phylum Firmicutes as well as elucidated the isolation and classification states of novel Firmicutes species isolated from Korean territory. The hierarchical classification system of the phylum Firmicutes has been developed since 1872 when the genus Bacillus was first reported and has been generally adopted since 2001. However, this taxonomic hierarchy is still being modified. Until Feb. 2017, the phylum Firmicutes consisted of seven classes (Bacilli, Clostridia, Erysipelotrichia, Limnochordia, Negativicutes, Thermolithobacteria, and Tissierellia), 13 orders, 45 families, and 421 genera. Firmicutes species isolated from various environments in Korea have been reported from 2000, and 187 species have been approved as of Feb. 2017. All Firmicutes species were affiliated with three classes (Bacilli, Clostridia, and Erysipelotrichia), four orders (Bacillales, Lactobacillales, Clostridiales, and Erysipelotrichales), 17 families, and 54 genera. A total of 173 species belong to the class Bacilli, of which 151 species were affiliated with the order Bacillales and the remaining 22 species with the order Lactobacillales. Twelve species belonging to the class Clostridia were affiliated within only one order, Clostridiales. The most abundant family was Bacillaceae (67 species), followed by the family Paenibacillaceae (56 species). Thirteen novel genera were created using isolates from the Korean environment. A number of Firmicutes species were isolated from natural environments in Korean territory. In addition, a considerable number of species were isolated from artificial resources such as fermented foods. Most Firmicutes species, belonging to the families Bacillaceae, Planococcaceae, and Staphylococcaceae, isolated from Korean fermented foods and solar salterns were halophilic or halotolerant. Firmicutes species were isolated from the whole territory of Korea, especially large numbers from Provinces Gyeonggi, Chungnam, and Daejeon.

Journal Articles

[PROTOCOL] Structural analysis of N-/O-glycans assembled on proteins in yeasts: Eun Jung Thak , Jungho Kim , Dong-Jik Lee , Jeong Yoon Kim , Hyun Ah Kang; J. Microbiol. 2018;56(1):11-23. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7468-x

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20 Citations

Abstract: Protein glycosylation, the most universal and diverse posttranslational modification, can affect protein secretion, stability, and immunogenicity. The structures of glycans attached to proteins are quite diverse among different organisms and even within yeast species. In yeast, protein glycosylation plays key roles in the quality control of secretory proteins, and particularly in maintaining cell wall integrity. Moreover, in pathogenic yeasts, glycans assembled on cell-surface glycoproteins can mediate their interactions with host cells. Thus, a comprehensive understanding of protein glycosylation in various yeast species and defining glycan structure characteristics can provide useful information for their biotechnological and clinical implications. Yeast-specific glycans are a target for glyco-engineering; implementing human-type glycosylation pathways in yeast can aid the production of recombinant glycoproteins with therapeutic potential. The virulenceassociated glycans of pathogenic yeasts could be exploited as novel targets for antifungal agents. Nowadays, several glycomics techniques facilitate the generation of species- and strain-specific glycome profiles and the delineation of modified glycan structures in mutant and engineered yeast cells. Here, we present the protocols employed in our laboratory to investigate the N- and O-glycan chains released from purified glycoproteins or cell wall mannoproteins in several yeast species.

Baekduia soli gen. nov., sp. nov., a novel bacterium isolated from the soil of Baekdu Mountain and proposal of a novel family name, Baekduiaceae fam. nov.: Dong-Shan An , Muhammad Zubair Siddiqi , Kyoung-Ho Kim , Hong-Shan Yu , Wan-Taek Im; J. Microbiol. 2018;56(1):24-29. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7107-6

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Abstract: A taxonomic study was conducted on BR7-21T, a bacterial strain isolated from the soil of a ginseng field in Baekdu Mountain. Comparative studies of the 16S rRNA gene sequence showed that the isolate was most closely related to Conexibacter woesei DSM 14684T, Solirubrobacter pauli ATCC BAA-492T, Patulibacter minatonensis JCM 12834T, with 93.8%, 92.4%, and 91.5% sequence similarity, respectively; each genus represented a family in the order Solirubrobacterales. Strain BR7-21T was Gram-reaction positive, non-spore forming, aerobic, non-motile, and short rod-shaped. It grew well on half-strength R2A medium. The G + C content of the genomic DNA was 73.9%. It contained meso-diaminopimelic acid in the cell wall and the major menaquinones were MK-7(H4) and MK-8(H4). The major fatty acids were summarized as (C16:1 ω7c/iso-C15:0 2-OH), iso-C16:0, and C17:0 cyclo. On the basis of polyphasic evidence, it was proposed that strain BR7- 21T should be placed in a new genus and species, for which the name Baekduia soli gen. nov., sp. nov. was proposed with the type strain BR7-21T (= KCTC 22257T = LMG 24797T). The family Baekduiaceae fam. nov. is proposed to encompass the genus Baekduia gen. nov.

Larkinella roseus sp. nov., a species of the family Cytophagaceae isolated from beach soil: Jae-Bong Lee , Sumin Hong , Seung-Yeol Lee , Su-Jin Park , Kyeung Il Park , Seok-Gwan Choi , Myung Kyum Kim , Leonid N. Ten , Hee-Young Jung; J. Microbiol. 2018;56(1):30-35. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7476-x

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Abstract: The taxonomic position of bacterial strain, designated 15J16- 1T3AT, recovered from a soil sample was established using a polyphasic approach. Phylogenic analysis based on the 16S rRNA gene sequence showed that strain 15J16-1T3AT belonged to the family Cytophagaceae, phylum Bacteroidetes, and was most closely related to ‘Larkinella harenae’ 15J9-9 (95.9% similarity), Larkinella ripae 15J11-11T (95.6%), Larkinella bovis M2TB15T (94.7%), Larkinella arboricola Z0532T (93.9%), and Larkinella insperata LMG 22510T (93.5%). Cells were rod-shaped, Gram-stain-negative, aerobic, and nonmotile. The isolate grew on NA, R2A, TSA, but not on LB agar. The strain was able to grow at temperature range from 10°C to 30°C with an optimum at 25°C and pH 6–8. Menaquinone MK-7 was the predominant respiratory quinone. The major cellular fatty acids comprised C16:1 ω5c (48.6%) and C15:0 iso (24.1%). Phosphatidylethanolamine, phosphatidylserine, and an unidentified lipid were the major polar lipids. The G + C content of the genomic DNA was 49.5 mol%. Strain 15J16-1T3AT could be distinguished from its closest phylogenetic neighbors based on its phenotypic, genotypic, and chemotaxonomic features. Therefore, the isolate is considered to represent a novel species in the genus Larkinella, for which the name Larkinella roseus sp. nov. is proposed. The type strain is 15J16-1T3AT (= KCTC 52004T = JCM 31991T).

Kurthia ruminicola sp. nov., isolated from the rumen contents of a Holstein cow: Myung Kyum Kim , Eun Tae Kim , Sang Bum Kim , Ha Yeon Jeong , Beom Young Park , Sathiyaraj Srinivasan; J. Microbiol. 2018;56(1):36-41. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7285-2

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Abstract: Gram-staining-positive, motile, rod-shaped bacteria, designated as H31022T and H31024 was isolated from rumen contents of a Holstein cow. Optimum growth occurred at 25°C and pH 7.0 on R2A agar medium. Oxidase and catalase activities are positive. The 16S rRNA gene sequence (1,452 bp) of the new isolates revealed they belong to the genus Kurthia of the phylum Firmicutes. Highest gene sequence similarities were assessed to be with Kurthia massiliensis JC30T (98.4%), Kurthia senegalensis JC8ET (97.5%), and Kurthia populi 10y- 14T (97.4%). Kurthia sibirica DSM 4747T (97.3%), Kurthia zopfii NBRC 101529T (97.0%), and Kurthia gibsonii NCIMB 9758T (96.7%). DNA G + C content of strains H31022T and H31024 were 34.4% and 39.7%. Strains H31022T and H31024 has the following chemotaxonomic characteristics; the major fatty acids are iso-C15:0, iso-C14:0 and anteiso-C15; polar lipid profile contained diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), unknown aminophospholipids (APL), unknown glycolipids (GL), unknown phospholipids (PL), and unknown polar lipids (L); the major quinone is MK-7. Based on polyphasic taxonomic analysis, strains H31022T (= KCTC 33923T = JCM 19640T) and H31024 (= KCTC 33924T = JCM 19641T) identified a novel species in the genus Kurthia for which the name Kurthia ruminicola sp. nov. is proposed.

Host-specificity of symbiotic mycorrhizal fungi for enhancing seed germination, protocorm formation and seedling development of over-collected medicinal orchid, Dendrobium devonianum: Hui Huang , Xiao-Meng Zi , Hua Lin , Jiang-Yun Gao; J. Microbiol. 2018;56(1):42-48. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7225-1

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52 Citations

Abstract: All orchids maintain an obligate relationship with mycorrhizal symbionts during seed germination. In most cases, germination-enhancing fungi have been isolated from roots of mature plants for conservation and cultivation purposes. To understand the germination biology of Dendrobium devonianum, an over-collected medicinal orchid, the seeds of D. devonianum were inoculated with a fungal strain (FDd1) isolated from naturally occurring protocorms of D. devonianum and two other germination-enhancing fungal strains (FDaI7 and FCb4) from D. aphyllum and Cymbidium mannii, respectively. The fungal strain was isolated from five protocorms of D. devonianum and identified as a species of the genus Epulorhiza. In germination trials, treatments with all of the three fungal strains showed a significant promoting effect on seed germination and protocorm formation, compared with the control treatment (no inoculation). However, FDd1 fungal strain showed the greatest effectiveness followed by FDaI7 and FCb4. For all inoculation and control treatments, seeds developed to protocorms regardless of the presence of illumination, whereas protocorms did not develop to seedlings unless illumination was provided. The results of our manipulative experiments confirmed the hypothesis that mycorrhizae associated with orchid seedlings are highly host-specific, and the degree of specificity may be life stagespecific under in vitro conditions. The specific mycorrhizal symbionts from protocorms can enhance restoration efforts and the conservation of orchids such as D. devonianum.

Diversity of fungi associated with roots of Calanthe orchid species in Korea: Myung Soo Park , John A. Eimes , Sang Hoon Oh , Hwa Jung Suh , Seung-Yoon Oh , Seobihn Lee , Ki Hyeong Park , Hyuk Joon Kwon , Soo-Young Kim , Young Woon Lim; J. Microbiol. 2018;56(1):49-55. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7319-9

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Abstract: While symbiotic fungi play a key role in the growth of endangered Calanthe orchid species, the relationship between fungal diversity and Calanthe species remains unclear. Here, we surveyed root associated fungal diversity of six Calanthe orchid species by sequencing the internal transcribed spacer (ITS) region using 454 pyrosequencing. Our results revealed that Paraboeremia and Coprinopsis are dominant fungal genera among Calanthe species. In terms of overall relative abundance, Paraboeremia was the most common fungal genus associated with Calanthe roots, followed by Coprinopsis. Overall fungal diversity showed a significant degree of variation depending on both location and Calanthe species. In terms of number of different fungal genera detected within Calanthe species, C. discolor had the most diverse fungal community, with 10 fungal genera detected. This study will contribute toward a better understanding of those fungi that are required for successful cultivation and conservation of Korean Calanthe species.

Application of high-salinity stress for enhancing the lipid productivity of Chlorella sorokiniana HS1 in a two-phase process: Ramesh Kakarla , Jung-Woon Choi , Jin-Ho Yun , Byung-Hyuk Kim , Jina Heo , Sujin Lee , Dae-Hyun Cho , Rishiram Ramanan , Hee-Sik Kim; J. Microbiol. 2018;56(1):56-64. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7488-6

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45 Citations

Abstract: Increased lipid accumulation of algal cells as a response to environmental stress factors attracted much attention of researchers to incorporate this stress response into industrial algal cultivation process with the aim of enhancing algal lipid productivity. This study applies high-salinity stress condition to a two-phase process in which microalgal cells are initially grown in freshwater medium until late exponential phase and subsequently subjected to high-salinity condition that induces excessive lipid accumulation. Our initial experiment revealed that the concentrated culture of Chlorella sorokiniana HS1 exhibited the intense fluorescence of Nile red at the NaCl concentration of 60 g/L along with 1 g/L of supplemental bicarbonate after 48 h of induction period without significantly compromising cultural integrity. These conditions were further verified with the algal culture grown for 7 days in a 1 L bottle reactor that reached late exponential phase; a 12% increment in the lipid content of harvested biomass was observed upon inducing high lipid accumulation in the concentrated algal culture at the density of 5.0 g DW/L. Although an increase in the sum of carbohydrate and lipid contents of harvested biomass indicated that the external carbon source supplemented during the induction period increased overall carbon assimilation, a decrease in carbohydrate content suggested the potential reallocation of cellular carbon that promoted lipid droplet formation under high-salinity stress. These
results
thus emphasize that the two-phase process can be successfully implemented to enhance algal lipid productivity by incorporating high-salinity stress conditions into the pre-concentrated sedimentation ponds of industrial algal production system.

Vacuolar zinc transporter Zrc1 is required for detoxification of excess intracellular zinc in the human fungal pathogen Cryptococcus neoformans: Minsu Cho , Guanggan Hu , Mélissa Caza , Linda C. Horianopoulos , James W. Kronstad , Won Hee Jung; J. Microbiol. 2018;56(1):65-71. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7475-y

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14 Citations

Abstract: Zinc is an important transition metal in all living organisms and is required for numerous biological processes. However, excess zinc can also be toxic to cells and cause cellular stress. In the model fungus Saccharomyces cerevisiae, a vacuolar zinc transporter, Zrc1, plays important roles in the storage and detoxification of excess intracellular zinc to protect the cell. In this study, we identified an ortholog of the S. cerevisiae ZRC1 gene in the human fungal pathogen Cryptococcus neoformans. Zrc1 was localized in the vacuolar membrane in C. neoformans, and a mutant lacking ZRC1 showed significant growth defects under high-zinc conditions. These results suggested a role for Zrc1 in zinc detoxification. However, contrary to our expectation, the expression of Zrc1 was induced in cells grown in zinc-limited conditions and decreased upon the addition of zinc. These expression patterns were similar to those of Zip1, the high-affinity zinc transporter in the plasma membrane of C. neoformans. Furthermore, we used the zrc1 mutant in a murine model of cryptococcosis to examine whether a mammalian host could inhibit the survival of C. neoformans using zinc toxicity. We found that the mutant showed no difference in virulence compared with the wildtype strain. This result suggests that Zrc1-mediated zinc detoxification is not required for the virulence of C. neoformans, and imply that zinc toxicity may not be an important aspect of the host immune response to the fungus.

Functional analysis of Mpk1-mediated cell wall integrity signaling pathway in the thermotolerant methylotrophic yeast Hansenula polymorpha: Hyunah Kim , Eun Jung Thak , Ji Yoon Yeon , Min Jeong Sohn , Jin Ho Choo , Jeong-Yoon Kim , Hyun Ah Kang; J. Microbiol. 2018;56(1):72-82. Published online January 4, 2018; DOI: https://doi.org/10.1007/s12275-018-7508-6

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Abstract: Understanding the characteristics and regulation mechanisms of cell wall integrity (CWI) in yeast is important not only for basic research but also in biotechnological applications. We found significantly different CWIs in two representative strains of the thermotolerant methylotrophic yeast Hansenula polymorpha. Compared to the A16 strain (classified as Ogataea polymorpha), the DL1-L strain (classified as Ogataea parapolymorpha) has a thinner cell wall that was found to be more fragile following long-term cultivation and more sensitive to zymolyase. To gain a deeper insight into this difference, we compared the characteristics of the Mpk1pmediated CWI signaling pathway in the two strains. While a DL1-L mutant deficient in Mpk1p (mpk1Δ) showed severe growth retardation at both normal and high growth temperatures and in the presence of cell-wall disrupting agents, the A16 mpk1Δ mutant displayed only a mild defect in cell growth. Sorbitol effect on rescuing growth retardation was different in the two mpk1Δ strains, which could partly be ascribed to subtle differences in the activation of HOG pathway. Among the cell wall disruptors evaluated, only caffeine clearly increased phosphorylation of Mpk1p in DL1-L, but not in A16. A transcriptome analysis of the DL1-L strain revealed that caffeine significantly increased the expression of a subset of cell-wall related genes in an Mpk1p-dependent manner, but not the expected Rlm1-target genes. Taken together, our data support an essential role for Mpk1p in maintaining CWI in H. polymorpha, although the requirement for Mpk1p and its regulation under diverse stress conditions varies depending on the strain background.

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