Xiao-Mei Fang

Diversity and Composition of Airborne Fungal Community Associated with Particulate Matters in Beijing during Haze and Non-haze Days

To assess the diversity and composition of airborne fungi associated with particulate matters (PMs) in Beijing, China, a total of 81 PM samples were collected, which were derived from PM2.5, PM10 fractions, and total suspended particles during haze and non-haze days. The airborne fungal community in these samples was analyzed using the Illumina Miseq platform with fungi-specific primers targeting the internal transcribed spacer 1 region of the large subunit rRNA gene. A total of 797,040 reads belonging to 1633 operational taxonomic units were observed. Of these, 1102 belonged to Ascomycota, 502 to Basidiomycota, 24 to Zygomycota, and 5 to Chytridiomycota. The dominant orders were Pleosporales (29.39%), Capnodiales (27.96%), Eurotiales (10.64%), and Hypocreales (9.01%). The dominant genera were Cladosporium, Alternaria, Fusarium, Penicillium, Sporisorium, and Aspergilus. Analysis of similarities revealed that both particulate matter sizes (R = 0.175, p = 0.001) and air quality levels (R = 0.076, p = 0.006) significantly affected the airborne fungal community composition. The relative abundance of many fungal genera was found to significantly differ among various PM types and air quality levels. Alternaria and Epicoccum were more abundant in total suspended particles samples, Aspergillus in heavy-haze days and PM2.5 samples, and Malassezia in PM2.5 samples and heavy-haze days. Canonical correspondence analysis and permutation tests showed that temperature (p < 0.01), NO2 (p < 0.01), PM10 (p < 0.01), SO2(p < 0.01), CO (p < 0.01), and relative humidity (p < 0.05) were significant factors that determine airborne fungal community composition. The results suggest that diverse airborne fungal communities are associated with particulate matters and may provide reliable data for studying the responses of human body to the increasing level of air pollution in Beijing.

Williamsia sterculiae sp. nov., isolated from a Chinese medicinal plant.

Two actinobacterial strains, CPCC 203464(T) and CPCC 203448, isolated from surface-sterilized stems of medicinal plants were subjected to a polyphasic taxonomic study. These two aerobic organisms formed pale yellow colonies on tryptic soy agar (TSA). Cells were Gram-stain-positive, non-acid-fast, non-motile, rod- or coccoid-like elements. Comparative 16S rRNA gene sequence analysis indicated that strains CPCC 203464(T) and CPCC 203448 were most closely related to the type strains of the species of the genus Williamsia. Chemotaxonomic properties such as containing meso-diaminopimelic acid in the cell wall, arabinose, galactose and ribose being the whole-cell hydrolysate sugars, phosphatidylethanolamine (PE), diphosphatidylglycerol (DPG), phosphatidylglycerol (PG) and phosphatidylinositol (PI) as the phospholipids, and C16 : 0, 10-methyl C18 : 0, C18 : 1ω9c, C16 : 1ω7c and/or iso-C15 : 0 2-OH as major fatty acids supported the affiliation of strains CPCC 203464(T) and CPCC 203448 to the genus Williamsia. The DNA-DNA hybridization values in combination with differentiating chemotaxonomic and physiological characteristics strongly suggested that these two isolates should be classified as representatives of a novel species of the genus Williamsia. The name Williamsia sterculiae sp. nov. is proposed, with strain CPCC 203464(T) ( = DSM 45741(T) = KCTC 29118(T)) as the type strain.

Surfactin derivatives from Micromonospora sp. CPCC 202787 and their anti-HIV activities

Micromonospora sp. CPCC 202787 is a strain isolated from soil, its EtOAc-soluble fraction of the extract from mycelia displayed significant inhibitory effect on HIV-1 replication in SupT1 Cells. That motivated us, and finally we obtained two new linear surfactins (1 and 2) and eight known cyclic surfactins (3− 10) (Figure 1). Surfactin, a kind of peptide consisting of seven amino acids and one 3β-hydroxy fatty acids, is not only supposed to be the effective lipopeptide biosurfactant, but also to be bioactive constituent deserving investigation due to its multiple bioactivities such as antibacterial, antiviral, antitumor and hemolytic etc.1 The linear surfactins have been reported for their surfactant property,2–4 whereas their NMR data and bioactivity apart from anti-TB5–7 are underreported. Strain CPCC 202787 was isolated from a soil sample collected in Daluo, Menghai, Jinghong of Yunnan province, China, and it was identified as Micromonospora sp. due to its identical ribosomal 16 s rRNA gene sequence to Micromonospora carbonacea (98%). Strain CPCC 202787 was cultured on Difco ISP medium 2 (0.4% yeast extract, 1.0% malt extract, 0.4% glucose and 2.0% agar power, pH 7.2) slant at 28 °C for 7 days. Then, the grown strain was inoculated in 500-ml Erlenmeyer flasks containing 100 ml of seed medium (0.5% glucose, 1% malt extract, 1% cottonseed powder, 2% soluble starch, 0.5% yeast extract, 0.05% K2HPO4, 0.5% (NH4)2SO4, 0.3% CaCO3 and 0.1% NaCl in deionized water; pH was adjusted to 7.5 before sterilization) at 28 °C on a rotary shaker (190 r.p.m.) for 72 h to prepare seed culture. Subsequently, 1 l seed culture was transferred into 25 l fermenter filled with 20 l of culture medium (same as above seed medium), and the fermentation was carried out at 28 °C for 90– 100 h with stirring at 200 r.p.m. The mycelia separated from 20 l culture by centrifugation were supersonic extracted with (CH3)2CO–H2O (v/v, 50:50) for three times, and the extract was further partitioned between EtOAc and H2O. The EtOAc-soluble fractioned was subjected to silica gel column chromatography eluted with petroleum ether− ethyl acetate (v/v, 10:1→ 7:1) to provide 22 fractions (Fr.1~Fr.22). By repeated separation on semi-preparative HPLC, compounds 1 (8.5 mg) and 2 (12.8 mg) were obtained from Fr.6–7, and compounds 3 (24.4 mg), 4 (7.5 mg), 5 (22.0 mg), 6 (7.7 mg), 7 (18.9 mg), 8 (32.6 mg), 9 (22.2 mg) and 10 (44.1 mg) were obtained from Fr.4− 5 finally. Compound 1 was a white solid with optical rotation of 1⁄2a 25 D + 7.3 (c= 0.1, CH3OH). The HRESIMS showed a [M–H] ion at m/z 1038.6736 (calcd for C52H92N7O14, 1038.6736), deducing the molecular formula of 1 to be C52H93N7O14. In the 1H NMR spectrum, seven characteristic NH doublets proton signals at δ 8.23 (1H, d, J= 7.4 Hz), 8.11 (1H, d, J= 8.2 Hz), 8.04 (1H, d, J= 8.0 Hz), 7.98 (1H, d, J= 8.0 Hz), 7.96 (1H, d, J= 7.7 Hz), 7.85 (1H, d, J= 8.6 Hz) and 7.73 (1H, d, J= 8.4 Hz) were observed, suggesting 1 had seven amino acid residues. Moreover, the large signal at δ 1.22 observed in the 1H NMR spectrum indicated the existence of a fatty acid chain in the molecular. The 13C NMR spectrum presented 10 carbonyl signals at δ 173.9, 173.8, 172.2, 172.0, 171.7, 171.6, 171.3, 171.2, 170.9 and 170.0. The above characteristic NMR data suggested 1 was a surfactin derivative. On the basis of combined analysis of 1H–1H COSY, TOCSY, HSQC and HMBC spectra (Figure 2), seven amino acid units consisting of one glutamic acid (Glu), one asparagine (Asp), one valine (Val) and four leucines (Leu) were identified. The sequence of the amino acid units was assigned by the ROESY correlations of δ 7.98 (Glu1-NH)/3.79 (fatty acid-C-3), 7.96 (Leu2-NH)/4.27 (Glu1-α-H), 8.04 (Leu3-NH)/4.26 (Leu2-α-H), 7.85 (Val4-NH)/4.34 (Leu3-α-H), 8.23 (Asp5-NH)/4.16 (Val4-α-H), 7.73 (Leu6-NH)/4.56 (Asp5-α-H), 8.11 (Leu7-NH)/4.34 (Leu6-α-H) (Figure 2), which was also supported by the observed HMBC correlations of δ 7.98 (Glu1-NH)/171.3 (fatty acid-C-3), 7.96 (Leu2-NH)/171.2 (Glu1-C=O), 8.04 (Leu3-NH)/172.0 (Leu2-C=O), 7.85 (Val4NH)/172.2 (Leu3-C=O), 8.23 (Asp5-NH)/170.9 (Val4-C=O), 7.73 (Leu6-NH)/170.0 (Asp5-C=O), 8.11 (Leu7-NH)/171.7 (Leu6-C=O). The molecular formula of 1 proved that a hydroxyl fatty acid chain length of 14 carbons were in the molecular, and the characteristic carbon signals at δ 38.5 (fatty acid-C-11), 27.4 (fatty acid-C-12), 22.5 (fatty acid-C-13) and 22.5 (fatty acid-C-14) suggested the iso-pattern of its terminal methyl.8 Consequently, 1 was elucidated as (CH3)2CH (CH2)8CH(OH) CH2CO-Glu-Leu-Leu-Val-Asp-Leu-Leu, a new [Leu7] linear surfactin with a iso-fatty acid chain length of 14 carbons and the fully assignment of NMR data were presented in Table 1.

Glycomyces paridis sp. nov., isolated from the medicinal plant Paris polyphylla

Three actinomycete strains originating from the surface-sterilized roots of Paris polyphylla were characterized by using a polyphasic approach. Phylogenetic analyses based on the 16S rRNA gene sequence showed that they formed a deep, monophyletic branch in the genus Glycomyces, and were most closely related to the type strains of the species Glycomyces harbinensis and Glycomycesscopariae. Morphological and chemotaxonomic data supported the affiliation of strains CPCC 204357T, CPCC 204354 and CPCC 204355 to the genus Glycomyces. The results of physiological and biochemical tests allowed phenotypic differentiation of strains CPCC 204357T, CPCC 204354 and CPCC 204355 from their closest phylogenetic related species in the genus Glycomyces. Low levels of DNA-DNA relatedness with its closest type strains of G. harbinensis and G. scopariaeindicated that strain CPCC 204357T represent a novel species, for which the name Glycomyces paridis sp. nov. is proposed, with CPCC 204357T (=DSM 102295T=KCTC 39745T) as the type strain.

Metabolites from the Plant Endophytic Fungus Aspergillus sp. CPCC 400735 and Their Anti-HIV Activities

Thirty-three metabolites including five phenalenone derivatives (1-5), seven cytochalasins (6-12), thirteen butenolides (13-25), and eight phenyl derivatives (26-33) were isolated from Aspergillus sp. CPCC 400735 cultured on rice. The structures of all compounds were elucidated by NMR, MS, and CD experiments, of which 1-5 (asperphenalenones A-E), 6 (aspochalasin R), and 13 (aspulvinone R) were identified as new compounds. Specifically, asperphenalenones A-E (1-5) represent an unusual structure composed of a linear diterpene derivative linked to a phenalenone derivative via a C-C bond. Compounds 1, 4, 10, and 26 exhibited anti-HIV activity with IC50 values of 4.5, 2.4, 9.2, and 6.6 μM, respectively (lamivudine 0.1 μM; efavirenz, 0.4 × 10-3 μM).

Paenibacillus eucommiae sp. nov., isolated from a traditional Chinese medicinal herbal plant Eucommia ulmoides Oliver.

The taxonomic status of a novel bacterium, designated strain CPCC 100226T, isolated from a traditional Chinese medicinal herbal plant, Eucommia ulmoides Oliver, was characterized by using a polyphasic approach. The aerobic isolate formed pale white colonies on tryptic soy agar. Cells were Gram-stain-positive, rod-shaped, motile and endospore-forming. Chemotaxonomic investigations revealed the presence of meso-diaminopimelic acid as the diagnostic diamino acid, MK-7 as the predominant menaquinone, anteiso-C15 : 0, iso-C15 : 0, iso-C16 : 0 and C16 : 0 as the major fatty acids, and the strain had a phospholipid pattern of phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and unidentified aminophospholipids. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that the isolate was closely related to Paenibacillus aestuarii DSM 23861T with 95.1 % similarity. The G+C content of the genomic DNA was 47.9 mol%. On the basis of the genotypic and phenotypic data, the isolate is considered to represent a novel species of the genus Paenibacillus. The name proposed for this taxon is Paenibacillus eucommiae sp. nov. with CPCC 100226T (=DSM 26048T=KCTC 33054T) as the type strain.

Ornithinimicrobium flavum sp. nov., isolated from the leaf of Paris polyphylla.

A Gram-positive bacterium originating from the surface-sterilized leaf of Paris polyphylla var. yunnanensis (Franch.) was characterized by using a polyphasic approach. The isolate formed yellow, smooth, circular colonies on nutrient agar with 0.2 % starch (NSA). Cells were non-motile, non-sporulating, irregular rods or cocci. Strain CPCC 203535T had the highest 16S rRNA gene sequence similarity to the type strain of Ornithinimicrobium kibberense (96.9 %) and formed the deepest branch in the genus Ornithinimicrobium in the neighbour-joining (NJ) phylogenetic tree based on 16S rRNA gene sequences. The major menaquinones of strain CPCC 203535T were MK-8(H4), MK-8(H2) and MK-8. The peptidoglycan contained ornithine as the diagnostic diamino acid. The polar lipid profile consisted of diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylinositol (PI) and unknown lipid (UL). The major fatty acids iso-C14 : 0, iso-C15 : 0, iso-C16 : 0 and anteiso-C15 : 0 were consistent with the fatty acid patterns reported for members of the genus Ornithinimicrobium. The DNA G+C composition is 71.4 mol%. The results of physiological and biochemical tests allowed phenotypic differentiation of strain CPCC 203535T from its closest phylogenetic species in the genus Ornithinimicrobium. Strain CPCC 203535T represents a novel species of the genus Ornithinimicrobium, for which the name Ornithinimicrobium flavum sp. nov. is proposed, with CPCC 203535T (=NBRC 109452 T=KCTC 29164T) as the type strain.

Roseomonas globiformis sp. nov., an airborne bacteria isolated from an urban area of Beijing

A novel dark pink pigmented bacterium, designated strain CPCC 100847T (deposited with strain code 0113-15), was isolated from the urban air of Beijing, China. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain CPCC 100847T was related to members of the genus Roseomonas and had the highest 16S rRNA gene sequence similarity to Roseomonas aestuarii JC17T (97.5 %). A low level of DNA-DNA relatedness (18.7 %) with its closest type strain R. aestuarii JC17T (KCTC 22692T) proved that strain CPCC 100847T belonged to a unique genomic species. CPCC 100847T had many common characteristics of the genus Roseomonas, but also had a range of cultural, physiological and biochemical characteristics that separated it from related Roseomonas species. Cells were Gram-negative, cocci- to oval-shaped, non-motile, non-endospore-forming and strictly aerobic. The respiratory ubiquinone was Q-10. The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, an unidentified aminolipid and an unidentified phospholipid. The major fatty acids (>5 %) were C18 : 1ω7c, anteiso-C15 : 0, C16 : 0, iso-C15 : 0 and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The combined genotypic and phenotypic data indicated that the isolate represents a novel species of the genus Roseomonas. The name proposed for this species is Roseomonasglobiformis sp. nov., with CPCC 100847T (=KCTC 52094T) as the type strain. The DNA G+C composition is 65.2 mol%.

Structural Variation in the Bacterial Community Associated with Airborne Particulate Matter in Beijing, China, during Hazy and Nonhazy Days

ABSTRACT The structural variation of the bacterial community associated with particulate matter (PM) was assessed in an urban area of Beijing during hazy and nonhazy days. Sampling for different PM fractions (PM2.5 [<2.5 μm], PM10 [<10 μm], and total suspended particulate) was conducted using three portable air samplers from September 2014 to February 2015. The airborne bacterial community in these samples was analyzed using the Illumina MiSeq platform with bacterium-specific primers targeting the 16S rRNA gene. A total of 1,707,072 reads belonging to 6,009 operational taxonomic units were observed. The airborne bacterial community composition was significantly affected by PM fractions (R = 0.157, P < 0.01). In addition, the relative abundances of several genera significantly differed between samples with various haze levels; for example, Methylobacillus, Tumebacillus, and Desulfurispora spp. increased in heavy-haze days. Canonical correspondence analysis and permutation tests showed that temperature, SO2 concentration, relative humidity, PM10 concentration, and CO concentration were significant factors that associated with airborne bacterial community composition. Only six genera increased across PM10 samples (Dokdonella, Caenimonas, Geminicoccus, and Sphingopyxis) and PM2.5 samples (Cellulomonas and Rhizobacter), while a large number of taxa significantly increased in total suspended particulate samples, such as Paracoccus, Kocuria, and Sphingomonas. Network analysis indicated that Paracoccus, Rubellimicrobium, Kocuria, and Arthrobacter were the key genera in the airborne PM samples. Overall, the findings presented here suggest that diverse airborne bacterial communities are associated with PM and provide further understanding of bacterial community structure in the atmosphere during hazy and nonhazy days. IMPORTANCE The results presented here represent an analysis of the airborne bacterial community associated with particulate matter (PM) and advance our understanding of the structural variation of these communities. We observed a shift in bacterial community composition with PM fractions but no significant difference with haze levels. This may be because the bacterial differences are obscured by high bacterial diversity in the atmosphere. However, we also observed that a few genera (such as Methylobacillus, Tumebacillus, and Desulfurispora) increased significantly on heavy-haze days. In addition, Paracoccus, Rubellimicrobium, Kocuria, and Arthrobacter were the key genera in the airborne PM samples. Accurate and real-time techniques, such as metagenomics and metatranscriptomics, should be developed for a future survey of the relationship of airborne bacteria and haze.