Yunfu Gu

Culturable Heavy Metal-Resistant and Plant Growth Promoting Bacteria in V-Ti Magnetite Mine Tailing Soil from Panzhihua, China

To provide a basis for using indigenous bacteria for bioremediation of heavy metal contaminated soil, the heavy metal resistance and plant growth-promoting activity of 136 isolates from V-Ti magnetite mine tailing soil were systematically analyzed. Among the 13 identified bacterial genera, the most abundant genus was Bacillus (79 isolates) out of which 32 represented B. subtilis and 14 B. pumilus, followed by Rhizobium sp. (29 isolates) and Ochrobactrum intermedium (13 isolates). Altogether 93 isolates tolerated the highest concentration (1000 mg kg−1) of at least one of the six tested heavy metals. Five strains were tolerant against all the tested heavy metals, 71 strains tolerated 1,000 mg kg−1 cadmium whereas only one strain tolerated 1,000 mg kg−1 cobalt. Altogether 67% of the bacteria produced indoleacetic acid (IAA), a plant growth-promoting phytohormone. The concentration of IAA produced by 53 isolates was higher than 20 µg ml−1. In total 21% of the bacteria produced siderophore (5.50–167.67 µg ml−1) with two Bacillus sp. producing more than 100 µg ml−1. Eighteen isolates produced both IAA and siderophore. The results suggested that the indigenous bacteria in the soil have beneficial characteristics for remediating the contaminated mine tailing soil.

Genetic characterisation of endophytic actinobacteria isolated from the medicinal plants in Sichuan

The genetic diversity of 41 endophytic actinobacteria isolated from 13 native medicinal plants growing in diverse ecoclimatic zones in Sichuan, China, was investigated by using PCR-RFLP of the ribosomal 16S rRNA gene and the intergenic spacer (IGS) region between the 16S rRNA and 23S rRNA genes, BOX fingerprinting and full sequence analysis of the 16S rRNA gene. The results revealed 29 and 40 genotypes from 16S and 16S–23S rRNA IGS genes PCR-RFLP data, respectively, which indicated considerable genetic diversity among the isolates. Analysis of 16S rRNA gene sequences demonstrated that the strains were most closely related to representatives of the generaStreptomyces andMicromonospora among actinobacteria. Native medicinal plants are thus important resources for endophytic actinobacteria.

Determination of the fungal community of pit mud in fermentation cellars for Chinese strong-flavor liquor, using DGGE and Illumina MiSeq sequencing

Chinese strong-flavor liquor (CSFL) is fermented in cellars lined with pit mud (PM). This PM, specific fermented clay, contains microbes that play important roles in CSFL production. However, there is limited information about fungal community structure and cellar-age-related changes in PM. In this study, PM samples were removed from the cellars used for 5 and 100years and characterized using denaturing gradient gel electrophoresis (DGGE) and Illumina MiSeq sequencing. Both methods revealed there were no significant differences in fungal species diversity (Shannon index, Chao1, and observed species) between the 5- and 100-year PM samples (p>0.05), but the communities were more stable in the 100-year PM samples (UPGMA). Illumina MiSeq sequencing allowed identification of 111 fungal genera belonging to 4 phyla (Ascomycota, Zygomycota, Basidiomycota, and Chytridiomycota) in the PM samples, with the predominant phylum being Ascomycota. The results also indicated that the compositions of dominant genera in the PM samples were significantly changed during long-term CSFL fermentation. There were relatively more Rhizopus, Phoma, and Trichosporon in the 5-year PM samples, and Aspergillus and Candida were most highly represented in the 100-year PM samples (p<0.05). Of these, Candida increased its relative abundance significantly in the 100-year samples (p<0.05). Overall, the results provide novel insights into the fungal community associated with CSFL production, and may suggest why fermentation in a cellar with PM that has been in usage for a longer time allows better quality CSFL production.

An indoleacetic acid-producing Ochrobactrum sp MGJ11 counteracts cadmium effect on soybean by promoting plant growth

To analyse whether some indoleacetic acid (IAA)‐secreting plant growth‐promoting bacteria can alleviate cadmium (Cd) stress, the role of an Ochrobactrum sp. MGJ11 from rhizosphere of soybean in promoting plant growth, and to evaluate the counteracting Cd effects on soybean.

Diversity of nifH gene in rhizosphere and non-rhizosphere soil of tobacco in Panzhihua, China

To gain a better understanding of the effects of tobacco root secretions on the nifH gene community of rhizosphere soil, the diversity of the nifH gene in the rhizosphere and non-rhizosphere soil of tobacco was investigated using denaturing gradient gel electrophoresis (DGGE) and sequencing. Shannon diversity (H), richness (S) and evenness (EH) indices were used to analyze the DGGE results. The results showed that the nifH gene diversity for rhizosphere soil was lower than that of the non-rhizosphere at three sampling sites. The increase in H of non-rhizosphere soil was due mainly to the increase in pH and the decrease in available N. Richness was correlated closely with H. There was a great difference of evenness between the rhizosphere and the non-rhizosphere samples. A range of sequence divergence was observed in the eight sequenced nifH clones. The sequences were divided into three clusters in the phylogenetic tree. The majority of the clones were similar to the nifH genes of Betaproteobacteria, Zoogloea oryzae, Dechloromonas sp. and Azovibrio restrictus. There was a difference regarding dominant species between the rhizosphere and the non-rhizosphere samples.

Long-term Fertilization Structures Bacterial and Archaeal Communities along Soil Depth Gradient in a Paddy Soil

Soil microbes provide important ecosystem services. Though the effects of changes in nutrient availability due to fertilization on the soil microbial communities in the topsoil (tilled layer, 0–20 cm) have been extensively explored, the effects on communities and their associations with soil nutrients in the subsoil (below 20 cm) which is rarely impacted by tillage are still unclear. 16S rRNA gene amplicon sequencing was used to investigate bacterial and archaeal communities in a Pup-Calric-Entisol soil treated for 32 years with chemical fertilizer (CF) and CF combined with farmyard manure (CFM), and to reveal links between soil properties and specific bacterial and archaeal taxa in both the top- and subsoil. The results showed that both CF and CFM treatments increased soil organic carbon (SOC), soil moisture (MO) and total nitrogen (TN) while decreased the nitrate_N content through the profile. Fertilizer applications also increased Olsen phosphorus (OP) content in most soil layers. Microbial communities in the topsoil were significantly different from those in subsoil. Compared to the CF treatment, taxa such as Nitrososphaera, Nitrospira, and several members of Acidobacteria in topsoil and Subdivision 3 genera incertae sedis, Leptolinea, and Bellilinea in subsoil were substantially more abundant in CFM. A co-occurrence based network analysis demonstrated that SOC and OP were the most important soil parameters that positively correlated with specific bacterial and archaeal taxa in topsoil and subsoil, respectively. Hydrogenophaga was identified as the keystone genus in the topsoil, while genera Phenylobacterium and Steroidobacter were identified as the keystone taxa in subsoil. The taxa identified above are involved in the decomposition of complex organic compounds and soil carbon, nitrogen, and phosphorus transformations. This study revealed that the spatial variability of soil properties due to long-term fertilization strongly shapes the bacterial and archaeal community composition and their interactions at both high and low taxonomic levels across the whole soil profile.

Identification and evaluation of reference genes for qRT-PCR studies in Lentinula edodes

Lentinula edodes (shiitake mushroom) is a common edible mushroom with a number of potential therapeutic and nutritional applications. It contains various medically important molecules, such as polysaccharides, terpenoids, sterols, and lipids, were contained in this mushroom. Quantitative real-time polymerase chain reaction (qRT-PCR) is a powerful tool to analyze the mechanisms underlying the biosynthetic pathways of these substances. qRT-PCR is used for accurate analyses of transcript levels owing to its rapidity, sensitivity, and reliability. However, its accuracy and reliability for the quantification of transcripts rely on the expression stability of the reference genes used for data normalization. To ensure the reliability of gene expression analyses using qRT-PCR in L. edodes molecular biology research, it is necessary to systematically evaluate reference genes. In the current study, ten potential reference genes were selected from L. edodes genomic data and their expression levels were measured by qRT-PCR using various samples. The expression stability of each candidate gene was analyzed by three commonly used software packages: geNorm, NormFinder, and BestKeeper. Base on the results, Rpl4 was the most stable reference gene across all experimental conditions, and Atu was the most stable gene among strains. 18S was found to be the best reference gene for different development stages, and Rpl4 was the most stably expressed gene under various nutrient conditions. The present work will contribute to qRT-PCR studies in L. edodes.

Endophytes isolated from ginger rhizome exhibit growth promoting potential for Zea mays

ABSTRACT Plant growth promoting bacteria (PGPB) are known to colonize ginger rhizome. A total of 57 strains were isolated and identified by 16S rDNA PCR-RFLP fingerprinting, and were grouped into genera Serratia, Enterobacter, Acinetobacter, Pseudomonas, Stenotrophomonas, Agrobacterium, Ochrobactrum, Bacillus and Tetrathiobacter. Out of 34 representative strains, 14 were positive for indole-3-acetic acid (IAA) production (1.02–49.66 µg ml–1), 16 were able to solubilize phosphate (9.3–233.05 µg ml–1) and 18 showed siderophore activities (9.47–70.66%). Six representative strains with plant growth promoting (PGP) activities boosted the growth of Zea mays, where not only the plant height, leaf area, and biomass yield were significantly improved, plant N, P and K uptakes were also increased. Plants co-inoculated with both PGPB and chemical fertilizer also showed an increased yield. Our study thus indicates that there is an abundance of endophytic bacteria in ginger rhizome, some of which possess promising capabilities for enhancing the growth of Z. mays and have the potential to serve as bio-fertilizers for crops.

Isolation and antimicrobial activities of actinobacteria closely associated with liquorice plants Glycyrrhiza glabra L. and Glycyrrhiza inflate BAT. in Xinjiang, China.

A total of 218 actinobacteria strains were isolated from wild perennial liquorice plants Glycyrrhiza glabra L. and Glycyrrhiza. inflate BAT. Based on morphological characteristics, 45 and 32 strains from G. inflate and G. glabra, respectively, were selected for further analyses. According to 16S rRNA sequence analysis, most of the strains belonged to genus Streptomyces and a few strains represented the rare actinobacteria Micromonospora, Rhodococcus and Tsukamurella. A total of 39 strains from G. inflate and 27 strains from G. glabra showed antimicrobial activity against at least one indicator organism. The range of the antimicrobial activity of the strains isolated from G. glabra and G. inflate was similar. A total of 34 strains from G. inflate and 29 strains from G. glabra carried at least one of the genes encoding polyketide synthases, non-ribosomal peptide synthetase and FADH2-dependent halogenase. In the type II polyketide synthase KSα gene phylogenetic analysis, the strains were divided into two major clades: one included known spore pigment production-linked KSα sequences and other sequences were linked to the production of different types of aromatic polyketide antibiotics. Based on the antimicrobial range, the isolates that carried different KSα types were not separated from each other or from the isolates that did not carry KSα. The incongruent phylogenies of 16S rRNA and KSα genes indicated that the KSα genes were possibly horizontally transferred. In all, the liquorice plants were a rich source of biocontrol agents that may produce novel bioactive compounds.

Degradation shaped bacterial and archaeal communities with predictable taxa and their association patterns in Zoige wetland at Tibet plateau

Soil microbes provide important ecosystem services. Zoige Plateau wetland, the largest alpine peat wetland in the world, has suffered from serious degradation in the past 30 years. We studied the composition of the Zoige Plateau alpine wetland soil microbiota and relations among specific taxa using 16S rRNA amplicon sequencing combined with association network analysis. Compared to the pristine swamp soil, taxons DA101, Aeromicrobium, Bradyrhizobium, and Candidatus Nitrososphaera were enriched and several methanogenic Euryarchaeota were depleted in the moderately degraded meadow soil and highly degraded sandy soil. Soil total potassium contents in soils with different degradation levels were significantly different, being the highest in meadow soil and lowest in swamp soil. The association network analysis showed that total potassium positively correlated with specific bacterial and archaeal taxa. Jiangella, Anaerolinea, Desulfobulbus, Geobacter, Flavobacterium, Methanobacterium and Methanosaeta were identified as the keystone genera in the networks. Soil degradation affected soil properties, and caused changes in the bacterial and archaeal community composition and the association patterns of community members. The changes could serve as early warning signals of soil degradation in alpine wetlands.