Identification of keystone taxa in rhizosphere microbial communities using different methods and their effects on compounds of the host Cinnamomum migao.

Exploring keystone taxa affecting microbial community stability and host function is crucial for understanding ecosystem functions. However, identifying keystone taxa from humongous microbial communities remains challenging. We collected 344 rhizosphere and bulk soil samples from the endangered plant C. migao for 2 years consecutively. Used high-throughput sequencing 16S rDNA and ITS to obtain the composition of bacterial and fungal communities. We explored keystone taxa and the applicability and limitations of five methods (SPEC-OCCU, Zi-Pi, Subnetwork, Betweenness, and Module), as well as the impact of microbial community domain, time series, and rhizosphere boundary on the identification of keystone taxa in the communities. Our results showed that the five methods, identified abundant keystone taxa in rhizosphere and bulk soil microbial communities. However, the keystone taxa shared by the rhizosphere and bulk soil microbial communities over time decreased rapidly decrease in the five methods. Among five methods on the identification of keystone taxa in the rhizosphere community, Module identified 113 taxa, SPEC-OCCU identified 17 taxa, Betweenness identified 3 taxa, Subnetwork identified 3 taxa, and Zi-Pi identified 4 taxa. The keystone taxa are mainly conditionally rare taxa, and their ecological functions include chemoheterotrophy, aerobic chemoheterotrophy, nitrate reduction, and anaerobic photoautotrophy. The results of the random forest model and structural equation model predict that keystone taxa Mortierella and Ellin6513 may have an effects on the accumulation of 1, 4, 7, - Cycloundecatriene, 1, 5, 9, 9-tetramethyl-, Z, Z, Z-, beta-copaene, bicyclogermacrene, 1,8-Cineole in C. migao fruits, but their effects still need further evidence. Our study evidence an unstable microbial community in the bulk soil, and the definition of microbial boundary and ecologically functional affected the identification of keystone taxa in the community. Subnetwork and Module are more in line with the definition of keystone taxa in microbial ecosystems in terms of maintaining community stability and hosting function.

Rhizobacterial compositions and their relationships with soil properties and medicinal bioactive ingredients in Cinnamomum migao.

Introduction: Rhizobacterial communities and their metabolites can affect plant growth, development, and stress resistance, as well as the biosynthesis and accumulation of bioactive compounds in medicinal plants. This relationship has been well-characterized in many medicinal herbs, although much less commonly in medicinal trees. Methods: Here, we analyzed the composition and structure of Cinnamomum migao rhizobacterial communities across nine growing regions in Yunnan, Guizhou and Guangxi, China, as well as differences in soil properties and fruit bioactive compounds. Results: Results showed that the C. migao rhizobacterial communities exhibited high species richness, but location-specific differences in structure. Site-specific differences in soil properties and bioactive compounds were also observed. Furthermore, rhizobacterial community compositions were correlated with both soil properties and fruit bioactive compounds, metabolism-related functions were most common in C. migao rhizobacteria. Discussion: Several bacterial genera, including Acidothermus, Acidibacter, Bryobacter, Candidatus_Solibacter, and Acidimicrobiales, potentially promote the biosynthesis and accumulation of 1,8-cineole, cypressene, limonene, and α-terpineol, Nitrospira and Alphaproteobacteria may play an inhibitory role. Finally, our results emphasized the critical role that soil pH and nitrogen levels play in driving rhizobacterial community structure, and specific functional bacteria can also counteract with soil properties, Acidibacter and Nitrospira can affect soil pH and nitrogen effectiveness. Overall, this study provides additional insight into the complex correlation of rhizosphere microorganisms with bioactive ingredients and soil properties of medicinal plants.

Arbuscular Mycorrhizal Fungi Improve the Growth, Water Status, and Nutrient Uptake of Cinnamomum migao and the Soil Nutrient Stoichiometry under Drought Stress and Recovery.

Drought greatly influences the growth and ecological stoichiometry of plants in arid and semi-arid regions such as karst areas, where Cinnamomum migao (C. migao) is an endemic tree species that is used as a bioenergy resource. Arbuscular mycorrhizal fungi (AMF) play a key role in nutrient uptake in the soil-plant continuum, increasing plant tolerance to drought. However, few studies have examined the contribution of AMF in improving the growth of C. migao seedlings and the soil nutrient stoichiometry under drought-stress conditions. A pot experiment was conducted under natural light in a plastic greenhouse to investigate the effects of individual inoculation and Co-inoculation of AMF [Funneliformis mosseae (F. mosseae) and Claroideoglomus etunicatum (C. etunicatum)] on the growth, water status, and nutrient uptake of C. migao as well as the soil nutrient stoichiometry under well-watered (WW) and drought-stress (DS) conditions. The results showed that compared with non-AMF control (CK), AM symbiosis significantly stimulated plant growth and had higher dry mass. Mycorrhizal plants had better water status than corresponding CK plants. AMF colonization notably increased the total nitrogen and phosphorus content of C. migao seedlings compared with CK. Mycorrhizal plants had higher leaf and stem total carbon concentrations than CK. The results indicated that AM symbiosis protects C. migao seedlings against drought stress by improving growth, water status, and nutrient uptake. In general, the C. migao seedlings that formed with C. etunicatum showed the most beneficial effect on plant growth, water status, and nutrient uptake among all treatments. In the future, we should study more about the biological characteristics of each AMF in the field study to understand more ecological responses of AMF under drought stress, which can better provide meaningful guidance for afforestation projects in karst regions.

Arbuscular mycorrhizal fungi improve the growth and drought tolerance of Cinnamomum migao by enhancing physio-biochemical responses.

Drought is the main limiting factor for plant growth in karst areas with a fragile ecological environment. Cinnamomum migao H.W. Li is an endemic medicinal woody plant present in the karst areas of southwestern China, and it is endangered due to poor drought tolerance. Arbuscular mycorrhizal fungi (AMF) are known to enhance the drought tolerance of plants. However, few studies have examined the contribution of AMF in improving the drought tolerance of C. migao seedlings. Therefore, we conducted a series of experiments to determine whether a single inoculation and coinoculation of AMF (Claroideoglomus lamellosum and Claroideoglomus etunicatum) enhanced the drought tolerance of C. migao. Furthermore, we compared the effects of single inoculation and coinoculation with different inoculum sizes (20, 40, 60, and 100 g; four replicates per treatment) on mycorrhizal colonization rate, plant growth, photosynthetic parameters, antioxidant enzyme activity, and malondialdehyde (MDA) and osmoregulatory substance contents. The results showed that compared with nonmycorrhizal plants, AMF colonization significantly improved plant growing status; net photosynthetic rate; superoxide dismutase, catalase, and peroxidase activities; and soluble sugar, soluble protein, and proline contents. Furthermore, AMF colonization increased relative water content and reduced MDA content in cells. These combined cumulative effects of AMF symbiosis ultimately enhanced the drought tolerance of seedlings and were closely related to the inoculum size. With an increase in inoculum size, the growth rate and drought tolerance of plants first increased and then decreased. The damage caused by drought stress could be reduced by inoculating 40-60 g of AMF, and the effect of coinoculation was significantly better than that of single inoculation at 60 g of AMF, while the effect was opposite at 40 g of AMF. Additionally, the interaction between AMF and inoculum sizes had a significant effect on drought tolerance. In conclusion, the inoculation of the AMF (Cl. lamellosum and Cl. etunicatum) improved photosynthesis, activated antioxidant enzymes, regulated cell osmotic state, and enhanced the drought tolerance of C. migao, enabling its growth in fragile ecological environments.

A Case Study Demonstrates That the Litter of the Rare Species Cinnamomum migao Composed of Different Tissues Can Affect the Chemical Properties and Microbial Community Diversity in Topsoil.

The decomposition of litter plays an important role in the return of forest soil nutrients, as well as the growth and productivity of plants. With this study, we aimed to determine the impact of litter mulching on different tissues of Cinnamomum migao, a rare Chinese endemic species. In particular, seeds and pericarp are easily overlooked components of C. migao litter. In this study, we tested control (uncovered litter) and litter (leaf, branch, seed, and pericarp) mulching conditions and conducted a one-year litter decomposition experiment. The enzyme activities of urease enzyme (UE) and invertase enzyme (INV) were significantly improved by litter mulching. Catalase (CAT) enzyme activities in leaf, branch, and seed litter mulching were lower than in the control, whereas CAT activity in pericarp mulching was significantly higher than in the control. Although Mortierella, Cladophialophora, Acidothermus, Sphingomonas, and Burkholderia were the dominant microbes of topsoil in different mulching treatments, there were differences in the number and connectivity of microbial communities, and this change was correlated with soil organic carbon (SOC) and CAT enzyme activity. Compared with leaves and branches, seeds and pericarp as litter are also very important for nutrient return and affect topsoil microbes in C. migao forest, which may be of significance for the growth feedback of C. migao in biennial bearing.

Seed Dormancy Release and Germination Requirements of Cinnamomum migao, an Endangered and Rare Woody Plant in Southwest China.

Seed dormancy is a complex adaptive trait of plants that are influenced by several physiological and environmental factors. The endangered plant Cinnamomum migao is also known to exhibit seed dormancy and low germination, which may influence its regeneration; however, these characteristics remain unexplored. To our knowledge, this study is the first to examine the type of dormancy and improve the germination percentage of C. migao seeds. We evaluated the structure and characteristics of the embryo and endocarp of C. migao seeds as well as the effects of endogenous inhibitors. Furthermore, we assessed the effects of light, stratification, alternating temperature, and gibberellic acid 3 (GA3) on the dormancy release of these seeds. The embryo was well developed the endocarp was water-permeable, and no obvious mechanical hindrance to germination was observed. However, the endocarp and embryo contained phenols and other germination inhibitors. The seed extracts of C. migao delayed the germination of cabbage and ryegrass seeds, which indicates the presence of endogenous inhibitors. These findings suggest that C. migao seeds exhibit physiological dormancy. Light and an alternating temperature (15/20°C) did not influence germination. However, GA3 pretreatment, alternating temperatures, and warm stratification relieved dormancy. GA3 pretreatment combined with the 15°C stratification treatment was most effective in rapidly releasing the C. migao seed dormancy. Our findings may facilitate the storage and conservation of this endangered plant, which is currently underrepresented in ex situ collections.

Different Arbuscular Mycorrhizal Fungi Established by Two Inoculation Methods Improve Growth and Drought Resistance of Cinnamomum Migao Seedlings Differently.

Drought stress is one of the greatest obstacles affecting field crop productivity in arid and semi-arid regions, and its severity and frequency are expected to increase due to human-induced changes to the environment and climate. Drought has led to rocky desertification in Karst regions. Cinnamomum migao is a unique, fast-growing medicinal plant of Southwest China that only thrives in Karst regions. Arbuscular mycorrhizal fungi (AMF) symbiosis alleviates drought stress in plants; however, establishment and function of the symbiotic interaction between AMF host plant in relation to the inoculation method remain unclear. Therefore, we conducted an experiment to investigate the effects of AMF species (Glomus etunicatum and Funneliformis mosseae) and two inoculation methods (seed vs. seedling inoculation) under drought stress on C. migao seedlings, and quantified mycorrhizal colonization, AMF spore density, root vigor, relative water content, C. migao growth, antioxidant enzyme activities, and osmotic adjustment. Inoculation with AMF (G. etunicatum and F. mosseae) positively affected the growth and root vigor of Cinnamomum migao under drought stress, regardless of the inoculation method. Additionally, both AMF species markedly upregulated antioxidant enzyme activities and osmotic adjustment substances, regardless of the inoculation method. Our results showed that the collective stimulatory effect of G. etunicatum is more efficient than that of F. mosseae. AMF application could promote afforestation with C. migao to prevent rocky desertification in Karst regions where water is the greatest limiting factor on plant growth and yield.

Two Arbuscular Mycorrhizal Fungi Alleviates Drought Stress and Improves Plant Growth in Cinnamomum migao Seedlings.

Cinnamomum migao plants often face different degrees of drought in karst habitats, which can lead to plants' death, especially in the seedling stage. Widespread of arbuscular mycorrhizal (AM) fungi in karst soils have the potential to address this drought, which is a threat to C. migao seedlings. We inoculated C. migao seedlings with spores from Glomus lamellosum and Glomus etunicatum, two AM fungi widely distributed in karst soils, to observe seedling growth response after simulated drought. Our results showed that 40 g of G. lamellosum and G. etunicatum significantly promoted the growth of C. migao seedlings, 120 days after inoculation. Following a 15-day drought treatment, root colonization of the seedlings with G. lamellosum or G. etunicatum had lower the accumulation of malondialdehyde (MDA) and increased the accumulation of enzymes and osmotic substances in the seedlings. The relative water content in different organs (roots, stems, and leaves) of the drought-stressed seedlings was higher in plants with G. lamellosum or G. etunicatum than in plants without AM fungi colonization. Our results showed that inoculation with AM fungi was an effective means to improve the drought resistance of C. migao seedlings.

Effects of rhizosphere fungi on the chemical composition of fruits of the medicinal plant Cinnamomum migao endemic to southwestern China.

BACKGROUND: This study examined how rhizosphere fungi influence the accumulation of chemical components in fruits of a small population species of Cinnamomum migao. RESULTS: Ascomycota and Basidiomycota were dominant in the rhizosphere fungal community of C. migao. Pestalotiopsis and Gibellulopsis were associated with α-Terpineol and sabinene content, and Gibellulopsis was associated with crude fat and carbohydrate content. There were significant differences in rhizosphere fungal populations between watersheds, and there was no obvious change between fruiting periods. Gibberella, Ilyonectria, Micropsalliota, and Geminibasidium promoted sabinene accumulation, and Clitocybula promoted α-Terpineol accumulation. CONCLUSION: The climate-related differentiation of rhizosphere fungal communities in watershed areas is the main driver of the chemical composition of C. migao fruit. The control of the production of biologically active compounds by the rhizosphere fungal community provides new opportunities to increase the industrial and medicinal value of the fruit of C. migao.

Transcriptome analysis of Cinnamomum migao seed germination in medicinal plants of Southwest China.

BACKGROUND: Cinnamomum migao is an endangered evergreen woody plant species endemic to China. Its fruit is used as a traditional medicine by the Miao nationality of China and has a high commercial value. However, its seed germination rate is extremely low under natural and artificial conditions. As the foundation of plant propagation, seed germination involves a series of physiological, cellular, and molecular changes; however, the molecular events and systematic changes occurring during C. migao seed germination remain unclear. RESULTS: In this study, combined with the changes in physiological indexes and transcription levels, we revealed the regulation characteristics of cell structures, storage substances, and antioxidant capacity during seed germination. Electron microscopy analysis revealed that abundant smooth and full oil bodies were present in the cotyledons of the seeds. With seed germination, oil bodies and other substances gradually degraded to supply energy; this was consistent with the content of storage substances. In parallel to electron microscopy and physiological analyses, transcriptome analysis showed that 80-90 % of differentially expressed genes (DEGs) appeared after seed imbibition, reflecting important development and physiological changes. The unigenes involved in material metabolism (glycerolipid metabolism, fatty acid degradation, and starch and sucrose metabolism) and energy supply pathways (pentose phosphate pathway, glycolysis pathway, pyruvate metabolism, tricarboxylic acid cycle, and oxidative phosphorylation) were differentially expressed in the four germination stages. Among these DEGs, a small number of genes in the energy supply pathway at the initial stage of germination maintained high level of expression to maintain seed vigor and germination ability. Genes involved in lipid metabolism were firstly activated at a large scale in the LK (seed coat fissure) stage, and then genes involved in carbohydrates (CHO) metabolism were activated, which had their own species specificity. CONCLUSIONS: Our study revealed the transcriptional levels of genes and the sequence of their corresponding metabolic pathways during seed germination. The changes in cell structure and physiological indexes also confirmed these events. Our findings provide a foundation for determining the molecular mechanisms underlying seed germination.

Simulated atmospheric nitrogen deposition inhibited the leaf litter decomposition of Cinnamomum migao H. W. Li in Southwest China.

Atmospheric nitrogen (N) deposition could affect various ecological processes in forest ecosystems, including plant litter decomposition and nutrient cycling. However, the mechanism of underlying litter decomposition and nutrient cycling of Cinnamomum migao under N deposition remains unclear. Therefore, we conducted a simulated N deposition experiment including four onsite treatments to assess the effects of N input on C. migao leaf litter decomposition, nutrient release, and soil enzyme activity. The results showed that simulated N deposition significantly increased the amount of total residual mass and lignin and cellulose, decreased the decomposition rate, and suppressed net nutrient release. N input increased C, N, and P ratios as decomposition progressed, and the proportion of mass remaining was positively correlated with the proportions of lignin and cellulose remaining at the later stage of decomposition. The differences in soil enzyme activity were primarily due to enzyme type and sampling time. We conclude that simulated N deposition significantly suppressed the leaf litter decomposition of C. migao by mainly altering the chemical properties and suppressing the decomposition of the organic matter in leaf litter. Lignin might have played an important role in the loss of leaf litter biomass at the later stage of decomposition.

Selection of 29 highly informative InDel markers for human identification and paternity analysis in Chinese Han population by the SNPlex genotyping system.

The interest of forensic researchers in single nucleotide polymorphism (SNP) has been attracted because of its potential advantages, such as low mutation rates, amenable to high-throughput automated platform and the improved application in the analysis of degraded samples. In this paper, 29 highly informative insertion/deletion (InDel, a special kind of SNP) markers were selected from the dbSNP ( http://www.ncbi.nlm.nih.gov/SNP/ ) according to the given criteria. 109 unrelated Chinese Han subjects were genotyped for the 29 InDels with SNPlex genotyping system. The allele frequency data revealed that the combined power of discrimination for the 29 InDel markers was 0.999999999990867 and the combined probability of paternity exclusion (PE) was 0.9930. Sensitivity studies were performed to evaluate the flexibility of the SNPlex genotyping system on the set of 29 InDels. Highly reproducible results could be obtained with 40-100 ng genomic DNA and the proportion of total allele drop-in was significantly increased when the amount of DNA added to PCR was lower than 35 ng. These results suggested that the set of 29 InDels was useful in paternity analysis or human identification in the future.

Diversification and molecular evolution of ATOH8, a gene encoding a bHLH transcription factor.

ATOH8 is a bHLH domain transcription factor implicated in the development of the nervous system, kidney, pancreas, retina and muscle. In the present study, we collected sequence of ATOH8 orthologues from 18 vertebrate species and 24 invertebrate species. The reconstruction of ATOH8 phylogeny and sequence analysis showed that this gene underwent notable divergences during evolution. For those vertebrate species investigated, we analyzed the gene structure and regulatory elements of ATOH8. We found that the bHLH domain of vertebrate ATOH8 was highly conserved. Mammals retained some specific amino acids in contrast to the non-mammalian orthologues. Mammals also developed another potential isoform, verified by a human expressed sequence tag (EST). Comparative genomic analyses of the regulatory elements revealed a replacement of the ancestral TATA box by CpG-islands in the eutherian mammals and an evolutionary tendency for TATA box reduction in vertebrates in general. We furthermore identified the region of the effective promoter of human ATOH8 which could drive the expression of EGFP reporter in the chicken embryo. In the opossum, both the coding region and regulatory elements of ATOH8 have some special features, such as the unique extended C-terminus encoded by the third exon and absence of both CpG islands and TATA elements in the regulatory region. Our gene mapping data showed that in human, ATOH8 was hosted in one chromosome which is a fusion product of two orthologous chromosomes in non-human primates. This unique chromosomal environment of human ATOH8 probably subjects its expression to the regulation at chromosomal level. We deduce that the great interspecific differences found in both ATOH8 gene sequence and its regulatory elements might be significant for the fine regulation of its spatiotemporal expression and roles of ATOH8, thus orchestrating its function in different tissues and organisms.

[Study on the treatment turquoise using Raman spectroscopy].

Due to a variety of the enhancement and treatment turquoises discovered in gem markets, the identification of turquoise is becoming more and more difficult. By using laser Raman spectroscopy analysis, the characteristics of Raman spectra of the pressed and filled turquoises were studied. The results show that laser Raman spectroscopy is an effective technique to identify the enhancement and treatment turquoises and the natural ones, moreover, it's a non-destructive testing method. The Raman spectra of the enhancement and treatment turquoises are resulted mainly from the vibrational mode and frequency of water, hydroxyl units, PO4 tetrahedron and CH2 units. Besides, they have the characteristic Raman spectra peaks at 2,937, 2,883 and 1,451 cm(-1) which are attributed to the stretching vibration and the bending vibration of CH2, respectively. These characteristic Raman vibration bands, it will help to distinguish the natural turquoises and the treatment ones. The study provides a new train of thought on the rapid, accurate, and non-destructive identification of turquoise.

[Raman spectrum study on turquoise].

The Raman spectrum has been employed to discuss the explanation of the structure of water and the vibration of [PO4(3-)] tetrahedron of the turquoise from Hubei and Anhui province. The Raman spectra are resulted mainly from vibrations of water, hydroxyl units and [PO4(3-)] tetrahedron of turquoise. The 3 510-3 440 cm(-1) bands with the main Raman spectra peak at 3 470 cm(-1) were assigned to the upsilon(OH) stretching vibrations and the 3 290-3 070 cm(-1) bands assigned to the upsilon(H2O) stretching vibrations. The bands observed at 1 200-1 030 cm(-1) with the strong peak at 1 039 cm(-1) were assigned to upsilon3 (PO4) antisymmetric stretching vibrations. And the Raman spectra peaks at 650-540 cm(-1) and 500-410 cm(-1) were attributed to the upsilon4 and upsilon2 bending vibrations of [PO4(3-)] tetrahedron, respectively. The spectra of the turquoise from different localities are basically similar and subtly different.

Bis(1,10-phenanthroline-κN,N')(phenyl-acetato-κO)copper(II) phenyl-acetate hexa-hydrate.

In the title compound, [Cu(C(8)H(7)O(2))(C(12)H(8)N(2))(2)](C(8)H(7)O(2))·6H(2)O, the Cu atom is in a distorted square-pyramidal coordination environment. The six crystallographically independent uncoordinated water mol-ecules are inter-connected by hydrogen bonds, completing dodeca-water (H(2)O)(12) clusters which are hydrogen bonded to the carboxyl-ate groups of phenyl-acetate anions, building up one-dimensional anionic chains propagating along [100]. Between the cationic and anionic chains are hydrogen bonds from water mol-ecules to the carboxyl-ate O atoms belonging to the phenyl-acetato ligands.

Cloning and expression pattern of a spermatogenesis-related gene, BEX1, mapped to chromosome Xq22.

Through screening a human fetal brain cDNA library, a cDNA similar to the mouse Bex1 was isolated. This new gene was named brain expressed X-linked protein 1 (BEX1). Northern blot analysis revealed a 1.0 kb transcript highly expressed in brain, pancreas, testis, and ovary, with lower levels present in heart, placenta, liver, kidney, spleen, thymus, prostate, small intestine, colon (no mucus), thyroid, spinal cord, and adrenal gland. No hybridization signal was seen in lung, skeletal muscle, peripheral blood leukocyte, stomach, lymph node, trachea, and bone marrow. The BEX1 gene was localized to chromosome band Xq22 between markers between DXS990 and DXS1059 by screening Stanford radiation hybrid G3 panels. In situ hybridization of mouse testis using BEX1 as a probe detected the signal in the pachytene spermatocytes and spermatids but not in spermatogonia. Furthermore, it was not detected at 6, 9, and 12 days postpartum, was present in low amount on Days 15 and 18 and its expression increased sharply after the initiation of puberty (about 21 days) in mouse testis.