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The best practice for microbiome analysis using R

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Protein & Cell 2023年第10期14卷 713-725页

作者： Tao Wen Guoqing Niu Tong Chen Qirong shen Jun Yuan Yong-Xin Liu shenzhen Branch Guangdong Laboratory of Lingnan Modern AgricultureGenome Analysis Laboratory of the Ministry of Agriculture and Rural AffairsAgricultural Genomics Institute at ShenzhenChinese Academy of Agricultural SciencesShenzhen 518120China The Key Laboratory of Plant Immunity Jiangsu Provincial Key Lab for Organic Solid Waste Utilization Jiangsu Collaborative Innovation Center for Solid Organic Waste Resource Utilization National Engineering Research Center for Organic-based FertilizersNanjing Agricultural UniversityNanjing 210095China National Resource Center for Chinese Materia Medica China Academy of Chinese Medical SciencesBeijing 100700China

With the gradual maturity of sequencing technology,many microbiome studies have published,driving the emergence and advance of related analysis tools.R language is the widely used platform for microbiome data analysis for powerful functions.However,tens of thousands of R packages and numerous similar analysis tools have brought major challenges for many researchers to explore microbiome data.How to choose suitable,efficient,convenient,and easy-to-learn tools from the numerous R packages has become a problem for many microbiome researchers.We have organized 324 common R packages for microbiome analysis and classified them according to application categories(diversity,difference,biomarker,correlation and network,functional prediction,and others),which could help researchers quickly find relevant R packages for microbiome analysis.Furthermore,we systematically sorted the integrated R packages(phyloseq,microbiome,MicrobiomeAnalystR,Animalcules,microeco,and amplicon)for microbiome analysis,and summarized the advantages and limitations,which will help researchers choose the appropriate tools.Finally,we thoroughly reviewed the R packages for microbiome analysis,summarized most of the common analysis content in the microbiome,and formed the most suitable pipeline for microbiome analysis.This paper is accompanied by hundreds of examples with 10,000 lines codes in GitHub,which can help beginners to learn,also help analysts compare and test different tools.This paper systematically sorts the application of R in microbiome,providing an important theoretical basis and practical reference for the development of better microbiome tools in the future.All the code is available at GitHub ***/taowenmicro/EasyMicrobiomeR.

关键词： R package microbiome data analysis visualization amplicon metagenome

Enrichment of beneficial cucumber rhizosphere microbes mediated by organic acid secretion

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Horticulture Research 2020年第1期7卷 662-674页

作者： Tao Wen Jun Yuan Xiaoming He Yue Lin Qiwei Huang Qirong shen Jiangsu Provincial Key Lab for Organic Solid Waste Utilization Key Laboratory of Plant ImmunityJiangsu Collaborative Innovation Center for Solid Organic WastesEducational Ministry Engineering Center of Resource-saving FertilizersNanjing Agricultural University210095 NanjingChina Vegetable Research Institute Guangdong Academy of Agricultural SciencesGuangzhou510640 GuangdongChina

Resistant cultivars have played important roles in controlling Fusarium wilt disease,but the roles of rhizosphere interactions among different levels of resistant cultivars are still unknown.Here,two phenotypes of cucumber,one resistant and one with increased susceptibility to Fusarium oxysporum f.***.cucumerinum(Foc),were grown in the soil and hydroponically,and then 16S rRNA gene sequencing and nontargeted metabolomics techniques were used to investigate rhizosphere microflora and root exudate profiles.Relatively high microbial community evenness for the Foc-susceptible cultivar was detected,and the relative abundances of Comamonadaceae and Xanthomonadaceae were higher for the Foc-susceptible cultivar than for the other cultivar.FishTaco analysis revealed that specific functional traits,such as protein synthesis and secretion,bacterial chemotaxis,and small organic acid metabolism pathways,were significantly upregulated in the rhizobacterial community of the Foc-susceptible cultivar.A machinelearning approach in conjunction with FishTaco plus metabolic pathway analysis revealed that four organic acids(citric acid,pyruvate acid,succinic acid,and fumarate)were released at higher abundance by the Foc-susceptible cultivar compared with the resistant cultivar,which may be responsible for the recruitment of Comamonadaceae,a potential beneficial microbial group.Further validation demonstrated that Comamonadaceae can be“cultured”by these organic acids.Together,compared with the resistant cultivar,the susceptible cucumber tends to assemble beneficial microbes by secreting more organic acids.

关键词： group susceptibility synthesis

Effect of bacterial intra-species community interactions on the production and activity of volatile organic compounds

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Soil Ecology Letters 2021年第1期3卷 32-41页

作者： Jianing Wang Xinlan Mei Zhong Wei Waseem Raza Qirong shen Jiangsu Provincial Key Lab for Organic Solid Waste Utilization National Engineering Research Center for Organic-based FertilizersJiangsu Collaborative Innovation Center for Solid Organic Waste Resource UtilizationNanjing Agricultural UniversityNanjing 210095China

Microorganisms experience intra-and inter-species interactions in the soil,and how these interactions affect the production of microbial volatile organic compounds(VOCs)is still not well-known.Here we evaluated the production and activity of microbial VOCs as driven by bacterial intra-species community interactions.We set up bacterial communities of increasing biodiversity out of 1–4 strains each of the Gram-positive Bacillus and Gram-negative Pseudomonas genera.We evaluated the ability of each community to provide two VOCmediated services,pathogen suppression and plant-growth promotion and then correlated these services to the production of VOCs by each community.The results showed that an increase in community richness from 1 to 4 strains of both genera increased VOC-mediated pathogen suppression and plant-growth promotion on agar medium and in the soil,which was positively correlated with the production of pathogen suppressing and plant growth-promoting VOCs.Pseudomonas strains maintained while Bacillus strains reduced community productivity with an increase in community richness and produced eight novel VOCs compared with the monocultures.These results revealed that intra-species interactions may vary between Gram-negative and Gram-positive species but improved VOC-mediated functioning with respect to pathogen suppression and plant-growth promotion by affecting the amount and diversity of produced VOCs potentially affecting plant disease outcomes.

关键词： Biodiversity Community richness Ecosystem functioning Intra-species interactions Plant-microbe interactions

Microbial diversity assembled from series-diluted suspensions of disease-suppressive soil determines pathogen invasion resistance

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Pedosphere 2021年第1期31卷 221-225页

作者： Yannan OU Zongzhuan shen Beibei WANG Menghui DONG Cece QIAO Rong LI Qirong shen liangru Provincial Key Lab jor Solid Organic Wase Uilianion Nariomal Engineering Research Ceouer fow Orpanie based FernilisersJiangsne Collaborarive Ionovantion Cener for Solid Organie Wase Resource Uiliaion.Nnpjing Agriculural Univeniy.Naning 210095Chine Hainan key Inahoratory for Sastainable Uilisatiomn of Tropical Blo-reources Colleger of Tropical Crops Hainen Universdry.Haikou 570228China

Dear Editor,Soil microbial biodiversity loss caused by agricultural intensification, climate change, and the application of chemical fertilizer has become a serious problem that threatens humans (Wall et al., 2015). One phenomenon responsible for economic and food security issues is soil-borne diseases(Fisher et al., 2012), which were reported to be associated with microbial diversity loss (shen et al., 2013;Fu et al.,2017).

关键词： soil diluted resistance

Root exudates mediate plant defense against foliar pathogens by recruiting beneficial microbes

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Soil Ecology Letters 2021年第1期3卷 42-51页

作者： Tao Wen Mengli Zhao Jun Yuan George A.Kowalchuk Qirong shen Jiangsu Provincial Key Lab for Organic Solid Waste Utilization National Engineering Research Center for Organic-based FertilizersJiangsu Collaborative Innovation Center for Solid Organic Waste Resource UtilizationNanjing Agricultural UniversityNanjing 210095China Ecology and Biodiversity Group Department of BiologyInstitute of Environmental BiologyUtrecht UniversityPadualaan 83584 CH UtrechtThe Netherlands

Plants are capable of releasing specific root exudates to recruit beneficial rhizosphere microbes upon foliar pathogen invasion attack,including long-chain fatty acids,amino acids,short-chain organic acids and sugars.Although long-chain fatty acids and amino acids application have been linked to soil legacy effects that improve future plant performance in the presence of the pathogen,the precise mechanisms involved are to a large extent still unknown.Here,we conditioned soils with long-chain fatty acids and amino acids application(L+A)or short-chain organic acids and sugars(S+S)to examine the direct role of such exudates on soil microbiome structure and function.The L+A treatment recruited higher abundances of Proteobacteria which were further identified as members of the genera Sphingomonas,Pseudomonas,Roseiflexus,and Flavitalea.We then isolated the enriched bacterial strains from these groups,identifying ten Pseudomonas strains that were able to help host plant to resist foliar pathogen infection.Further investigation showed that the L+A treatment resulted in growth promotion of these Pseudomonas strains.Collectively,our data suggest that long-chain fatty acids and amino acids stimulated by foliar pathogen infection can recruit specific Pseudomonas populations that can help protect the host plant or future plant generations.

关键词： Foliar pathogen resistance Pseudomonas Recruitment Root exudates

Effects of Local Nitrogen Supply on Water Uptake of Bean Plants in a Split Root System

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Journal of Integrative Plant Biology 2007年第4期49卷 472-480页

作者： Shiwei Guo Qirong shen Holger Brueckt College of Resource and Environmental Science Nanjing Agricultural University Nanjing 210095 China Institute of Plant Nutrition and Soil Science University of Kiel Kiel D-24118 Germany

To study the effects of local nitrogen supply on water and nutrient absorption, French bean （Phaseolus vulgaris L.） plants were grown in a split root system. Five treatments supplied with different nitrogen forms were compared： homogeneous nitrate （NN） and homogenous ammonium （AA） supply, spatially separated supply of nitrate and ammonium （NA）, half of the root system supplied with N-free nutrient solution, the other half with either nitrate （NO） or ammonium （AO）. The results showed that 10 d after onset of treatments, root dry matter （DM） in the nitratesupplied vessels treated with NA was more than two times higher than that in the ammonium-supplied vessels. Water uptake from the nitrate-supplied vessels treated with NA was 281% higher than under ammonium supply. In treatments NO and AO, the local supply of N resulted in clearly higher root DM, and water uptake from the nitratesupplied vessels was 82% higher than in the -N vessels. However, in AO plants, water uptake from the -N nutrient solution was 129% higher than from the ammonium-supplied vessels. This indicates a compensatory effect, which resulted in almost identical rates of total water uptake of treatments AA and AO, which had comparable shoot DM and leaf area. Ammonium supply reduced potassium and magnesium absorption. Water uptake was positively correlated with N, Mg and K uptake.

关键词： nitrogen form nutrient uptake rates Phaseolus vulgaris split root system water uptake rates.

Rhizosphere immunity: targeting the underground for sustainable plant health management

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Frontiers of Agricultural Science and Engineering 2020年第3期7卷 317-328页

作者： Zhong WEI Vill-Petri FRIMAN Thomas POMMIER Stefan GEISEN Alexandre JOUSSET Qirong shen Key Laboratory of Plant Immunity Jiangsu Key Laboratory for Organic Solid Waste UtilizationNational Engineering Research Center for Organic-Based FertilizersNanjing Agricultural UniversityNanjing 210095China Department of Biology University of YorkYorkYO105DDUK Ecologie Microbienne UMR1418French National Institute for Agricultural Research(INRA)University Lyon IF-69622 VilleurbanneFrance Laboratory of Nematology Wageningen University and Research6700ES Wageningenthe Netherlands Institute of Environmental Biology Ecology and BiodiversityUtrecht University3584CH Utrechtthe Netherlands

Managing plant health is a great challenge formodern food production and is further complicated by thelack of common ground between the many disciplinesinvolved in disease control. Here we present the concept ofrhizosphere immunity, in which plant health is consideredas an ecosystem level property emerging from networks ofinteractions between plants, microbiota and the surround-ing soil matrix. These interactions can potentially extendthe innate plant immune system to a point where therhizosphere immunity can fulfil all four core functions ofafull immune system: pathogen prevention, recognition,response and homeostasis. We suggest that consideringplant health from a meta-organism perspective will help indeveloping multidisciplinary pathogen management stra-tegies that focus on steering the whole plant-microbe-soilnetworks instead of individual components. This might beachieved by bringing together the latest discoveries inphytopathology, microbiome research, soil science andagronomy to pave the way toward more sustainable andproductive agriculture.

关键词： rthizosphere soil microbiome plant immunity microbial ecology plant health soilborme pathogens

Growth substrates alter aboveground plant microbial and metabolic properties thereby influencing insect herbivore performance

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Science China(Life Sciences) 2023年第8期66卷 1728-1741页

作者： Jun Yuan Tao Wen shengdie Yang Chao Zhang Mengli Zhao Guoqing Niu Penghao Xie Xiaoyu Liu Xinyuan Zhao Qirong shen T.Martijn Bezemer The Key Laboratory of Plant Immunity Jiangsu Provincial Key Lab for Organic Solid Waste UtilizationJiangsu Collaborative Innovation Center for Solid Organic WastesEducational Ministry Engineering Center of Resource-saving fertilizersNanjing Agricultural UniversityNanjing 210095China Institute of Biology Above-Belowground Interactions groupLeiden UniversityP.O.Box 95052300 RA LeidenThe Netherlands Department of Terrestrial Ecology Netherlands Institute of EcologyP.O.Box 506700 AB WageningenThe Netherlands

The gut microbiome of plant-eaters is affected by the food they eat,but it is currently unclear how the plant metabolome and microbiome are influenced by the substrate the plant grows in and how this subsequently impacts the feeding behavior and gut microbiomes of insect herbivores.Here,we use Plutella xylostella caterpillars and show that the larvae prefer leaves of cabbage plants growing in a vermiculite substrate to those from plants growing in conventional soil systems.From a plant metabolomics analysis,we identified 20 plant metabolites that were related to caterpillar feeding performance.In a bioassay,the effects of these plant metabolites on insects'feeding were tested.Nitrate and compounds enriched with leaves of soilless cultivation promoted the feeding of insects,while compounds enriched with leaves of plants growing in natural soil decreased feeding.Several microbial groups(e.g.,Sporolactobacillus,Haliangium)detected inside the plant correlated with caterpillar feeding performance and other microbial groups,such as Ramlibacter and Methylophilus,correlated with the gut microbiome.Our results highlight the role of growth substrates on the food metabolome and microbiome and on the feeding performance and the gut microbiome of plant feeders.It illustrates how belowground factors can influence the aboveground properties of plant-animal systems,which has important implications for plant growth and pest control.

关键词： cultivation pattern gut microbiome herbivory leaf microbiome leaf metabolome

Changes in bulk soil affect the disease-suppressive rhizosphere microbiome against Fusarium wilt disease

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Frontiers of Agricultural Science and Engineering 2020年第3期7卷 307-316页

作者： Lin FU Wu XIONG Francisco DINI-ANDREOTE Beibei WANG Chengyuan TAO Yunze RUAN Zongzhuan shen Rong LI Qirong shen Jiangsu Provincial Key Laboratory of Solid Organic Waste Uilization Jiangsu Collaborative Innovation Center of Solid Organic WastesEducation Ministry Engineering Center of Resource Saving FertilizersNanjing Agricultural UniversityNanjing 210095China School of Life Sciences Liaoning UniversityShenyang 110036China Ecology and Biodiversity Group Department of BiologyInstitute of Environmental BiologyUtrecht UniversityUtrecht3584 CHThe Netherlands Department of Plant Science The Pennsylvania State UniversityUniversity ParkPA 16802USA Huck Institutes of the Life Sciences The Pennsylvania State UniversityUniversity ParkPA 16802USA Hainan Key Laboratory for Sustainable Uilization of Tropical Bio-Resources Institute of Tropical Agriculture and ForestryHainan UniversityHaikou 570228China

Harnessing disease suppressive microbiomes constitutes a promising strategy for optimizing plant growth.However,relatively lttle information is available about the relationship between bulk and rhizosphere soil microbiomes.Here,the assembly of banana bulk soil and rhizosphere microbiomes was investigated in a mono-culture system consisting of bio-organic(BIO)and organic management practices.Applying BIO practice in newly reclaimed fields resulted in a high-efficiency biocontrol rate,thus providing a promising strategy for pre-control of Fusarium wilt disease.The soil microbiota was further characterized by MiSeq sequencing and quantitative PCR.The results indicate that disease suppression was mediated by the structure of a suppressive rhizosphere microbiome with respect to distinct community composition,diversity and abundance.Overall microbiome suppressiveness was primarily related to a particular set of enriched bacterial taxa affiliated with Pseudomonas,Terrimonas,Cupriavi-dus,Gp6,Ohtaekwangia and Duganella.Finally,struc-tural equation modeling was used to show that the changes in bulk soil bacterial community determined its induced rhizosphere bacterial community,which serves as an important and direct factor in restraining the pathogen.Collectively,this study provides an integrative approach to disentangle the biological basis of disease-suppressive microbiomes in the context of agricultural practice and soil management.

关键词： agricultural practice bulk soil disease suppression rhizosphere ecology