Endophytic bacterial communities and spatiotemporal variations in cotton roots in Xinjiang, China

Endophytic bacteria may be important for plant health and other ecologically relevant functions of cotton. However, the endophytic bacterial community structure and diversity in cotton is still poorly characterized. We investigated the community structure of endophytic bacteria in cotton roots growing in Xinjiang, China, using the Illumina amplicon sequencing. A total of 60.84 M effective sequences of 16S rRNA gene V3 region were obtained from cotton samples. These sequences revealed huge amount of operational taxonomic units (OTUs) in cotton, that is, 81-338 OTUs in a cotton sample, at 3% cutoff level and sequencing depth of 50000 sequences. We identified 23 classes from the resulting 2,723,384 sequences. Gammaproteobacteria were the dominant class in all cottons, followed by Alphaproteobacteria, Actinobacteria and Bacilli. A marked difference in the diversity of endophytic bacteria in cotton for different growth periods was evident. The greatest number of OTUs was detected during seedling (654 OTUs) and budding (381 OTUs). Endophytic bacteria diversity was reduced during flowering (350 OTUs) and boll-opening (351 OTUs). 217 OTUs were common to all four periods. There were more tags of Pantoea in Shihezi than other locations. While there were more tags of Erwinia in Hami than other locations. The dynamics of endophytic bacteria communities were influenced by plant growth stage. These results show the complexity of the bacterial populations present in inner tissues of cotton.

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Host identity is more important in structuring bacterial epiphytes than endophytes in a tropical mangrove forest

FEMS Microbiology Ecology ◽

10.1093/femsec/fiaa038 ◽

2020 ◽

Vol 96 (4) ◽

Author(s):

Hui Yao ◽

Xiang Sun ◽

Chao He ◽

Xing-Chun Li ◽

Liang-Dong Guo

Keyword(s):

Community Structure ◽

Endophytic Bacteria ◽

Illumina Miseq ◽

Rrna Gene ◽

Network Structures ◽

Community Structures ◽

Mangrove Species ◽

Host Identity ◽

Biodiversity Maintenance ◽

Endophytic Bacterial Community

ABSTRACT Interactions between plants and microbes are involved in biodiversity maintenance, community stability and ecosystem functioning. However, differences in the community and network structures between phyllosphere epiphytic and endophytic bacteria have rarely been investigated. Here, we examined phyllosphere epiphytic and endophytic bacterial communities of six mangrove species using Illumina MiSeq sequencing of the 16S rRNA gene. The results revealed that the community structure of epiphytic and endophytic bacteria was different. Plant identity significantly affected the diversity and community structure of both epiphytic and endophytic bacteria, with a greater effect on the community structure of the former than the latter. Network analysis showed that both plant–epiphytic and plant–endophytic bacterial network structures were characterized by significantly highly specialized and modular but lowly connected and anti-nested properties. Furthermore, the epiphytic bacterial network was more highly specialized and modular but less connected and more strongly anti-nested than the endophytic bacterial network. This study reveals that the phyllosphere epiphytic and endophytic bacterial community structures differ and plant identity has a greater effect on the epiphytic than on the endophytic bacteria, which may provide a comprehensive insight into the role of plant identity in driving the phyllosphere epiphytic and endophytic microbial community structures in mangrove ecosystems.

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Handling of spurious sequences affects the outcome of high-throughput 16S rRNA gene amplicon profiling

ISME Communications ◽

10.1038/s43705-021-00033-z ◽

2021 ◽

Vol 1 (1) ◽

Author(s):

Sandra Reitmeier ◽

Thomas C. A. Hitch ◽

Nicole Treichel ◽

Nikolaos Fikas ◽

Bela Hausmann ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Diversity Analysis ◽

Careful Attention ◽

Operational Taxonomic Units ◽

Basic Concepts ◽

Gnotobiotic Mice ◽

Mock Communities

Abstract16S rRNA gene amplicon sequencing is a popular approach for studying microbiomes. However, some basic concepts have still not been investigated comprehensively. We studied the occurrence of spurious sequences using defined microbial communities based on data either from the literature or generated in three sequencing facilities and analyzed via both operational taxonomic units (OTUs) and amplicon sequence variants (ASVs) approaches. OTU clustering and singleton removal, a commonly used approach, delivered approximately 50% (mock communities) to 80% (gnotobiotic mice) spurious taxa. The fraction of spurious taxa was generally lower based on ASV analysis, but varied depending on the gene region targeted and the barcoding system used. A relative abundance of 0.25% was found as an effective threshold below which the analysis of spurious taxa can be prevented to a large extent in both OTU- and ASV-based analysis approaches. Using this cutoff improved the reproducibility of analysis, i.e., variation in richness estimates was reduced by 38% compared with singleton filtering using six human fecal samples across seven sequencing runs. Beta-diversity analysis of human fecal communities was markedly affected by both the filtering strategy and the type of phylogenetic distances used for comparison, highlighting the importance of carefully analyzing data before drawing conclusions on microbiome changes. In summary, handling of artifact sequences during bioinformatic processing of 16S rRNA gene amplicon data requires careful attention to avoid the generation of misleading findings. We propose the concept of effective richness to facilitate the comparison of alpha-diversity across studies.

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Bacterial community associated with worker honeybees (Apis mellifera) affected by European foulbrood

PeerJ ◽

10.7717/peerj.3816 ◽

2017 ◽

Vol 5 ◽

pp. e3816 ◽

Cited By ~ 20

Author(s):

Tomas Erban ◽

Ondrej Ledvinka ◽

Martin Kamler ◽

Bronislava Hortova ◽

Marta Nesvorna ◽

...

Keyword(s):

Apis Mellifera ◽

Clinical Symptoms ◽

Clinical Signs ◽

Illumina Miseq ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Melissococcus Plutonius ◽

European Foulbrood ◽

Bacterial Groups ◽

V3 Region

BackgroundMelissococcus plutoniusis an entomopathogenic bacterium that causes European foulbrood (EFB), a honeybee (Apis melliferaL.) disease that necessitates quarantine in some countries. In Czechia, positive evidence of EFB was absent for almost 40 years, until an outbreak in the Krkonose Mountains National Park in 2015. This occurrence of EFB gave us the opportunity to study the epizootiology of EFB by focusing on the microbiome of honeybee workers, which act as vectors of honeybee diseases within and between colonies.MethodsThe study included worker bees collected from brood combs of colonies (i) with no signs of EFB (EFB0), (ii) without clinical symptoms but located at an apiary showing clinical signs of EFB (EFB1), and (iii) with clinical symptoms of EFB (EFB2). In total, 49 samples from 27 honeybee colonies were included in the dataset evaluated in this study. Each biological sample consisted of 10 surface-sterilized worker bees processed for DNA extraction. All subjects were analyzed using conventional PCR and by metabarcoding analysis based on the 16S rRNA gene V1–V3 region, as performed through Illumina MiSeq amplicon sequencing.ResultsThe bees from EFB2 colonies with clinical symptoms exhibited a 75-fold-higher incidence ofM. plutoniusthan those from EFB1 asymptomatic colonies.Melissococcus plutoniuswas identified in all EFB1 colonies as well as in some of the control colonies. The proportions ofFructobacillus fructosus,Lactobacillus kunkeei,Gilliamella apicola,Frischella perrara, andBifidobacterium coryneformewere higher in EFB2 than in EFB1, whereasLactobacillus melliswas significantly higher in EFB2 than in EFB0.Snodgrassella alviandL. melliventris,L. helsingborgensisand,L. kullabergensisexhibited higher proportion in EFB1 than in EFB2 and EFB0. The occurrence ofBartonella apisandCommensalibacter intestiniwere higher in EFB0 than in EFB2 and EFB1.Enterococcus faecalisincidence was highest in EFB2.ConclusionsHigh-throughput Illumina sequencing permitted a semi-quantitative analysis of the presence ofM. plutoniuswithin the honeybee worker microbiome. The results of this study indicate that worker bees from EFB-diseased colonies are capable of transmittingM. plutoniusdue to the greatly increased incidence of the pathogen. The presence ofM. plutoniussequences in control colonies supports the hypothesis that this pathogen exists in an enzootic state. The bacterial groups synergic to both the colonies with clinical signs of EFB and the EFB-asymptomatic colonies could be candidates for probiotics. This study confirms thatE. faecalisis a secondary invader toM. plutonius; however, other putative secondary invaders were not identified in this study.

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Gut microbiota composition is associated with narcolepsy type 1

Neurology Neuroimmunology & Neuroinflammation ◽

10.1212/nxi.0000000000000896 ◽

2020 ◽

Vol 7 (6) ◽

pp. e896

Author(s):

Alexandre Lecomte ◽

Lucie Barateau ◽

Pedro Pereira ◽

Lars Paulin ◽

Petri Auvinen ◽

...

Keyword(s):

Community Structure ◽

Gut Microbiota ◽

Bacterial Communities ◽

Beta Diversity ◽

Alpha Diversity ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Differential Abundance ◽

Alpha And Beta Diversity

ObjectiveTo test the hypothesis that narcolepsy type 1 (NT1) is related to the gut microbiota, we compared the microbiota bacterial communities of patients with NT1 and control subjects.MethodsThirty-five patients with NT1 (51.43% women, mean age 38.29 ± 19.98 years) and 41 controls (57.14% women, mean age 36.14 ± 12.68 years) were included. Stool samples were collected, and the fecal microbiota bacterial communities were compared between patients and controls using the well-standardized 16S rRNA gene amplicon sequencing approach. We studied alpha and beta diversity and differential abundance analysis between patients and controls, and between subgroups of patients with NT1.ResultsWe found no between-group differences for alpha diversity, but we discovered in NT1 a link with NT1 disease duration. We highlighted differences in the global bacterial community structure as assessed by beta diversity metrics even after adjustments for potential confounders as body mass index (BMI), often increased in NT1. Our results revealed differential abundance of several operational taxonomic units within Bacteroidetes, Bacteroides, and Flavonifractor between patients and controls, but not after adjusting for BMI.ConclusionWe provide evidence of gut microbial community structure alterations in NT1. However, further larger and longitudinal multiomics studies are required to replicate and elucidate the relationship between the gut microbiota, immunity dysregulation and NT1.

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The Wheat Microbiome Under Four Management Strategies, and Potential for Endophytes in Disease Protection

Phytobiomes Journal ◽

10.1094/pbiomes-05-17-0023-r ◽

2017 ◽

Vol 1 (3) ◽

pp. 158-168 ◽

Cited By ~ 51

Author(s):

Kristi Gdanetz ◽

Frances Trail

Keyword(s):

Bacterial Communities ◽

Management Strategy ◽

Crop Yields ◽

Management Strategies ◽

Growing Season ◽

Amplicon Sequencing ◽

Antagonistic Activity ◽

Rrna Gene ◽

Operational Taxonomic Units ◽

Plant Organ

Manipulating plant-associated microbes to reduce disease or improve crop yields requires a thorough understanding of interactions within the phytobiome. Plants were sampled from a wheat/maize/soybean crop rotation site that implements four different crop management strategies. We analyzed the fungal and bacterial communities of leaves, stems, and roots of wheat throughout the growing season using 16S and fungal internal transcribed spacer 2 rRNA gene amplicon sequencing. The most prevalent operational taxonomic units (OTUs) were shared across all samples, although levels of the low-abundance OTUs varied. Endophytes were isolated from plants, and tested for antagonistic activity toward the wheat pathogen Fusarium graminearum. Antagonistic strains were assessed for plant protective activity in seedling assays. Our results suggest that microbial communities were strongly affected by plant organ and plant age, and may be influenced by management strategy.

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Intrinsic challenges in ancient microbiome reconstruction using 16S rRNA gene amplification

Scientific Reports ◽

10.1038/srep16498 ◽

2015 ◽

Vol 5 (1) ◽

Cited By ~ 69

Author(s):

Kirsten A. Ziesemer ◽

Allison E. Mann ◽

Krithivasan Sankaranarayanan ◽

Hannes Schroeder ◽

Andrew T. Ozga ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Dna Amplification ◽

Amplicon Sequencing ◽

Universal Primers ◽

Rrna Gene ◽

Dental Calculus ◽

Shotgun Metagenomics ◽

Variable Regions ◽

V3 Region

Abstract To date, characterization of ancient oral (dental calculus) and gut (coprolite) microbiota has been primarily accomplished through a metataxonomic approach involving targeted amplification of one or more variable regions in the 16S rRNA gene. Specifically, the V3 region (E. coli 341–534) of this gene has been suggested as an excellent candidate for ancient DNA amplification and microbial community reconstruction. However, in practice this metataxonomic approach often produces highly skewed taxonomic frequency data. In this study, we use non-targeted (shotgun metagenomics) sequencing methods to better understand skewed microbial profiles observed in four ancient dental calculus specimens previously analyzed by amplicon sequencing. Through comparisons of microbial taxonomic counts from paired amplicon (V3 U341F/534R) and shotgun sequencing datasets, we demonstrate that extensive length polymorphisms in the V3 region are a consistent and major cause of differential amplification leading to taxonomic bias in ancient microbiome reconstructions based on amplicon sequencing. We conclude that systematic amplification bias confounds attempts to accurately reconstruct microbiome taxonomic profiles from 16S rRNA V3 amplicon data generated using universal primers. Because in silico analysis indicates that alternative 16S rRNA hypervariable regions will present similar challenges, we advocate for the use of a shotgun metagenomics approach in ancient microbiome reconstructions.

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Microbiota Analysis of an Environmental Slurry and Its Potential Role as a Reservoir of Bovine Digital Dermatitis Pathogens

Applied and Environmental Microbiology ◽

10.1128/aem.00244-17 ◽

2017 ◽

Vol 83 (11) ◽

Cited By ~ 3

Author(s):

Kirstine Klitgaard ◽

Mikael L. Strube ◽

Anastasia Isbrand ◽

Tim K. Jensen ◽

Martin W. Nielsen

Keyword(s):

Dairy Herd ◽

Amplicon Sequencing ◽

Dairy Herds ◽

Rrna Gene ◽

Digital Dermatitis ◽

Bacterial Populations ◽

Associated Bacteria ◽

Content Type ◽

Next Generation Sequencing Technology ◽

Route Of Transmission

ABSTRACT At present, very little information exists regarding what role the environmental slurry may play as an infection reservoir and/or route of transmission for bovine digital dermatitis (DD), a disease which is a global problem in dairy herds. To investigate whether DD-related bacteria belong to the indigenous microbiota of the dairy herd environment, we used deep amplicon sequencing of the 16S rRNA gene in 135 slurry samples collected from different sites in 22 dairy farms, with and without DD-infected cows. Both the general bacterial populations and digital dermatitis-associated Treponema were targeted in this study. The results revealed significant differences in the bacterial communities between the herds, with only 12 bacterial taxa shared across at least 80% of all the individual samples. These differences in the herd microbiota appeared to reflect mainly between-herd variation. Not surprisingly, the slurry was dominated by ubiquitous gastrointestinal bacteria, such as Ruminococcaceae and Lachnospiraceae. Despite the low relative abundance of spirochetes, which ranged from 0 to 0.6%, we were able to detect small amounts of bacterial DNA from DD-associated treponemes in the slurry. However, the DD-associated Treponema spp. were detected only in samples from herds with reported DD problems. These data indicate that treponemes involved in the pathogenesis of DD are not part of the normal environmental microflora in dairy herds without clinical DD and, consequently, that slurry is not a primary reservoir of infection. IMPORTANCE Bovine digital dermatitis (DD), a dermal disease which causes lameness in dairy cattle, is a serious problem worldwide. To control this disease, the infection reservoirs and transmission routes of DD pathogens need to be clarified. The dairy herd slurry may be a pathogen reservoir of DD-associated bacteria. The rationale for the present study was, therefore, to examine whether DD-associated bacteria are always present in slurry or if they are found only in DD-afflicted herds. The results strongly indicated that DD Treponema spp. are not part of the indigenous slurry and, therefore, do not comprise an infection reservoir in healthy herds. This study applied next-generation sequencing technology to decipher the microbial compositions of environmental slurry of dairy herds with and without digital dermatitis.

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Press xenobiotic disturbance favors deterministic assembly with a shift in function and structure of bacterial communities in sludge bioreactors

10.1101/2020.10.15.341966 ◽

2020 ◽

Author(s):

Ezequiel Santillan ◽

Hari Seshan ◽

Stefan Wuertz

Keyword(s):

Community Structure ◽

Bacterial Communities ◽

Temporal Dynamics ◽

Treatment Plant ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Batch Reactors ◽

Function Structure ◽

Community Function ◽

And Function

AbstractDisturbance is thought to affect community assembly mechanisms, which in turn shape community structure and the overall function of the ecosystem. Here, we tested the effect of a continuous (press) xenobiotic disturbance on the function, structure, and assembly of bacterial communities within a wastewater treatment system. Two sets of four-liter sequencing batch reactors were operated in triplicate with and without the addition of 3-chloroaniline for a period of 132 days, following 58 days of acclimation after inoculation with sludge from a full-scale treatment plant. Temporal dynamics of bacterial community structure were derived from 16S rRNA gene amplicon sequencing. Community function, structure and assembly differed between press disturbed and undisturbed reactors. Temporal partitioning of assembly mechanisms via phylogenetic and non-phylogenetic null modelling analysis revealed that deterministic assembly prevailed for disturbed bioreactors, while the role of stochastic assembly was stronger for undisturbed reactors. Our findings are relevant because research spanning various disturbance types, environments and spatiotemporal scales is needed for a comprehensive understanding of the effects of press disturbances on assembly mechanisms, structure, and function of microbial communities.Graphical abstract

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Bacterial diversity and community structure in nitrate-contaminated shallow groundwater in the Poyang Lake basin, China

E3S Web of Conferences ◽

10.1051/e3sconf/20199801012 ◽

2019 ◽

Vol 98 ◽

pp. 01012 ◽

Cited By ~ 1

Author(s):

Yihui Dong ◽

Jiale Li ◽

Zhanxue Sun ◽

Evgeniya Soldatova ◽

Jinjing Zan

Keyword(s):

Community Structure ◽

Bacterial Diversity ◽

Poyang Lake ◽

Shallow Groundwater ◽

Jiangxi Province ◽

Lake Basin ◽

Rrna Gene ◽

Bacterial Populations ◽

Poyang Lake Basin ◽

Groundwater Environment

The Poyang Lake basin in the Jiangxi province of China has been exposed to nitrate pollution caused by irrigation practices, leading to high groundwater nitrate concentration. Eight groundwater samples were collected from shallow wells for hydrochemical, bacterial diversity, and community structure analysis in November 2017. Shallow groundwaters of the basin are weakly acid and in an oxidizing state, with EC ranging from 87.6 to 279.5 µS/cm and TDS varying between 53 to 344 mg/L and averaging of 164 mg/L. The NO3-N form is the dominant nitrogen species in groundwater, with сoncentrations of NO3-N, NO2-N and NH4-N ranging between 2.5 to 164 mg/L, 0.01 to 0.10 mg/L, <0.01 to 0.08 mg/L, respectively. Groundwater communities are dominated by actinobacteria, alphaproteobacterial, gammaproteobacteria and betaproteobacteria both in high- and low-nitrate groundwaters. The results of a 16S rRNA gene clone library indicate that the bacterial community structure of the high-nitrate groundwater is different from that of the low-nitrate groundwater. The bacterial populations Denitratisoma and Sulfuritalea detected in low-nitrate groundwater suggest that these bacteria are capable of denitrification in anaerobic groundwater environment. Bacterial populations Flavobacteria and Cytophagia in high-nitrate groundwater are common in the s wetlands examined and likely capable of nitrification.

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Community Structure and Co-Occurrence Network Analysis of Bacteria and Fungi in Wheat Fields vs Fruit Orchards

10.21203/rs.3.rs-1142341/v1 ◽

2021 ◽

Author(s):

Xinyu Cui ◽

Huan He ◽

Fengxiao Zhu ◽

Xiaobo Liu ◽

You Ma ◽

...

Keyword(s):

Community Structure ◽

Amplicon Sequencing ◽

Yangtze River Delta ◽

Vital Role ◽

Fungal Communities ◽

Rrna Gene ◽

Network Analyses ◽

Planting Methods ◽

Bacteria And Fungi ◽

Occurrence Patterns

Abstract Soil microorganisms play a vital role in biogeochemical processes and nutrient turnover in agricultural ecosystems. However, the information on how the structure and co-occurrence patterns of microbial communities respond to the change of planting methods is still limited. In this study, a total of 34 soil samples were collected from 17 different fields of two planting types (wheat and orchards) along the Taige Canal in Yangtze River Delta. The distribution and diversity of bacterial and fungal communities in soil were determined using amplicon sequencing targeting the 16S rRNA gene and ITS gene, respectively. The dominated bacteria were Proteobacteria, Acidobacteriota, Actinobacteriota, Chloroflexi, Bacteroidota, and Firmicutes. The relative abundance of Actinobacteriota and Firmicutes was higher in the orchards, while Chloroflexi and Nitrospirota were more abundant in wheat fields. Ascomycota, Mortierellomycota, and Basidiomycota were the predominant fungi in both types of soils. The diversity of bacterial and fungal communities was greater in the wheat fields than in the orchards. The statistical analyses showed that pH was the main factor shaping the community structure. Moreover, high co-occurrence patterns of bacteria and fungi were confirmed in both wheat fields and orchards. Network analyses showed that both the wheat fields and orchards occurred modular structure, which mainly contained nodes of Acidobacteriota, Chloroflexi, Gemmatimonadota, Nitrospirota and Ascomycota. In summary, our work showed the co-occurrence network and the convergence/divergence of microbial community structure in wheat fields and orchards, giving a comprehensive understanding of the microbe-microbe interaction during planting methods changes.

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