scholarly journals Dilution-to-Stimulation/Extinction Method: a Combination Enrichment Strategy To Develop a Minimal and Versatile Lignocellulolytic Bacterial Consortium

2020 ◽  
Vol 87 (2) ◽  
Author(s):  
Laura Díaz-García ◽  
Sixing Huang ◽  
Cathrin Spröer ◽  
Rocío Sierra-Ramírez ◽  
Boyke Bunk ◽  
...  
Keyword(s):  
Microbial Consortium ◽  
Plant Biomass ◽  
Bacterial Species ◽  
Bacterial Consortium ◽  
Priority Effects ◽  
Plant Residues ◽  
Agricultural Plant ◽  
Enrichment Strategy ◽  
Sequence Types ◽  
Bacterial Sequence

ABSTRACT The engineering of complex communities can be a successful path to understand the ecology of microbial systems and improve biotechnological processes. Here, we developed a strategy to assemble a minimal and effective lignocellulolytic microbial consortium (MELMC) using a sequential combination of dilution-to-stimulation and dilution-to-extinction approaches. The consortium was retrieved from Andean forest soil and selected through incubation in liquid medium with a mixture of three types of agricultural plant residues. After the dilution-to-stimulation phase, approximately 50 bacterial sequence types, mostly belonging to the Sphingobacteriaceae, Enterobacteriaceae, Pseudomonadaceae, and Paenibacillaceae, were significantly enriched. The dilution-to-extinction method demonstrated that only eight of the bacterial sequence types were necessary to maintain microbial growth and plant biomass consumption. After subsequent stabilization, only two bacterial species (Pseudomonas sp. and Paenibacillus sp.) became highly abundant (>99%) within the MELMC, indicating that these are the key players in degradation. Differences in the composition of bacterial communities between biological replicates indicated that selection, sampling, and/or priority effects could shape the consortium structure. The MELMC can degrade up to ∼13% of corn stover, consuming mostly its (hemi)cellulosic fraction. Tests with chromogenic substrates showed that the MELMC secretes an array of endoenzymes able to degrade xylan, arabinoxylan, carboxymethyl cellulose, and wheat straw. Additionally, the metagenomic profile inferred from the phylogenetic composition along with an analysis of carbohydrate-active enzymes of 20 bacterial genomes support the potential of the MELMC to deconstruct plant polysaccharides. This capacity was mainly attributed to the presence of Paenibacillus sp. IMPORTANCE The significance of our study mainly lies in the development of a combined top-down enrichment strategy (i.e., dilution to stimulation coupled to dilution to extinction) to build a minimal and versatile lignocellulolytic microbial consortium. We demonstrated that mainly two selectively enriched bacterial species (Pseudomonas sp. and Paenibacillus sp.) are required to drive the effective degradation of plant polymers. Our findings can guide the design of a synthetic bacterial consortium that could improve saccharification (i.e., the release of sugars from agricultural plant residues) processes in biorefineries. In addition, they can help to expand our ecological understanding of plant biomass degradation in enriched bacterial systems.

scholarly journals Top-Down Enrichment Strategy to Co-cultivate Lactic Acid and Lignocellulolytic Bacteria From the Megathyrsus maximus Phyllosphere

2021 ◽  
Vol 12 ◽  
Author(s):  
Laura Díaz-García ◽  
Dayanne Chaparro ◽  
Hugo Jiménez ◽  
Luis Fernando Gómez-Ramírez ◽  
Adriana J. Bernal ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Plant Biomass ◽  
Selection Process ◽  
Carbon Sources ◽  
Bacterial Consortium ◽  
Top Down ◽  
Relevant Role ◽  
Ph Decrease ◽  
Silage Process ◽  
Enrichment Strategy

Traditionally, starting inoculants have been applied to improve ensiling of forage used for livestock feed. Here, we aimed to build up a bioinoculant composed of lactic acid-producing and lignocellulolytic bacteria (LB) derived from the Megathyrsus maximus (guinea grass) phyllosphere. For this, the dilution-to-stimulation approach was used, including a sequential modification of the starting culture medium [Man, Rogosa, and Sharpe (MRS) broth] by addition of plant biomass (PB) and elimination of labile carbon sources. Along 10 growth-dilution steps (T1–T10), slight differences were observed in terms of bacterial diversity and composition. After the sixth subculture, the consortium started to degrade PB, decreasing its growth rate. The co-existence of Enterobacteriales (fast growers and highly abundance), Actinomycetales, Bacillales, and Lactobacillales species was observed at the end of the selection process. However, a significant structural change was noticed when the mixed consortium was cultivated in higher volume (500ml) for 8days, mainly increasing the proportion of Paenibacillaceae populations. Interestingly, Actinomycetales, Bacillales, and Lactobacillales respond positively to a pH decrease (4–5), suggesting a relevant role within a further silage process. Moreover, gene-centric metagenomic analysis showed an increase of (hemi)cellulose-degrading enzymes (HDEs) during the enrichment strategy. Reconstruction of metagenome-assembled genomes (MAGs) revealed that Paenibacillus, Cellulosimicrobium, and Sphingomonas appear as key (hemi)cellulolytic members (harboring endo-glucanases/xylanases, arabinofuranosidases, and esterases), whereas Enterococcus and Cellulosimicrobium have the potential to degrade oligosaccharides, metabolize xylose and might produce lactic acid through the phosphoketolase (PK) pathway. Based on this evidence, we conclude that our innovative top-down strategy enriched a unique bacterial consortium that could be useful in biotechnological applications, including the development/design of a synthetic bioinoculant to improve silage processes.

Diffusion-Linked Microbial Metabolism in the Vadose Zone

1999 ◽  
Author(s):  
J. E. Watson ◽  
R. F. Harris
Keyword(s):  
Vadose Zone ◽  
Plant Biomass ◽  
Higher Plants ◽  
High Energy ◽  
Essential Elements ◽  
Biofilm Reactor ◽  
Plant Residues ◽  
Soil Organisms ◽  
Transport And Transformation ◽  
Low Energy Electrons

Figure 7.1 is a schematic of nutrient and contaminant transformations and cycling in the vadose zone. As detailed in Harris and Arnold (1995), higher plants take up C, N, P, and S in their most oxidized forms and use, via photosynthesis, the Sun’s energy and low-energy electrons from the oxygen in water to convert the oxidized forms of these essential elements into the relatively high energy reduced forms comprising plant biomass. Following plant death, the biomass residues enter the soil and are attacked by soil organisms as a source of food. The plant residues are depolymerized and the reduced, high-energy monomers are assimilated in part into soil organism biomass, and in part are used as electron donors to combine with the most thermodynamically efficient electron acceptors for dissimilatory energy generation to drive growth and maintenance reactions. In aerobic zones, oxygen is the preferred electron acceptor as long as it is nonlimiting. Death of soil organisms produces dead biomass which re-enters the biological reactor. Ultimately, via respiration in aerobic soils, all the reduced C, N, P, and S materials are released as their oxidized forms, and oxygen is reduced to water to complete the cycle. Ideally, the cycle is conservative, particularly from the standpoint of nonleakage of nutrients, such as nitrate, into the groundwater. Similarly, contaminants entering the vadose zone, either as a function of agronomic use or by accident, should ideally be integrated into the natural nutrient cycles and converted to harmless by-products for assimilation and dissimilation by soil organisms and higher plants (Liu, 1994). Management of nutrient and contaminant transformations by the soil organisms requires a thorough understanding of the ecophysiological and solute transport ground rules that control the nature and rates of transformation options available to the soil organisms. In models of chemical transport and transformation through the vadose zone, colonies of microorganisms are frequently treated as a homogeneous biofilm reactor (Grant and Rochette, 1994). Often, modeling efforts are focused on environmental conditions external to the microbial colony. This consideration of the colony as a biofilm with relatively constant nutrient uptake rates ignores the growth differentiation that occurs as the colony develops

scholarly journals Carbohydrate and Amino Acid Profiles of Cotton Plant Biomass Products

Agriculture ◽  
2019 ◽  
Vol 10 (1) ◽  
pp. 2 ◽  
Author(s):  
Zhongqi He ◽  
Dan C. Olk ◽  
Haile Tewolde ◽  
Hailin Zhang ◽  
Mark Shankle
Keyword(s):  
Amino Acids ◽  
Cotton Plant ◽  
Management Practices ◽  
Plant Biomass ◽  
Plant Residues ◽  
Cotton Leaf ◽  
Essential Information ◽  
Plant Parts ◽  
Processing Strategies ◽  
Biomass Materials

To achieve the optimal and diverse utilization of cotton (Gossypium hirsutum) plant residues in various agricultural, industrial, and environmental applications, the chemical composition of cotton biomass tissues across different plant parts (e.g., seed, boll, bur, leaves, stalk, stem, and root) is of essential information. Thus, in this work, we collected field-grown whole mature cotton plants and separated them into distinct biomass fractions including main stems, leaf blades, branches, petioles, roots, and reproductive parts (mid-season growth stage) or bur, peduncles/bract, and seed cotton (pre-defoliation stage). The contents of selected carbohydrates and amino acids in these cotton biomass materials were determined. Both essential and nonessential amino acids were enriched in cotton leaf blades and reproductive parts. The distribution pattern of the selected carbohydrates differed from that of amino acids—higher contents of carbohydrate were found in roots, main stems, and branches. Although glucose was the most abundant non-structural carbohydrate in cotton plant parts at mid-season, xylose was the most abundant in most plant parts at the pre-defoliation stage. Nutritional carbohydrates and amino acids were further accumulated in seeds at pre-defoliation. The information reported in this work would be helpful in exploring and optimizing management practices and processing strategies for utilizing cotton crop biomass materials as valuable and renewable natural resources.

Physiological Characterization of a Bacterial Consortium Reductively Dechlorinating 1,2,3- and 1,2,4-Trichlorobenzene

1998 ◽  
Vol 64 (2) ◽  
pp. 496-503 ◽  
Author(s):  
Lorenz Adrian ◽  
Werner Manz ◽  
Ulrich Szewzyk ◽  
Helmut Görisch
Keyword(s):  
Energy Source ◽  
Sulfate Reduction ◽  
Reductive Dechlorination ◽  
Microbial Consortium ◽  
Bacterial Consortium ◽  
Oligonucleotide Probes ◽  
Electron Sources ◽  
Physiological Characterization

ABSTRACT A bacterial mixed culture reductively dechlorinating trichlorobenzenes was established in a defined, synthetic mineral medium without any complex additions and with pyruvate as the carbon and energy source. The culture was maintained over 39 consecutive transfers of small inocula into fresh media, enriching the dechlorinating activity. In situ probing with fluorescence-labeled rRNA-targeted oligonucleotide probes revealed that two major subpopulations within the microbial consortium were phylogenetically affiliated with a sublineage within the Desulfovibrionaceaeand the gamma subclass of Proteobacteria. The bacterial consortium grew by fermentation of pyruvate, forming acetate, propionate, CO2, formate, and hydrogen. Acetate and propionate supported neither the reduction of trichlorobenzenes nor the reduction of sulfate when sulfate was present. Hydrogen and formate were used for sulfate reduction to sulfide. Sulfate strongly inhibited the reductive dechlorination of trichlorobenzenes. However, when sulfate was depleted in the medium due to sulfate reduction, dechlorination of trichlorobenzenes started. Similar results were obtained when sulfite was present in the cultures. Molybdate at a concentration of 1 mM strongly inhibited the dechlorination of trichlorobenzenes. Cultures supplied with molybdate plus sulfate did not reduce sulfate, but dechlorination of trichlorobenzenes occurred. Supplementation of electron-depleted cultures with various electron sources demonstrated that formate was used as a direct electron donor for reductive dechlorination, whereas hydrogen was not.

scholarly journals Molecular Basis of a Bacterial Consortium: Interspecies Catabolism of Atrazine

1998 ◽  
Vol 64 (1) ◽  
pp. 178-184 ◽  
Author(s):  
Mervyn L. de Souza ◽  
David Newcombe ◽  
Sam Alvey ◽  
David E. Crowley ◽  
Anthony Hay ◽  
...  
Keyword(s):  
Nitrogen Source ◽  
Southern Hybridization ◽  
Cyanuric Acid ◽  
Bacterial Species ◽  
Bacterial Consortium ◽  
Triazine Ring ◽  
Carbon And Nitrogen ◽  
Consortium Member ◽  
Ring Compounds ◽  
Pure Cultures

ABSTRACT Pseudomonas sp. strain ADP contains the genes,atzA, -B, and -C, that encode three enzymes which metabolize atrazine to cyanuric acid. Atrazine-catabolizing pure cultures isolated from around the world contain genes homologous to atzA, -B, and -C. The present study was conducted to determine whether the same genes are present in an atrazine-catabolizing bacterial consortium and how the genes and metabolism are subdivided among member species. The consortium contained four or more bacterial species, but two members, Clavibacter michiganese ATZ1 andPseudomonas sp. strain CN1, collectively mineralized atrazine. C. michiganese ATZ1 released chloride from atrazine, produced hydroxyatrazine, and contained a homolog to theatzA gene that encoded atrazine chlorohydrolase. C. michiganese ATZ1 stoichiometrically metabolized hydroxyatrazine to N-ethylammelide and contained genes homologous toatzB and atzC, suggesting that either a functional AtzB or -C catalyzed N-isopropylamine release from hydroxyatrazine. C. michiganese ATZ1 grew on isopropylamine as its sole carbon and nitrogen source, explaining the ability of the consortium to use atrazine as the sole carbon and nitrogen source. A second consortium member, Pseudomonassp. strain CN1, metabolized the N-ethylammelide produced byC. michiganese ATZ1 to transiently form cyanuric acid, a reaction catalyzed by AtzC. A gene homologous to the atzCgene of Pseudomonas sp. strain ADP was present, as demonstrated by Southern hybridization and PCR. Pseudomonassp. strain CN1, but not C. michiganese, metabolized cyanuric acid. The consortium metabolized atrazine faster than didC. michiganese individually. Additionally, the consortium metabolized a much broader set of triazine ring compounds than did previously described pure cultures in which the atzABCgenes had been identified. These data begin to elucidate the genetic and metabolic bases of catabolism by multimember consortia.

scholarly journals Raw Cow Milk Bacterial Consortium as Bioindicator of Circulating Anti-Microbial Resistance (AMR)

Animals ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 2378
Author(s):  
Cristian Piras ◽  
Viviana Greco ◽  
Enrico Gugliandolo ◽  
Alessio Soggiu ◽  
Bruno Tilocca ◽  
...  
Keyword(s):  
Ecological Niche ◽  
Bacterial Species ◽  
Raw Milk ◽  
Bacterial Consortium ◽  
Cow Milk ◽  
Milk Analysis ◽  
Animal Products ◽  
Bacterial Consortia ◽  
Expression Of Genes ◽  
Ecological Pressure

The environment, including animals and animal products, is colonized by bacterial species that are typical and specific of every different ecological niche. Natural and human-related ecological pressure promotes the selection and expression of genes related to antimicrobial resistance (AMR). These genes might be present in a bacterial consortium but might not necessarily be expressed. Their expression could be induced by the presence of antimicrobial compounds that could originate from a given ecological niche or from human activity. In this work, we applied (meta)proteomics analysis of bacterial compartment of raw milk in order to obtain a method that provides a measurement of circulating AMR involved proteins and gathers information about the whole bacterial composition. Results from milk analysis revealed the presence of 29 proteins/proteoforms linked to AMR. The detection of mainly β-lactamases suggests the possibility of using the milk microbiome as a bioindicator for the investigation of AMR. Moreover, it was possible to achieve a culture-free qualitative and functional analysis of raw milk bacterial consortia.

scholarly journals Effects of Plant Biomass, Plant Diversity, and Water Content on Bacterial Communities in Soil Lysimeters: Implications for the Determinants of Bacterial Diversity

2007 ◽  
Vol 73 (21) ◽  
pp. 6916-6929 ◽  
Author(s):  
Delita Zul ◽  
Sabine Denzel ◽  
Andrea Kotz ◽  
J�rg Overmann
Keyword(s):  
Species Composition ◽  
Water Content ◽  
Bacterial Diversity ◽  
Plant Species ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Plant Biomass ◽  
Bacterial Species ◽  
Bacterial Community Composition ◽  
Ecological Niches ◽  
Soil Lysimeters

ABSTRACT Soils may comprise tens of thousands to millions of bacterial species. It is still unclear whether this high level of diversity is governed by functional redundancy or by a multitude of ecological niches. In order to address this question, we analyzed the reproducibility of bacterial community composition after different experimental manipulations. Soil lysimeters were planted with four different types of plant communities, and the water content was adjusted. Group-specific phylogenetic fingerprinting by PCR-denaturing gradient gel electrophoresis revealed clear differences in the composition of Alphaproteobacteria, Betaproteobacteria, Bacteroidetes, Chloroflexi, Planctomycetes, and Verrucomicrobia populations in soils without plants compared to that of populations in planted soils, whereas no influence of plant species composition on bacterial diversity could be discerned. These results indicate that the presence of higher plant species affects the species composition of bacterial groups in a reproducible manner and even outside of the rhizosphere. In contrast, the environmental factors tested did not affect the composition of Acidobacteria, Actinobacteria, Archaea, and Firmicutes populations. One-third (52 out of 160) of the sequence types were found to be specifically and reproducibly associated with the absence or presence of plants. Unexpectedly, this was also true for numerous minor constituents of the soil bacterial assemblage. Subsequently, one of the low-abundance phylotypes (beta10) was selected for studying the interdependence under particular experimental conditions and the underlying causes in more detail. This so-far-uncultured phylotype of the Betaproteobacteria species represented up to 0.18% of all bacterial cells in planted lysimeters compared to 0.017% in unplanted systems. A cultured representative of this phylotype exhibited high physiological flexibility and was capable of utilizing major constituents of root exudates. Our results suggest that the bacterial species composition in soil is determined to a significant extent by abiotic and biotic factors, rather than by mere chance, thereby reflecting a multitude of distinct ecological niches.

scholarly journals Rapid Mineralization of Benzo[a]pyrene by a Microbial Consortium Growing on Diesel Fuel

2000 ◽  
Vol 66 (10) ◽  
pp. 4205-4211 ◽  
Author(s):  
Robert A. Kanaly ◽  
Richard Bartha ◽  
Kazuya Watanabe ◽  
Shigeaki Harayama
Keyword(s):  
Incubation Period ◽  
Diesel Fuel ◽  
Microbial Consortium ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Cell Protein ◽  
Rrna Gene ◽  
Gradient Gel Electrophoresis ◽  
Balance Analysis ◽  
Pyrene Concentration ◽  
Sequence Types

ABSTRACT A microbial consortium which rapidly mineralized the environmentally persistent pollutant benzo[a]pyrene was recovered from soil. The consortium cometabolically converted [7-14C]benzo[a]pyrene to14CO2 when it was grown on diesel fuel, and the extent of benzo[a]pyrene mineralization was dependent on both diesel fuel and benzo[a]pyrene concentrations. Addition of diesel fuel at concentrations ranging from 0.007 to 0.2% (wt/vol) stimulated the mineralization of 10 mg of benzo[a]pyrene per liter 33 to 65% during a 2-week incubation period. When the benzo[a]pyrene concentration was 10 to 100 mg liter−1 and the diesel fuel concentration was 0.1% (wt/vol), an inoculum containing 1 mg of cell protein per liter (small inoculum) resulted in mineralization of up to 17.2 mg of benzo[a]pyrene per liter in 16 days. This corresponded to 35% of the added radiolabel when the concentration of benzo[a]pyrene was 50 mg liter−1. A radiocarbon mass balance analysis recovered 25% of the added benzo[a]pyrene solubilized in the culture suspension prior to mineralization. Populations growing on diesel fuel most likely promoted emulsification of benzo[a]pyrene through the production of surface-active compounds. The consortium was also analyzed by PCR-denaturing gradient gel electrophoresis of 16S rRNA gene fragments, and 12 dominant bands, representing different sequence types, were detected during a 19-day incubation period. The onset of benzo[a]pyrene mineralization was compared to changes in the consortium community structure and was found to correlate with the emergence of at least four sequence types. DNA from 10 sequence types were successfully purified and sequenced, and that data revealed that eight of the consortium members were related to the classProteobacteria but that the consortium also included members which were related to the genera Mycobacterium andSphingobacterium.

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