Spike Formation Is a Turning Point Determining Wheat Root Microbiome Abundance, Structures and Functions.

Root selection of their associated microbiome composition and activities is determined by the plant's developmental stage and distance from the root. Total gene abundance, structure and functions of root-associated and rhizospheric microbiomes were studied throughout wheat growth season under field conditions. On the root surface, abundance of the well-known wheat colonizers Proteobacteria and Actinobacteria decreased and increased, respectively, during spike formation, whereas abundance of Bacteroidetes was independent of spike formation. Metagenomic analysis combined with functional co-occurrence networks revealed a significant impact of plant developmental stage on its microbiome during the transition from vegetative growth to spike formation. For example, gene functions related to biofilm and sensorial movement, antibiotic production and resistance and carbons and amino acids and their transporters. Genes associated with these functions were also in higher abundance in root vs. the rhizosphere microbiome. We propose that abundance of transporter-encoding genes related to carbon and amino acid, may mirror the availability and utilization of root exudates. Genes related to antibiotic resistance mechanisms were abundant during vegetative growth, while after spike formation, genes related to the biosynthesis of various antibiotics were enriched. This observation suggests that during root colonization and biofilm formation, bacteria cope with competitor's antibiotics, whereas in the mature biofilm stage, they invest in inhibiting new colonizers. Additionally, there is higher abundance of genes related to denitrification in rhizosphere compared to root-associated microbiome during wheat growth, possibly due to competition with the plant over nitrogen in the root vicinity. We demonstrated functional and phylogenetic division in wheat root zone microbiome in both time and space: pre- and post-spike formation, and root-associated vs. rhizospheric niches. These findings shed light on the dynamics of plant-microbe and microbe-microbe interactions in the developing root zone.

Quality of Irrigation Water Affects Soil Functionality and Bacterial Community Stability in Response to Heat Disturbance.

Anthropogenic activities alter the structure and function of a bacterial community. Furthermore, bacterial communities structured by the conditions the anthropogenic activities present may consequently reduce their stability in response to an unpredicted acute disturbance. The present mesocosm-scale study exposed soil bacterial communities to different irrigation water types, including freshwater, fertilized freshwater, treated wastewater, and artificial wastewater, and evaluated their response to a disturbance caused by heat. These effectors may be considered deterministic and stochastic forces common in agricultural operations of arid and semiarid regions. Bacterial communities under conditions of high mineral and organic carbon availability (artificial wastewater) differed from the native bacterial community and showed a proteobacterial dominance. These bacterial communities had a lower resistance to the heat treatment disturbance than soils under conditions of low resource availability (high-quality treated wastewater or freshwater). The latter soil bacterial communities showed a higher abundance of operational taxonomic units (OTUs) classified as Bacilli These results were elucidated by soil under conditions of high resource availability, which lost higher degrees of functional potential and had a greater bacterial community composition change. However, the functional resilience, after the disturbance ended, was higher under a condition of high resource availability despite the bacterial community composition shift and the decrease in species richness. The functional resilience was directly connected to the high growth rates of certain Bacteroidetes and proteobacterial groups. A high stability was found in samples that supported the coexistence of both resistant OTUs and fast-growing OTUs.IMPORTANCE This report presents the results of a study employing a hypothesis-based experimental approach to reveal the forces involved in determining the stability of a soil bacterial community to disturbance. The resultant postdisturbance bacterial community composition dynamics and functionality were analyzed. The paper demonstrates the relatedness of community structure and stability under cultivation conditions prevalent in an arid area under irrigation with water of different qualities. The use of common agricultural practices to demonstrate these features has not been described before. The combination of a fundamental theoretical issue in ecology with common and concerning disturbances caused by agricultural practice makes this study unique. Furthermore, the results of the present study have applicable importance regarding soil conservation, as it enables a better characterization and monitoring of stressed soil bacterial communities and possible intervention to reduce the stress. It will also be of valued interest in coming years, as fresh water scarcity and the use of alternative water sources are expected to rise globally.

Resilience of soil bacterial community to irrigation with water of different qualities under Mediterranean climate.

Limited freshwater (FW) availability due to climate change and increasing global populations is driving agriculture in arid and semi-arid regions to recycle vast quantities of water for irrigation. However, irrigation with treated wastewater (TWW), which contains dissolved organic matter, salts and microorganisms might alter soil microbial populations, and thus affect soil fertility. We characterized the effects of irrigation with TWW and FW on soil bacterial community composition for three consecutive years. Orchard samples were collected at the end of each of the rainy (winter) and irrigation (summer) season. Community composition, determined by 16S ribosomal DNA amplicon pyrosequencing, was highly similar in soil samples obtained at the end of three consecutive rainy seasons, regardless of irrigation season water quality. However, whereas composition in soil shifted slightly during irrigation seasons by FW irrigation, it was greatly influenced by TWW irrigation. During the irrigation season, a decrease in the relative abundance of Actinobacteria was observed; along with an increase in the relative abundance Gammaproteobacteria within TWW-irrigated soils. The return to the 'baseline state' during the rainy season demonstrates that the soil community is not resistant to anthropogenic impact imposed by irrigation water quality, yet is resilient in long term.

Host signature effect on plant root-associated microbiomes revealed through analyses of resident vs. active communities.

Plant roots create specific microbial habitat in the soil - the rhizosphere. In this study, we characterized the rhizosphere microbiome of four host plant species to get insight into the impact of the host (host signature effect) on resident vs. active communities. Results show a distinct plant host specific signature found among wheat, maize, tomato and cucumber, based on the following three parameters: (i) each plant promoted the activity of a unique suite of soil bacterial populations; (ii) significant variations were observed in the number and the degree of dominance of active populations; and (iii) the level of contribution of active (rRNA-based) populations to the resident (DNA-based) community profiles. In the rhizoplane of all four plants, a significant reduction of diversity was observed, relative to the bulk soil. Moreover, an increase in DNA-RNA correspondence indicated higher representation of active bacterial populations in the residing rhizoplane community. This study demonstrates that the host plant determines the bacterial community composition in its immediate vicinity, especially with respect to the active populations.

The role of the bacterial community in the nutritional ecology of the bulb mite Rhizoglyphus robini (Acari: Astigmata: Acaridae).

The biology of many arthropods can only be understood when their associated microbiome is considered. The nutritional requirements of the bulb mite Rhizoglyphus robini Claparede (Acari: Astigmata: Acaridae) in the laboratory have been shown to be very easily satisfied, and in the field the mites prefer fungus-infected over uninfected plants. To test whether symbiotic bacteria facilitate the survival of R. robini on a temporarily nutritionally unbalanced diet, we investigated the composition of its microbiome. Using 454 pyrosequencing of 16S rRNA gene fragments, 3 genera were found to dominate the bacterial community: Myroides (41.4%), Serratia (11.4%), and Alcaligenes (4.5%); the latter 2 are known to include chitinase-producing species. Laboratory experiments demonstrated that mite fecundity is significantly higher (2 times) on fungus than on controls (sterilized potato dextrose agar and filter paper). Also, when mite homogenate was applied to a chitin layer, the halo produced through degradation was clearly visible, while the saline control did not produce a halo. We thus concluded that R. robini utilizes fungal chitin, at least to a certain extent, as a food source with the help of its associated bacteria. This information supports the general concept of multigenome organisms and the involvement of bacteria in the mite's nutritional ecology.

Volatile organic compounds of the soil bacterium Bacillus halotolerans suppress pathogens and elicit defense-responsive genes in plants.

Volatile organic compounds (VOCs) produced by bacteria play an important, yet relatively unexplored role in interactions between plants and phytopathogens. In this study, the soil bacterium Bacillus halotolerans NYG5 was identified as a potent biocontrol agent against several phytopathogenic fungi (Macrophomina phaseolina, Rhizoctonia solani, Pythium aphanidermatum, and Sclerotinia sclerotiorum) through the production of VOCs. NYG5-emitted VOCs also inhibited the growth of bacterial pathogens (Agrobacterium tumefaciens, Xanthomonas campestris, Clavibacter michiganensis, and Pseudomonas syringae). When cultured in various growth media, NYG5 produced a variety of VOCs. Five distinct VOCs (2-methylbutanoic acid, 5-methyl-2-hexanone, 2,3-hexanedione, 2-ethyl-1-hexanol, and 6-methyl-2-heptanone) were identified using headspace GC-MS. 2,3-Hexanedione exhibited potent lethal effects on the tested phytopathogens and nematicidal activity against Meloidogyne javanica at a concentration of 50 ppm. In addition, 0.05 ppm 2,3-hexanedione stimulated the expression of pathogenesis-related genes 1 and 2 in Arabidopsis thaliana. Interestingly, 2,3-hexanedione is used as a food additive at higher concentrations than those tested in this study. Hence, 2,3-hexanedione is a promising biologically active compound that might serve as a sustainable alternative to common chemical pesticides and an elicitor of plant defense.

A pipeline for rapidly evaluating activity and inferring mechanisms of action of prospective antifungal compounds.

BACKGROUND: Fungal phytopathogens are a significant threat to crops and food security, and there is a constant need to develop safe and effective compounds that antagonize them. In-planta assays are complex and tedious and are thus not suitable for initial high-throughput screening of new candidate antifungal compounds. We propose an in vitro screening pipeline that integrates five rapid quantitative and qualitative methods to estimate the efficacy and mode of action of prospective antifungal compounds. RESULTS: The pipeline was evaluated using five documented antifungal compounds (benomyl, catechol, cycloheximide, 2,4-diacetylphloroglucinol, and phenylacetic acid) that have different modes of action and efficacy, against the model soilborne fungal pathogen Fusarium oxysporum f. sp. radicis cucumerinum. We initially evaluated the five compounds' ability to inhibit fungal growth and metabolic activity using green fluorescent protein (GFP)-labeled F. oxysporum and PrestoBlue staining, respectively, in multiwell plate assays. We tested the compounds' inhibition of both conidial germination and hyphal elongation. We then employed FUN-1 and SYTO9/propidium iodide staining, coupled to confocal microscopy, to differentiate between fungal growth inhibition and death at the cellular level. Finally, using a reactive oxygen species (ROS)-detection assay, we were able to quantify ROS production in response to compound application. CONCLUSIONS: Collectively, the proposed pipeline provides a wide array of quantitative and qualitative data on the tested compounds that can help pinpoint promising novel compounds; these can then be evaluated more vigorously using in planta screening assays. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Soil suppressiveness to fusarium disease: shifts in root microbiome associated with reduction of pathogen root colonization.

Soil suppressiveness to Fusarium disease was induced by incubating sandy soil with debris of wild rocket (WR; Diplotaxis tenuifolia) under field conditions. We studied microbial dynamics in the roots of cucumber seedlings following transplantation into WR-amended or nonamended soil, as influenced by inoculation with Fusarium oxysporum f. sp. radicis-cucumerinum. Disease symptoms initiated in nonamended soil 6 days after inoculation, compared with 14 days in WR-amended soil. Root infection by F. oxysporum f. sp. radicis-cucumerinum was quantified using real-time polymerase chain reaction (PCR). Target numbers were similar 3 days after inoculation for both WR-amended and nonamended soils, and were significantly lower (66%) 6 days after inoculation and transplanting into the suppressive (WR-amended) soil. This decrease in root colonization was correlated with a reduction in disease (60%) 21 days after inoculation and transplanting into the suppressive soil. Fungal community composition on cucumber roots was assessed using mass sequencing of fungal internal transcribed spacer gene fragments. Sequences related to F. oxysporum, Fusarium sp. 14005, Chaetomium sp. 15003, and an unclassified Ascomycota composed 96% of the total fungal sequences in all samples. The relative abundances of these major groups were highly affected by root inoculation with F. oxysporum f. sp. radicis-cucumerinum, with a 10-fold increase in F. oxysporum sequences, but were not affected by the WR amendment. Quantitative analysis and mass-sequencing methods indicated a qualitative shift in the root's bacterial community composition in suppressive soil, rather than a change in bacterial numbers. A sharp reduction in the size and root dominance of the Massilia population in suppressive soil was accompanied by a significant increase in the relative abundance of specific populations; namely, Rhizobium, Bacillus, Paenibacillus, and Streptomyces spp. Composition of the Streptomyces community shifted significantly, as determined by PCR denaturing gradient gel electrophoresis, resulting in an increase in the dominance of a specific population in suppressive soils after only 3 days. This shift was related mainly to the increase in Streptomyces humidus, a group previously described as antagonistic to phytopathogenic fungi. Thus, suitable soil amendment resulted in a shift in the root's bacterial communities, and infection by a virulent pathogen was contained by the root microbiome, leading to a reduced disease rate.

Comparative genomics of Bacillus cereus sensu lato spp. biocontrol strains in correlation to in-vitro phenotypes and plant pathogen antagonistic capacity.

Bacillus cereus sensu lato (Bcsl) strains are widely explored due to their capacity to antagonize a broad range of plant pathogens. These include B. cereus sp. UW85, whose antagonistic capacity is attributed to the secondary metabolite Zwittermicin A (ZwA). We recently isolated four soil and root-associated Bcsl strains (MO2, S-10, S-25, LSTW-24) that displayed different growth profiles and in-vitro antagonistic effects against three soilborne plant pathogens models: Pythium aphanidermatum (oomycete) Rhizoctonia solani (basidiomycete), and Fusarium oxysporum (ascomycete). To identify genetic mechanisms potentially responsible for the differences in growth and antagonistic phenotypes of these Bcsl strains, we sequenced and compared their genomes, and that of strain UW85 using a hybrid sequencing pipeline. Despite similarities, specific Bcsl strains had unique secondary metabolite and chitinase-encoding genes that could potentially explain observed differences in in-vitro chitinolytic potential and anti-fungal activity. Strains UW85, S-10 and S-25 contained a (~500 Kbp) mega-plasmid that harbored the ZwA biosynthetic gene cluster. The UW85 mega-plasmid contained more ABC transporters than the other two strains, whereas the S-25 mega-plasmid carried a unique cluster containing cellulose and chitin degrading genes. Collectively, comparative genomics revealed several mechanisms that can potentially explain differences in in-vitro antagonism of Bcsl strains toward fungal plant pathogens.

Bacterial volatile organic compounds as biopesticides, growth promoters and plant-defense elicitors: Current understanding and future scope.

Bacteria emit a large number of volatile organic compounds (VOCs) into the environment. VOCs are species-specific and their emission depends on environmental conditions, such as growth medium, pH, temperature, incubation time and interaction with other microorganisms. These VOCs can enhance plant growth, suppress pathogens and act as signaling molecules during plant-microorganism interactions. Some bacterial VOCs have been reported to show strong antimicrobial, nematicidal, pesticidal, plant defense, induced tolerance and plant-growth-promoting activities under controlled conditions. Commonly produced antifungal VOCs include dimethyl trisulfide, dimethyl disulfide, benzothiazole, nonane, decanone and 1-butanol. Species of Bacillus, Pseudomonas, Arthrobacter, Enterobacter and Burkholderia produce plant growth promoting VOCs, such as acetoin and 2,3-butenediol. These VOCs affect expression of genes involved in defense and development in plant species (i.e., Arabidopsis, tobacco, tomato, potato, millet and maize). VOCs are also implicated in altering pathogenesis-related genes, inducing systemic resistance, modulating plant metabolic pathways and acquiring nutrients. However, detailed mechanisms of action of VOCs need to be further explored. This review summarizes the bioactive VOCs produced by diverse bacterial species as an alternative to agrochemicals, their mechanism of action and challenges for employment of bacterial VOCs for sustainable agricultural practices. Future studies on technological improvements for bacterial VOCs application under greenhouse and open field conditions are warranted.

Corrigendum: Comparative genomics of Bacillus cereus sensu lato spp. biocontrol strains in correlation to in-vitro phenotypes and plant pathogen antagonistic capacity.

[This corrects the article DOI: 10.3389/fmicb.2023.996287.].

Short- and long-term effects of continuous compost amendment on soil microbiome community.

Organic amendment, and especially the use of composts, is a well-accepted sustainable agricultural practice. Compost increases soil carbon and microbial biomass, changes enzymatic activity, and enriches soil carbon and nitrogen stocks. However, relatively little is known about the immediate and long-term temporal dynamics of agricultural soil microbial communities following repeated compost applications. Our study was conducted at two field sites: Newe Ya'ar (NY, Mediterranean climate) and Gilat (G, semi-arid climate), both managed organically over 4 years under either conventional fertilization (0, zero compost) or three levels of compost amendment (20, 40 and 60 m3/ha or 2, 4, 6 L/m2). Microbial community dynamics in the soils was examined by high- and low-time-resolution analyses. Annual community composition in compost-amended soils was significantly affected by compost amendment levels in G (first, second and third years) and in NY (third year). Repeated sampling at high resolution (9-10 times over 1 year) showed that at both sites, compost application initially induced a strong shift in microbial communities, lasting for up to 1 month, followed by a milder response. Compost application significantly elevated alpha diversity at both sites, but differed in the compost-dose correlation effect. We demonstrate higher abundance of taxa putatively involved in organic decomposition and characterized compost-related indicator taxa and a compost-derived core microbiome at both sites. Overall, this study describes temporal changes in the ecology of soil microbiomes in response to compost vs. conventional fertilization.

Freshwater microbial metagenomes sampled across different water body characteristics, space and time in Israel.

Freshwater bodies are critical components of terrestrial ecosystems. The microbial communities of freshwater ecosystems are intimately linked water quality. These microbes interact with, utilize and recycle inorganic elements and organic matter. Here, we present three metagenomic sequence datasets (total of 182.9 Gbp) from different freshwater environments in Israel. The first dataset is from diverse freshwater bodies intended for different usages - a nature reserve, irrigation and aquaculture facilities, a tertiary wastewater treatment plant and a desert rainfall reservoir. The second represents a two-year time-series, collected during 2013-2014 at roughly monthly intervals, from a water reservoir connected to an aquaculture facility. The third is from several time-points during the winter and spring of 2015 in Lake Kinneret, including a bloom of the cyanobacterium Microcystis sp. These datasets are accompanied by physical, chemical, and biological measurements at each sampling point. We expect that these metagenomes will facilitate a wide range of comparative studies that seek to illuminate new aspects of freshwater microbial ecosystems and inform future water quality management approaches.

Colonization of cucumber seeds by bacteria during germination.

Detailed analysis revealed fundamental differences between bacterial association with cucumber (Cucumis sativus) seeds and seedlings roots. Seed colonization by bacteria seems to result from passive encounter between bacteria, conveyed by imbibed soil solution, and the germinating seed. In accordance, the seed-associated bacterial community composition directly reflected that of the germination medium and was characterized by low dominance. Transition from seed to root was marked by a shift in bacterial community composition and in an increase in dominance values. Furthermore, settlement of bacteria on roots was tightly controlled by the specific properties of each root segment. Size and richness of the seed-associated bacterial community were clearly determinate by the community in the germination medium. In contrast, for fully developed and active roots, the medium effect on these parameters was negligible. Perturbation of the seed environment by a pathogen (Pythium aphanidermatum) had major consequences on the seed bacterial community. However, those were mostly related to direct pathogen-bacteria rather than seed-bacteria interactions. In conclusion, simple, even passive processes may determine the initial stage of plant-microbe association during seed germination, prior to extension of the primary root. Therefore, seed germination is a unique phase in the plant life cycle, with respect to its interaction with the below-ground microbiome.

Isolation and bioaugmentation of an estradiol-degrading bacterium and its integration into a mature biofilm.

Bioaugmentation can alter the potential activity as well as the composition of the naturally occurring microbial biota during bioremediation of a contaminated site. The focus of the current study is the pollutant 17ß-estradiol (E2), which can cause endocrine effects and is potentially harmful to aquatic biota and to public health. The community composition and function of biofilms, originating from a wetland system, as affected by augmentation of an estradiol-degrading bacterium (EDB-LI1) under different conditions, were investigated. EDB-LI1 inoculation into biofilm from two wetland ponds representing early and advanced water treatment stages, respectively, yielded three significant observations, as follows: (i) EDB-LI1, enriched from a biofilm of a constructed wetland wastewater treatment system, was detected (by quantitative PCR [qPCR] analysis) in this environment in the augmented biofilm only; (ii) the augmented biofilm acquired the ability to remove estradiol; and (iii) the bacterial community composition (analyzed by PCR-denaturing gradient gel electrophoresis [DGGE]) of the augmented biofilm differed from that of the control biofilm. Furthermore, EDB-LI1 bioaugmentation showed a higher level of removal of estradiol with biofilms that originated from the advanced-treatment-stage wetland pond than those from the early-treatment-stage pond. Hence, the bioaugmentation efficiency of EDB-LI1 depends on both the quality of the feed water and the microbial community composition in the pond.

Microbial activity and organic matter dynamics during 4 years of irrigation with treated wastewater.

The global changes in rainfall frequency and quantity have subjected arid and semi-arid regions to long periods of drought. As this phenomenon corresponds to increasing trend of water shortage, the use of treated wastewater (TWW) has been suggested as an alternative for irrigation of agricultural crops in these areas. The aim of the study was to investigate the short- and middle-term effects of TWW irrigation on the soil microbial activities and organic carbon content. The microbial community activity was measured every 1-3 months for 4 years in a persimmon (Diospyros kaki) orchard. These activities were used here as an indicator for the soil health. The hydrolysis activity (detected by fluorescein diacetate hydrolysis (FDA) assay) increased during the irrigation season and was significantly higher in soils irrigated with TWW compared to those irrigated with freshwater (FW). This activity was also negatively correlated with dissolved organic carbon (DOC) concentrations during the irrigation season, suggesting that the community degraded the DOC in the soils regardless of its origin. The irrigation season was also characterized by an increase in nitrification potential in both TWW- and FW-irrigated soils, which coincided with high concentrations of nitrate (50 mg kg(-1) soil). Overall, there was an increase in all measured activities during the irrigation season, and they were higher in the TWW soils. However, it appears that after each irrigation season, the potential activity of the community returned to levels similar to or even slightly lower than those of FW-irrigated soil during the wet season, suggesting that the periodic irrigation did not significantly change the soil microbial activity.

Molecular analysis of bacterial communities in raw cow milk and the impact of refrigeration on its structure and dynamics.

The impact of refrigeration on raw cow milk bacterial communities in three farm bulk tanks and three dairy plant silo tanks was studied using two methods: DGGE and cloning. Both methods demonstrated that bacterial taxonomic diversity decreased during refrigeration. Gammaproteobacteria, especially Pseudomonadales, dominated the milk after refrigeration. Farm samples and dairy plant samples differed in their microbial community composition, the former showing prevalence of Gram-positive bacteria affiliated with the classes Bacilli, Clostridia and Actinobacteria, the latter showing prevalence of Gram-negative species belonging to the Gammaproteobacteria class. Actinobacteria prevalence in the farm milk samples immediately after collection stood at about 25% of the clones. A previous study had found that psychrotolerant Actinobacteria identified in raw cow milk demonstrated both lipolytic and proteolytic enzymatic activity. Thus, we conclude that although Pseudomonadales play an important role in milk spoilage after long periods of cold incubation, Actinobacteria occurrence may play an important role when assessing the quality of milk arriving at the dairy plant from different farms. As new cooling technologies reduce the initial bacterial counts of milk to very low levels, more sensitive and efficient methods to evaluate the bacterial quality of raw milk are required. The present findings are an important step towards achieving this goal.

Elevated CO2 and nitrate levels increase wheat root-associated bacterial abundance and impact rhizosphere microbial community composition and function.

Elevated CO2 stimulates plant growth and affects quantity and composition of root exudates, followed by response of its microbiome. Three scenarios representing nitrate fertilization regimes: limited (30 ppm), moderate (70 ppm) and excess nitrate (100 ppm) were compared under ambient and elevated CO2 (eCO2, 850 ppm) to elucidate their combined effects on root-surface-associated bacterial community abundance, structure and function. Wheat root-surface-associated microbiome structure and function, as well as soil and plant properties, were highly influenced by interactions between CO2 and nitrate levels. Relative abundance of total bacteria per plant increased at eCO2 under excess nitrate. Elevated CO2 significantly influenced the abundance of genes encoding enzymes, transporters and secretion systems. Proteobacteria, the largest taxonomic group in wheat roots (~ 75%), is the most influenced group by eCO2 under all nitrate levels. Rhizobiales, Burkholderiales and Pseudomonadales are responsible for most of these functional changes. A correlation was observed among the five gene-groups whose abundance was significantly changed (secretion systems, particularly type VI secretion system, biofilm formation, pyruvate, fructose and mannose metabolism). These changes in bacterial abundance and gene functions may be the result of alteration in root exudation at eCO2, leading to changes in bacteria colonization patterns and influencing their fitness and proliferation.

Pairwise Interactions of Three Related Pseudomonas Species in Plant Roots and Inert Surfaces.

Bacteria are social organisms that interact extensively within and between species while responding to external stimuli from their environments. Designing synthetic microbial communities can enable efficient and beneficial microbiome implementation in many areas. However, in order to design an efficient community, one must consider the interactions between their members. Using a reductionist approach, we examined pairwise interactions of three related Pseudomonas species in various microenvironments including plant roots and inert surfaces. Our results show that the step between monoculture and co-culture is already very complex. Monoculture root colonization patterns demonstrate that each isolate occupied a particular location on wheat roots, such as root tip, distance from the tip, or scattered along the root. However, pairwise colonization outcomes on the root did not follow the bacterial behavior in monoculture, suggesting various interaction patterns. In addition, we show that interspecies interactions on a microscale on inert surface take part in co-culture colonization and that the interactions are affected by the presence of root extracts and depend on its source. The understanding of interrelationships on the root may contribute to future attempts to manipulate and improve bacterial colonization and to intervene with root microbiomes to construct and design effective synthetic microbial consortia.

Microbial population and activity in wetland microcosms constructed for improving treated municipal wastewater.

The idea of using constructed wetlands for the treatment and improving of wastewater emerged in the second half of the last century. Despite relatively wide use of this environmentally friendly technology, relatively little is known about the microbial populations involved in biotransformation and removal of contaminants in this system. The aim of the current study was to investigate the assembly and function of microbial populations in vertical-flow constructed wetland microcosms designed to improve the quality of wastewater after activated sludge treatment. Also, the performance of 3-year-old wetland ponds was investigated. Even though the quality of the influent water was relatively high, improvement in water parameters such as coliform level, ammonia concentration, BOD, and TSS was observed. The performance of the wetland ponds was comparable to that of the microcosms. The microbial community composition of the biofilm formed on the surface of gravel particles in vegetated and plant-free microcosms was studied by denaturing gradient gel electrophoresis (DGGE) and sequencing of 16S rRNA gene fragments. Highly complex bacterial diversity was observed in the biofilm. Cluster analysis of DGGE patterns demonstrated that depth within the wetland microcosm has a stronger effect on microbial community composition of the biofilm formed on wetland matrix than vegetation. Measurements of fluorescein diacetate hydrolysis activity and nitrification potential revealed that hydrolytic activity was affected by both microcosm depth and vegetation presence, whereas nitrification potential was mostly influenced by depth. Resolving the bacterial assemblage of wetland biofilm, which often is considered a black box, will help to understand the interactions involved in the development of diverse and mature biofilm and its function.