Isolation of Methane Enriched Bacterial Communities and Application as Wheat Biofertilizer under Drought Conditions: An Environmental Contribution.

The search for methanotrophs as plant-growth-promoting rhizobacteria (PGPR) presents an important contribution to mitigating the impact of global warming by restoring the natural soil potential for consuming methane while benefiting plants during droughts. Our in silico simulations suggest that water, produced as a byproduct of methane oxidation, can satisfy the cell growth requirement. In addition to water, methanotrophs can produce metabolites that stimulate plant growth. Considering this, we proposed that applying methanotrophs as PGPR can alleviate the effect of droughts on crops, while stimulating atmospheric methane consumption. In this work, we isolated a series of methanotrophic communities from the rhizospheres of different crops, including Italian sweet pepper and zucchini, using an atmosphere enriched with pure methane gas, to determine their potential for alleviating drought stress in wheat plants. Subsequently, 23 strains of nonmethanotrophic bacteria present in the methanotrophic communities were isolated and characterized. We then analyzed the contribution of the methane-consuming consortia to the improvement of plant growth under drought conditions, showing that some communities contributed to increases in the wheat plants' lengths and weights, with statistically significant differences according to ANOVA models. Furthermore, we found that the presence of methane gas can further stimulate the plant-microbe interactions, resulting in larger plants and higher drought tolerance.

High-Throughput Microbial Community Analyses to Establish a Natural Fungal and Bacterial Consortium from Sewage Sludge Enriched with Three Pharmaceutical Compounds.

Emerging and unregulated contaminants end up in soils via stabilized/composted sewage sludges, paired with possible risks associated with the development of microbial resistance to antimicrobial agents or an imbalance in the microbial communities. An enrichment experiment was performed, fortifying the sewage sludge with carbamazepine, ketoprofen and diclofenac as model compounds, with the aim to obtain strains with the capability to transform these pollutants. Culturable microorganisms were obtained at the end of the experiment. Among fungi, Cladosporium cladosporioides, Alternaria alternata and Penicillium raistrickii showed remarkable degradation rates. Population shifts in bacterial and fungal communities were also studied during the selective pressure using Illumina MiSeq. These analyses showed a predominance of Ascomycota (Dothideomycetes and Aspergillaceae) and Actinobacteria and Proteobacteria, suggesting the possibility of selecting native microorganisms to carry out bioremediation processes using tailored techniques.

Seed-Encapsulation of Desiccation-Tolerant Microorganisms for the Protection of Maize from Drought: Phenotyping Effects of a New Dry Bioformulation.

Droughts and high temperatures deeply affect crop production. The use of desiccation-tolerant (or xerotolerant) microorganisms able to protect plants from droughts represents a promising alternative. These xerotolerant microorganisms have previously been used to modulate plant responses and improve their tolerance to drought. In addition, these microorganisms could be stored and used in dry formats, which would improve their viability and resilience at a much lower cost than current market alternatives. In the present study we analyze the possibility of using strains of xerotolerant Actinobacteria in encapsulated format on seeds. Under this formulation, we carried out greenhouse with farming soil with maize plants. Under greenhouse conditions, the plants showed greater resistance to drought, as well as increased growth and production yield, but not as well in field trials. This alternative could represent a useful tool to improve water efficiency in crops for drought-affected areas or affected by water scarcity.

Plant Beneficial Deep-Sea Actinobacterium, Dermacoccus abyssi MT1.1T Promote Growth of Tomato (Solanum lycopersicum) under Salinity Stress.

Salt stress is a serious agricultural problem threatens plant growth and development resulted in productivity loss and global food security concerns. Salt tolerant plant growth promoting actinobacteria, especially deep-sea actinobacteria are an alternative strategy to mitigate deleterious effects of salt stress. In this study, we aimed to investigate the potential of deep-sea Dermacoccus abyssi MT1.1T to mitigate salt stress in tomato seedlings and identified genes related to plant growth promotion and salt stress mitigation. D. abyssi MT1.1T exhibited plant growth promoting traits namely indole-3-acetic acid (IAA) and siderophore production and phosphate solubilization under 0, 150, 300, and 450 mM NaCl in vitro. Inoculation of D. abyssi MT1.1T improved tomato seedlings growth in terms of shoot length and dry weight compared with non-inoculated seedlings under 150 mM NaCl. In addition, increased total soluble sugar and total chlorophyll content and decreased hydrogen peroxide content were observed in tomato inoculated with D. abyssi MT1.1T. These results suggested that this strain mitigated salt stress in tomatoes via osmoregulation by accumulation of soluble sugars and H2O2 scavenging activity. Genome analysis data supported plant growth promoting and salt stress mitigation potential of D. abyssi MT1.1T. Survival and colonization of D. abyssi MT1.1T were observed in roots of inoculated tomato seedlings. Biosafety testing on D. abyssi MT1.1T and in silico analysis of its whole genome sequence revealed no evidence of its pathogenicity. Our results demonstrate the potential of deep-sea D. abyssi MT1.1T to mitigate salt stress in tomato seedlings and as a candidate of eco-friendly bio-inoculants for sustainable agriculture.

Impacts of Agriculture on the Environment and Soil Microbial Biodiversity.

Agriculture represents an important mechanism in terms of reducing plant, animal, and microbial biodiversity and altering the environment. The pressure to cope with the increasing food demands of the human population has intensified the environmental impact, and alternative ways to produce food are required in order to minimize the decrease in biodiversity. Conventional agricultural practices, such as floods and irrigation systems; the removal of undesired vegetation by fires, tilling, and plowing; the use of herbicides, fertilizers, and pesticides; and the intensification of these practices over the last 50 years, have led to one of the most important environmental threats-a major loss of biodiversity. In this study, we review the impact that agriculture and its intensification have had on the environment and biodiversity since its invention. Moreover, we demonstrate how these impacts could be reduced through the use of microorganisms as biostimulants.

Deep-Sea Actinobacteria Mitigate Salinity Stress in Tomato Seedlings and Their Biosafety Testing.

Soil salinity is an enormous problem affecting global agricultural productivity. Deep-sea actinobacteria are interesting due to their salt tolerance mechanisms. In the present study, we aim to determine the ability of deep-sea Dermacoccus (D. barathri MT2.1T and D. profundi MT2.2T) to promote tomato seedlings under 150 mM NaCl compared with the terrestrial strain D. nishinomiyaensis DSM20448T. All strains exhibit in vitro plant growth-promoting traits of indole-3-acetic acid production, phosphate solubilization, and siderophore production. Tomato seedlings inoculated with D. barathri MT2.1T showed higher growth parameters (shoot and root length, dry weight, and chlorophyll content) than non-inoculated tomato and the terrestrial strain under 150 mM NaCl. In addition, hydrogen peroxide (H2O2) in leaves of tomatoes inoculated with deep-sea Dermacoccus was lower than the control seedlings. This observation suggested that deep-sea Dermacoccus mitigated salt stress by reducing oxidative stress caused by hydrogen peroxide. D. barathri MT2.1T showed no harmful effects on Caenorhabditis elegans, Daphnia magna, Eisenia foetida, and Escherichia coli MC4100 in biosafety tests. This evidence suggests that D. barathri MT2.1T would be safe for use in the environment. Our results highlight the potential of deep-sea Dermacoccus as a plant growth promoter for tomatoes under salinity stress.

The Effects of Plant-Associated Bacterial Exopolysaccharides on Plant Abiotic Stress Tolerance.

Plant growth-promoting rhizobacteria (PGPR) are beneficial soil microorganisms that can stimulate plant growth and increase tolerance to biotic and abiotic stresses. Some PGPR are capable of secreting exopolysaccharides (EPS) to protect themselves and, consequently, their plant hosts against environmental fluctuations and other abiotic stresses such as drought, salinity, or heavy metal pollution. This review focuses on the enhancement of plant abiotic stress tolerance by bacterial EPS. We provide a comprehensive summary of the mechanisms through EPS to alleviate plant abiotic stress tolerance, including salinity, drought, temperature, and heavy metal toxicity. Finally, we discuss how these abiotic stresses may affect bacterial EPS production and its role during plant-microbe interactions.

Dealing with water stress and microbial preservation.

The relevance of preserving microorganisms has been well accepted for several decades. Interest is now shifting towards investigating adequate preservation methods to improve microbial survival rates and to preserve new taxa of previously considered unculturable microorganisms. In addition, a growing interest in preserving fragile microbial consortia or communities with biotechnological interest motivates the improvement of preservation methods. In the present study, we reviewed the effect of water availability in microbial diversity shift. We describe the effect of drought on microorganisms at the molecular level and their molecular responses to this life-threatening challenge focusing on the production of xeroprotectants. We also review the interspecies interactions of those drought-tolerant microorganisms with other sensitive organisms including neighbouring prokaryotes and eukaryotes such as plants, and the potential role of these microorganisms at determining the ecological composition of stressed environments. We emphasize the importance of applying the knowledge derived from the molecular mechanisms used by desiccation-tolerant microorganisms for the improvement of the preservation techniques. An overview of the current and newer techniques for preserving microorganisms and microbial communities is provided. The biotechnological interest in preserving pure cultures, microbial consortia and communities is also discussed.

Proteome Comparison Between Natural Desiccation-Tolerant Plants and Drought-Protected Caspicum annuum Plants by Microbacterium sp. 3J1.

Desiccation-tolerant plants are able to survive for extended periods of time in the absence of water. The molecular understanding of the mechanisms used by these plants to resist droughts can be of great value for improving drought tolerance in crops. This understanding is especially relevant in an environment that tends to increase the number and intensity of droughts. The combination of certain microorganisms with drought-sensitive plants can improve their tolerance to water scarcity. One of these bacteria is Microbacterium sp. 3J1, an actinobacteria able to protect pepper plants from drought. In this study, we supplemented drought-tolerant and drought-sensitive plant rhizospheres with Microbacterium sp. 3J1 and analyzed their proteomes under drought to investigate the plant-microbe interaction. We also compare this root proteome with the proteome found in desiccation-tolerant plants. In addition, we studied the proteome of Microbacterium sp. 3J1 subjected to drought to analyze its contribution to the plant-microbe interaction. We describe those mechanisms shared by desiccation-tolerant plants and sensitive plants protected by microorganisms focusing on protection against oxidative stress, and production of compatible solutes, plant hormones, and other more specific proteins.

Units for vigilance of emerging diseases based on wastewater treatment plants (WWTP-UVED).

Pandemics deeply affect the health and economy of the world population. A precise determination of affected communities is of great importance to establish containment measures and reduce the economic impact. Here, we propose the development of Units for Vigilance of Emerging Diseases based on the screening of pathogens released to wastewater treatment plants to follow the spread of the infectious agent to determine the location of infected people.

The involvement of McpB chemoreceptor from Pseudomonas aeruginosa PAO1 in virulence.

Pseudomonas aeruginosa is an opportunistic human pathogen causing infections in a variety of plant and animal hosts. The gene mcpB, part of the chemosensory gene cluster II, encodes a soluble chemoreceptor whose function remains unknown. Previous studies show that the cheB2 gene, also located in the chemosensory cluster II, is involved in a specific response during infection and it is required for full pathogenicity of P. aeruginosa. To determine whether the McpB (or Aer2) chemoreceptor is involved in virulence processes, we generated a mcpB mutant and tested its phenotype using a virulence-measuring system. This system was developed by our group and is based on different bioassays using organisms living at different soil trophic levels, including microbial, nematode, arthropod, annelid, and plant model systems. The deletion of mcpB resulted in an attenuation of bacterial virulence in different infection models, and wild-type virulence was restored following genetic complementation of the mutant strain. Our study indicates that the McpB chemoreceptor is linked to virulence processes and may constitute the basis for the development of alternative strategies against this pathogen.

Microbial Production of Ethanol From Sludge Derived From an Urban Wastewater Treatment Plant.

A collection of lipase-producing microorganisms was isolated from sludge derived from an urban wastewater treatment plant. The microorganisms with the highest levels of lipase activity were selected in order to use triglycerides present in the sludge effectively and were then transformed with pdc:adhB genes for the production of ethanol. The transgenic strains showed high growth rates in diluted sludge and produced lipase protein in order to utilize fat present in the sludge, which provides an abundant source of carbon. Using sludge derived from treated wastewater as nutrient source, ethanol was produced by certain transgenic species belonging to the genera Proteus. Different forms of sludge were tested for maximal ethanol production, with dehydrated sludge being found to produce the best performance.

Anthracene drives sub-cellular proteome-wide alterations in the degradative system of Penicillium oxalicum.

Polycyclic aromatic hydrocarbons (PAHs) are widely distributed in polluted environments and are included in the priority list of toxic compounds. Previous studies have shown that the fungus Penicillium oxalicum, isolated from a hydrocarbon-polluted pond, has a great capability to transform different PAHs in short periods under submerged fermentation conditions. Although cytochrome p450s (CYPs) seems to be the main responsible enzyme in this process, changes in proteome profile remains poorly understood. The aim of this work was to characterise molecular disturbances in the cytosolic and microsomal sub-proteomes of P. oxalicum by applying two-dimensional (2D) gel electrophoresis and label-free quantitative proteomics during anthracene biodegradation. Our results showed that by using 2D-gels, 10 and 8 differential proteins were over-expressed in the cytosolic and microsomal fractions, respectively. Most of them were related to stress response. Shotgun proteomics allowed the identification of 158 and 174 unique protein species that differentially accumulated during anthracene biotransformation, such as CYPs, epoxide hydrolases and transferases enzymes, belonging to Phase I and Phase II of the metabolism of xenobiotics, contributing to the anthracene biodegradation pathway. These results confirm the biological significance of ascomycetes fungi the rol of CYPs on biodegradation and the need of a deeper knowledge on fungal proteomics for the application of the appropriate microorganisms in biodegradation processes.

Protection of Pepper Plants from Drought by Microbacterium sp. 3J1 by Modulation of the Plant's Glutamine and α-ketoglutarate Content: A Comparative Metabolomics Approach.

Drought tolerance of plants such as tomato or pepper can be improved by their inoculation with rhizobacteria such as Microbacterium sp. 3J1. This interaction depends on the production of trehalose by the microorganisms that in turn modulate the phyto-hormone profile of the plant. In this work we describe the characterization of metabolic changes during the interaction of pepper plants with Microbacterium sp. 3J1 and of the microorganism alone over a period of drought. Our main findings include the observation that the plant responds to the presence of the microorganism by changing the C and N metabolism based on its glutamine and α-ketoglutarate content, these changes contribute to major changes in the concentration of molecules involved in the balance of the osmotic pressure. These include sugars and amino-acids; the concentration of antioxidant molecules, of metabolites involved in the production of phytohormones like ethylene, and of substrates used for lignin production such as ferulic and sinapic acids. Most of the altered metabolites of the plant when inoculated with Microbacterium sp. 3J1 in response to drought coincided with the profile of altered metabolites in the microorganism alone when subjected to drought, pointing to a response by which the plant relies on the microbe for the production of such metabolites. To our knowledge this is the first comparative study of the microbe colonized-plant and microbe alone metabolomes under drought stress.

Plant Drought Tolerance Enhancement by Trehalose Production of Desiccation-Tolerant Microorganisms.

A collection of desiccation-tolerant xeroprotectant-producing microorganisms was screened for their ability to protect plants against drought, and their role as plant growth-promoting rhizobacteria was investigated in two different crops (tomato and pepper). The most commonly described biochemical mechanisms for plant protection against drought by microorganisms including the production of phytohormones, antioxidants and xeroprotectants were analyzed. In particular, the degree of plant protection against drought provided by these microorganisms was characterized. After studying the findings and comparing them with results of the closest taxonomic relatives at the species and strain levels, we propose that trehalose produced by these microorganisms is correlated with their ability to protect plants against drought. This proposal is based on the increased protection of plants against drought by the desiccation-sensitive microorganism Pseudomonas putida KT2440, which expresses the otsAB genes for trehalose biosynthesis in trans.

A New Physiological Role for the DNA Molecule as a Protector against Drying Stress in Desiccation-Tolerant Microorganisms.

The DNA molecule is associated with the role of encoding information required to produce RNA which is translated into proteins needed by the cell. This encoding involves information transmission to offspring or to other organisms by horizontal transfer. However, despite the abundance of this molecule in both the cell and the environment, its physiological role seems to be restricted mainly to that of a coding and inheritance molecule. In this paper, we report a new physiological role for the DNA molecule as involved in protection against desiccation, in addition to its well-established main information transfer and other recently reported functions such as bio-film formation in eDNA form. Desiccation-tolerant microorganisms such as Microbacterium sp. 3J1 significantly upregulate genes involved in DNA synthesis to produce DNA as part of their defensive mechanisms to protect protein structures and functions from drying according to RNA-seq analysis. We have observed the intracellular overproduction of DNA in two desiccation-tolerant microorganisms, Microbacterium sp. 3J1 and Arthrobacter siccitolerans 4J27, in response to desiccation signals. In addition, this conclusion can be made from our observations that synthetic DNA protects two proteins from drying and when part of a xeroprotectant preparation, DNA from various organisms including desiccation-sensitive species, does the same. Removal of DNA by nuclease treatment results in absence of this additive protective effect. We validated this role in biochemical and biophysical assays in proteins and occurs in trans even with short, single chains of synthetically produced DNA.

Genome Sequence of Microbacterium sp. Strain 3J1, a Highly Desiccation-Tolerant Bacterium That Promotes Plant Growth.

The genome sequence for Microbacterium sp. strain 3J1, a desiccation-tolerant organism isolated from the Nerium oleander rhizosphere, is reported here. The genome is estimated to be approximately 3.5 Mb in size, with an average G+C content of 67.7% and a predicted number of protein-coding sequences of 3,310.

Genome Sequence of Arthrobacter koreensis 5J12A, a Plant Growth-Promoting and Desiccation-Tolerant Strain.

Arthrobacter koreensis 5J12A is a desiccation-tolerant organism isolated from the Nerium oleander rhizosphere. Here, we report its genome sequence, which may shed light on its role in plant growth promotion. This is believed to be the first published genome of a desiccation-tolerant plant growth promoter from the genus Arthrobacter.

Genome Sequence of Rhodococcus sp. 4J2A2, a Desiccation-Tolerant Bacterium Involved in Biodegradation of Aromatic Hydrocarbons.

The genome sequence for Rhodococcus sp. 4J2A2, a newly described desiccation-tolerant strain that removes aromatic hydrocarbons, is reported here. The genome is estimated to be around 7.5 Mb in size, with an average G+C content of 60.77% and a predicted number of protein-coding sequences of 6,354.