Functional and structural responses of a halophilic consortium to oily sludge during biodegradation.

Biotreatment of oily sludge and the involved microbial communities, particularly in saline environments, have been rarely investigated. We enriched a halophilic bacterial consortium (OS-100) from petroleum refining oily sludge, which degraded almost 86% of the aliphatic hydrocarbon (C10-C30) fraction of the oily sludge within 7 days in the presence of 100 g/L NaCl. Two halophilic hydrocarbon-degrading bacteria related to the genera Chromohalobacter and Halomonas were isolated from the OS-100 consortium. Hydrocarbon degradation by the OS-100 consortium was relatively higher compared to the isolated bacteria, indicating potential synergistic interactions among the OS-100 community members. Exclusion of FeCl2, MgCl2, CaCl2, trace elements, and vitamins from the culture medium did not significantly affect the hydrocarbon degradation efficiency of the OS-100 consortium. To the contrary, hydrocarbon biodegradation dropped from 94.1 to 54.4% and 5% when the OS-100 consortium was deprived from phosphate and nitrogen sources in the culture medium, respectively. Quantitative PCR revealed that alkB gene expression increased up to the 3rd day of incubation with 11.277-fold, consistent with the observed increments in hydrocarbon degradation. Illumina-MiSeq sequencing of 16 S rRNA gene fragments revealed that the OS-100 consortium was mainly composed of the genera Halomonas, Idiomarina, Alcanivorax and Chromohalobacter. This community structure changed depending on the culturing conditions. However, remarkable changes in the community structure were not always associated with remarkable shifts in the hydrocarbonoclastic activity and vice versa. The results show that probably synergistic interactions between community members and different subpopulations of the OS-100 consortium contributed to salinity tolerance and hydrocarbon degradation.

Rhizosphere Microbiome of Arid Land Medicinal Plants and Extra Cellular Enzymes Contribute to Their Abundance.

Revealing the unexplored rhizosphere microbiome of plants in arid environments can help in understanding their interactions between microbial communities and plants during harsh growth conditions. Here, we report the first investigation of rhizospheric fungal and bacterial communities of Adenium obesum, Aloe dhufarensis and Cleome austroarabica using next-generation sequencing approaches. A. obesum and A. dhufarensis grows in dry tropical and C. austroarabica in arid conditions of Arabian Peninsula. The results indicated the presence of 121 fungal and 3662 bacterial operational taxonomic units (OTUs) whilst microbial diversity was significantly high in the rhizosphere of A. obesum and A. dhufarensis and low in C. austroarabica. Among fungal phyla, Ascomycota and Basidiomycota were abundantly associated within rhizospheres of all three plants. However, Mucoromycota was only present in the rhizospheres of A. obesum and A. dhufarensis, suggesting a variation in fungal niche on the basis of host and soil types. In case of bacterial communities, Actinobacteria, Proteobacteria, Bacteroidetes, Planctomycetes, Acidobacteria, and Verrucomicrobia were predominant microbial phyla. These results demonstrated varying abundances of microbial structure across different hosts and locations in arid environments. Rhizosphere's extracellular enzymes analysis revealed varying quantities, where, glucosidase, cellulase, esterase, and 1-aminocyclopropane-1-carboxylate deaminase were significantly higher in the rhizosphere of A. dhufarensis, while phosphatase and indole-acetic acid were highest in the rhizosphere of A. obesum. In conclusion, current findings usher for the first time the core microbial communities in the rhizospheric regions of three arid plants that vary greatly with location, host and soil conditions, and suggest the presence of extracellular enzymes could help in maintaining plant growth during the harsh environmental conditions.

Anti-proliferative potential of cyclotetrapeptides from Bacillus velezensis RA5401 and their molecular docking on G-Protein-Coupled Receptors.

Elucidation of bioactive chemical compounds from rhizobacteria is highly utilized in pharmaceuticals and naturopathy, due to their health benefits to human and plants. In current study, four cyclopeptides along with one phenyl amide were isolated from the ethyl acetate extract of Bacillus velezensis sp. RA5401. Their structures were determined and characterized as cycle (L-prolyl-L-leucyl)2 (1), cyclo (L-prolyl-l-valine)2 (2), cycle (L-phenylanalyl-L-propyl)2 (3), cyclo (D-pro-L-tyr-L-pro-L-tyr)2 (4) and N-(2-phenylethyl)acetamide (5) on the basis of electron spray ionization mass spectrometry (ESI-MS), nuclear magnetic resonance (NMR) techniques and comparison with the literature data. The five compounds have been isolated for the first time from this species. The effect of various concentrations of these compounds on the proliferation of MDA-MB-231 breast cancer cells was examined. It was found that 1 and 2 induced concentration-independent anti-proliferative effects, while 3, 4 and 5 inhibited cancer cell proliferation in a concentration-dependent manner. Furthermore, to determine the suitable binding targets of these compounds within cancer cell line, detailed target prediction and comparative molecular-docking studies were performed. The compounds 1 and 2 hit intracellular anti-cancer targets of proteases family, while compounds 3, 4 and 5 interacted with different membrane receptors of G-Protein-Coupled Receptors (GPCRs). In conclusion, the Bacillus velezensis RA5401 can be an ideal strain to produce anti-proliferative constituents at industrial scale.

EDTA addition enhances bacterial respiration activities and hydrocarbon degradation in bioaugmented and non-bioaugmented oil-contaminated desert soils.

The low number and activity of hydrocarbon-degrading bacteria and the low solubility and availability of hydrocarbons hamper bioremediation of oil-contaminated soils in arid deserts, thus bioremediation treatments that circumvent these limitations are required. We tested the effect of Ethylenediaminetetraacetic acid (EDTA) addition, at different concentrations (i.e. 0.1, 1 and 10 mM), on bacterial respiration and biodegradation of Arabian light oil in bioaugmented (i.e. with the addition of exogenous alkane-degrading consortium) and non-bioaugmented oil-contaminated desert soils. Post-treatment shifts in the soils' bacterial community structure were monitored using MiSeq sequencing. Bacterial respiration, indicated by the amount of evolved CO2, was highest at 10 mM EDTA in bioaugmented and non-bioaugmented soils, reaching an amount of 2.2 ± 0.08 and 1.6 ± 0.02 mg-CO2 g(-1) after 14 days of incubation, respectively. GC-MS revealed that 91.5% of the C14-C30 alkanes were degraded after 42 days when 10 mM EDTA and the bacterial consortium were added together. MiSeq sequencing showed that 78-91% of retrieved sequences in the original soil belonged to Deinococci, Alphaproteobacteria, Gammaproteobacteia and Bacilli. The same bacterial classes were detected in the 10 mM EDTA-treated soils, however with slight differences in their relative abundances. In the bioaugmented soils, only Alcanivorax sp. MH3 and Parvibaculum sp. MH21 from the exogenous bacterial consortium could survive until the end of the experiment. We conclude that the addition of EDTA at appropriate concentrations could facilitate biodegradation processes by increasing hydrocarbon availability to microbes. The addition of exogenous oil-degrading bacteria along with EDTA could serve as an ideal solution for the decontamination of oil-contaminated desert soils.

Diversity of bacterial communities along a petroleum contamination gradient in desert soils.

Microbial communities in oil-polluted desert soils have been rarely studied compared to their counterparts from freshwater and marine environments. We investigated bacterial diversity and changes therein in five desert soils exposed to different levels of oil pollution. Automated rRNA intergenic spacer (ARISA) analysis profiles showed that the bacterial communities of the five soils were profoundly different (analysis of similarities (ANOSIM), R = 0.45, P < 0.0001) and shared less than 20 % of their operational taxonomic units (OTUs). OTU richness was relatively higher in the soils with the higher oil pollution levels. Multivariate analyses of ARISA profiles revealed that the microbial communities in the S soil, which contains the highest level of contamination, were different from the other soils and formed a completely separate cluster. A total of 16,657 ribosomal sequences were obtained, with 42-89 % of these sequences belonging to the phylum Proteobacteria. While sequences belonging to Betaproteobacteria, Gammaproteobacteria, Bacilli, and Actinobacteria were encountered in all soils, sequences belonging to anaerobic bacteria from the classes Deltaproteobacteria, Clostridia, and Anaerolineae were only detected in the S soil. Sequences belonging to the genus Terriglobus of the class Acidobacteria were only detected in the B3 soil with the lowest level of contamination. Redundancy analysis (RDA) showed that oil contamination level was the most determinant factor that explained variations in the microbial communities. We conclude that the exposure to different levels of oil contamination exerts a strong selective pressure on bacterial communities and that desert soils are rich in aerobic and anaerobic bacteria that could potentially contribute to the degradation of hydrocarbons.

Resistance of Lophelia pertusa to coverage by sediment and petroleum drill cuttings.

In laboratory experiments, the cold-water coral Lophelia pertusa was exposed to settling particles. The effects of reef sediment, petroleum drill cuttings and a mix of both, on the development of anoxia at the coral surface were studied using O2, pH and H2S microsensors and by assessing coral polyp mortality. Due to the branching morphology of L. pertusa and the release of coral mucus, accumulation rates of settling material on coral branches were low. Microsensors detected H2S production in only a few samples, and sulfate reduction rates of natural reef sediment slurries were low (<0.3 nmol S cm(-3) d(-1)). While the exposure to sediment clearly reduced the coral's accessibility to oxygen, L. pertusa tolerated both partial low-oxygen and anoxic conditions without any visible detrimental short-term effect, such as tissue damage or death. However, complete burial of coral branches for >24 h in reef sediment resulted in suffocation.

Bacterial diversity of a cyanobacterial mat degrading petroleum compounds at elevated salinities and temperatures.

Cyanobacterial mats of the Arabian Gulf coast of Saudi Arabia experience extreme conditions of temperature and salinity. Because they are exposed to continuous oil pollution, they form ideal models for biodegradation under extreme conditions. We investigated the bacterial diversity of these mats using denaturing gradient gel electrophoresis and 16S rRNA cloning, and tested their potential to degrade petroleum compounds at various salinities (fresh water to 16%) and temperatures (5 to 50 degrees C). Cloning revealed that c. 15% of the obtained sequences were related to unknown, possibly novel bacteria. Bacteria belonging to Beta-, Gamma- and Deltaproteobacteria, Cytophaga-Flavobacterium-Bacteroides group and Spirochetes, were detected. The biodegradation of petroleum compounds at different salinities by mat microorganisms showed that pristine and n-octadecane were optimally degraded at salinities between 5 and 12% (weight per volume NaCl) whereas the optimum degradation of phenanthrene and dibenzothiophene was at 3.5% salinity. The latter compounds were also degradable at 8% salinity. The same compounds were degraded at temperatures between 15 and 40 degrees C but not at 5 and 50 degrees C. The optimum temperature of degradation was 28-40 degrees C for both aliphatics and aromatics. We conclude that the studied microbial mats from Saudi Arabia are rich in novel halotolerant and thermotolerant microorganisms with the potential to degrade petroleum compounds at elevated salinities and temperatures.

Microbial diversity of a heavily polluted microbial mat and its community changes following degradation of petroleum compounds.

We studied the microbial diversity of benthic cyanobacterial mats inhabiting a heavily polluted site in a coastal stream (Wadi Gaza) and monitored the microbial community response induced by exposure to and degradation of four model petroleum compounds in the laboratory. Phormidium- and Oscillatoria-like cyanobacterial morphotypes were dominant in the field. Bacteria belonging to different groups, mainly the Cytophaga-Flavobacterium-Bacteriodes group, the gamma and beta subclasses of the class Proteobacteria, and the green nonsulfur bacteria, were also detected. In slurry experiments, these communities efficiently degraded phenanthrene and dibenzothiophene completely in 7 days both in the light and in the dark. n-Octadecane and pristane were degraded to 25 and 34% of their original levels, respectively, within 7 days, but there was no further degradation until 40 days. Both cyanobacterial and bacterial communities exhibited noticeable changes concomitant with degradation of the compounds. The populations enriched by exposure to petroleum compounds included a cyanobacterium affiliated phylogenetically with Halomicronema. Bacteria enriched both in the light and in the dark, but not bacteria enriched in any of the controls, belonged to the newly described Holophaga-Geothrix-Acidobacterium phylum. In addition, another bacterial population, found to be a member of green nonsulfur bacteria, was detected only in the bacteria treated in the light. All or some of the populations may play a significant role in metabolizing the petroleum compounds. We concluded that the microbial mats from Wadi Gaza are rich in microorganisms with high biodegradative potential.

Degradation of petroleum model compounds immobilized on clay by a hypersaline microbial mat.

In this study the degradation of hydrophobic petroleum model compounds (phenanthrene, pristane, octadecane and dibenzothiophene) added to a submersed hypersaline microbial mat was investigated. Montmorillonite with an artificially altered, hydrophobic surface was used as carrier material, forming an organo-clay complex (OCC) with the attached mixture of petroleum model compounds. 6 mg/cm2 OCC were applied to cyanobacterial mat pieces, containing approximately 33.3 microg/mg OCC of each compound. The degradation experiment was performed under controlled laboratory conditions and accompanied by chemical analyses by GC/GC-MS, molecular analyses by PCR and DGGE as well as functional analyses by microsensor measurements of oxygen, photosynthesis, sulfide, pH and light. All applied model compounds were degraded, but residues were still present after 18 weeks. The aromatic compounds phenanthrene (5.1 microg/mg OCC) and dibenzothiophene (4.3 microg/mg OCC) were preferentially degraded compared to the alkanes pristane (12.4 microg/mg OCC) and n-octadecane (13.4 microg/mg OCC). Metabolic changes during the degradation process could not be detected by microsensor measurements. The molecular population analyses did not reveal any significant community changes concomitant with the decrease of the petroleum model compounds. We conclude, that the pristine mats represent an intact, robust ecosystem in which the enzymatic requirements for the degradation of the applied pollutants exist. The slow degradation process did not affect the usual high internal turnover rates and did not favor a certain population in the community of the mats.