Bioremediation of oily hypersaline soil via autochthonous bioaugmentation with halophilic bacteria and archaea.

Kuwaiti hypersaline soil samples were contaminated with 5 % (w/w) weathered Kuwaiti light crude oil and bioaugmented with autochthonous halophilic hydrocarbonoclastic archaeal and bacterial strains, two each, individually and as consortia. Residual oil contents were determined, and microbial communities were analyzed by culture-dependent and culture-independent approaches initially and seasonally for one year. After one year of the bioremediation process, the mean oil degradation rate was similar across all treated soils including the controlled unbioaugmented one. Oil hydrocarbons were drastically reduced in all soil samples with values ranging from 82.7 % to 93 %. During the bioremediation process, the number of culturable oil-degrading bacteria increased to a range of 142 to 344 CFUx104 g-1 after 12 months of bioaugmentation. Although culture-independent analysis showed a high proportion of inoculants initially, none could be cultured throughout the bioremediation procedure. Within a year, microbial communities changed continually, and 33 species of halotolerant/halophilic hydrocarbonoclastic bacteria were isolated and identified belonged mainly to the three major bacterial phyla Actinobacteria, Proteobacteria, and Firmicutes. The archaeal phylum Halobacterota represented <1 % of the microbial community's relative abundance, which explains why none of its members were cultured. Improving the biodegradability of an already balanced environment by autochthonous bioaugmentation is more involved than just adding the proper oil degraders. This study emphasizes the possibility of a relatively large resistant population, a greater diversity of oil-degrading microorganisms, and the highly selective impacts of oil contamination on hypersaline soil bacterial communities.

Contribution of soil bacteria to the atmosphere across biomes.

The dispersion of microorganisms through the atmosphere is a continual and essential process that underpins biogeography and ecosystem development and function. Despite the ubiquity of atmospheric microorganisms globally, specific knowledge of the determinants of atmospheric microbial diversity at any given location remains unresolved. Here we describe bacterial diversity in the atmospheric boundary layer and underlying soil at twelve globally distributed locations encompassing all major biomes, and characterise the contribution of local and distant soils to the observed atmospheric community. Across biomes the diversity of bacteria in the atmosphere was negatively correlated with mean annual precipitation but positively correlated to mean annual temperature. We identified distinct non-randomly assembled atmosphere and soil communities from each location, and some broad trends persisted across biomes including the enrichment of desiccation and UV tolerant taxa in the atmospheric community. Source tracking revealed that local soils were more influential than distant soil sources in determining observed diversity in the atmosphere, with more emissive semi-arid and arid biomes contributing most to signatures from distant soil. Our findings highlight complexities in the atmospheric microbiota that are relevant to understanding regional and global ecosystem connectivity.

Corrigendum: Cross-Bioaugmentation Among Four Remote Soil Samples Contaminated With Oil Exerted Just Inconsistent Effects on Oil-Bioremediation.

[This corrects the article DOI: 10.3389/fmicb.2019.02827.].

Bioaugmentation failed to enhance oil bioremediation in three soil samples from three different continents.

Soil samples from Kuwait, Lebanon, Egypt and Germany were polluted with 3% crude oil. Series of samples were left unbioaugmented, others were bioaugmented with Kuwaiti desert soil with a long history of oil pollution and still others with Kuwaiti marine biofouling material. In the samples from Kuwait, Egypt, and Germany, bioaugmentation did not enhance oil removal, whereas it did in the sample from Lebanon. Taxa from the desert-soil bioaugmented batches, but none of those from the biofouling-material bioaugmented ones, succeeded in colonizing the four studied soils. The dynamics of the hydrocarbonoclastic communities during bioremediation were monitored. Those communities differed in composition, not only according to the type of soil, but also for the same soil; at various phases of bioremediation. Although each soil seemed to have its characteristic microflora, they all were similar in harboring lower and higher actinomycetes and pseudomonads in addition to many other taxa. None of the taxa prevailed through all phases of bioremediation. The most powerful isolate in oil-removal; was Rhodococcus erythropolis (Germany), and the weakest was Arthrobacter phenanthrenivorans (Lebanon). The pure hydrocarbonoclastic isolates tolerated unusually high oil concentrations, up to 30%.

Cross-Bioaugmentation Among Four Remote Soil Samples Contaminated With Oil Exerted Just Inconsistent Effects on Oil-Bioremediation.

Soil samples were collected from Kuwait, Lebanon, Egypt, and Germany, and artificially polluted with 3% (w/w) crude oil. Cross-bioaugmentation was done among them, and the oil-consumption and the constituent communities of hydrocarbonoclastic bacteria were monitored periodically through 6 months. The results showed that cross-bioaugmentation did not bring about reproducible effects on oil-removal in the four soils. After 6 months, oil-removal values reached between 82 and 95% in most of the samples including the unbioaugmented controls. The numbers of hydrocarbonoclastic bacteria showed significant increases followed by significant decreases during the course of bioremediation also in the unbioaugmented controls. In most cases, the inoculated bacterial taxa failed to colonize the soils, and oil-removal was achieved mainly by the native (autochthonous) soil bacterial communities. those belonged to the genera Mycolicibacterium, Mycobacterium, Xanthobacter, Pseudoxanthomonas, Pseudomonas, Zavarzinia, and others. The microbial communities in the four soils also comprised nitrogen fixing bacteria belonging to the genera Gordonia, Rhizobium, Kocuria, and Azospirillum. Such diazotrophs are known to enrich the soils with fixed nitrogen and thus, contribute to enhancing the microbiological hydrocarbon-consumption. It was concluded that cross-bioaugmentation leads to unpredictable and inconsistent effects on oil removal. Therefore, it could not beregarded as the technology of choice for oil-bioremediation.

Gelatinizing oil in water and its removal via bacteria inhabiting the gels.

When crude oil samples were shaken (200 rpm) in seawater samples from the Arabian Gulf at 30 °C, usually oil-gels were produced spontaneously leaving the water quite clear. The gelators could probably be based on cholesteryl derivatives. Microscopic examination of the established gels revealed nanofibrellar structures similar to those described by earlier workers for artificially synthesized gelators. Communities of bacteria including prosthetic and stalked members as well as oil-degrading bacteria were recorded in such gels. Chemical analysis revealed that 88.5% of the oil entrapped by gelation was biodegraded after 40 days at 30 °C. Individual bacterial species isolated from the oil-gels biodegraded in batch cultures between 17.8 and 33.3% of the oil added at time zero in 12 days at 30 °C. Gelation is a promising approach, not only for clean, physical removal of oil spilled in aquatic habitats, as so far suggested, but also in its effective microbiological biodegradation, as the current study revealed.

Calcium (II) - and dipicolinic acid mediated-biostimulation of oil-bioremediation under multiple stresses by heat, oil and heavy metals.

The oil-producing Arabian Gulf states have hot summer seasons of about 7-month in length. Therefore, environmental oil spills should be bioremediated by the activity of indigenous, hydrocarbonoclastic (hydrocarbon-degrading) microorganisms with optimum growth at about 50 °C. Soils in such arid countries harbor thermophilic bacteria, whose oil-consumption potential is enhanced by calcium (II) - and dipicolinic acid (DPA)-supplement. Those organisms are, however, subjected to additional stresses including toxic effects of heavy metals that may be associated with the spilled oil. Our study highlighted the resistance of indigenous, thermophilic isolates to the heavy metals, mercury (II), cadmium (II), arsenic (II) and lead (II) at 50 °C. We also detected the uptake of heavy metals by 15 isolates at 50 °C, and identified the merA genes coding for Hg2+-resistance in 4 of the studied Hg2+-resistant isolates. Hg2+ was the most toxic metal and the metal toxicity was commonly higher in the presence of oil. The addition of Ca2+ and DPA enhanced the Hg2+-resistance among most of the isolates at 50 °C. Crude oil consumption at 50 °C by 4 selected isolates was inhibited by the tested heavy metals. However, Ca2+ and DPA limited this inhibition and enhanced oil-consumption, which exceeded by far the values in the control cultures.

Capabilities and limitations of DGGE for the analysis of hydrocarbonoclastic prokaryotic communities directly in environmental samples.

Prokaryotic communities in pristine and oil-contaminated desert soil, seawater, and hypersaline coastal soil were analyzed using culture-dependent and culture-independent approaches. The former technique was the dilution-plating method. For the latter, total genomic DNA was extracted and the 16S rRNA genes were amplified using a universal bacterial primer pair and primer pairs specific for Actinobacteria, Gammaproteobacteria, and Archaea. The amplicons were resolved using denaturing gradient gel electrophoresis (DGGE) and sequenced, and the sequences were compared to those in GenBank. The plating method offered the advantages of capturing the targeted hydrocarbonoclastic microorganisms, counting them and providing cultures for further study. However, this technique could not capture more than a total of 15 different prokaryotic taxa. Those taxa belonged predominantly to the genera Alcanivorax, Pseudoxanthomonas, Bosea, Halomonas, and Marinobacter. The individual isolates in culture consumed between 19 and 50% of the available crude oil in 10 days. Although the culture-independent approach revealed much more microbial diversity, it was not problem-free. The subdivision primers exhibited satisfactory specificity, but they failed to capture all the available taxa. The universal bacterial primer pair ignored Actinobacteria altogether, although the primer pair specific for Actinobacteria captured many of them, for example, the genera Geodermatophilus, Streptomyces, Mycobacterium, Pontimonas, Rhodococcus, Blastococcus, Kocuria, and many others. Because most researchers worldwide use universal primers for PCR, this finding should be considered critically to avoid misleading interpretations.

Biofilm comprising phototrophic, diazotrophic, and hydrocarbon-utilizing bacteria: a promising consortium in the bioremediation of aquatic hydrocarbon pollutants.

Biofilms harboring simultaneously anoxygenic and oxygenic phototrophic bacteria, diazotrophic bacteria, and hydrocarbon-utilizing bacteria were established on glass slides suspended in pristine and oily seawater. Via denaturing gradient gel electrophoresis analysis on PCR-amplified rRNA gene sequence fragments from the extracted DNA from biofilms, followed by band amplification, biofilm composition was determined. The biofilms contained anoxygenic phototrophs belonging to alphaproteobacteria; pico- and filamentous cyanobacteria (oxygenic phototrophs); two species of the diazotroph Azospirillum; and two hydrocarbon-utilizing gammaproteobacterial genera, Cycloclasticus and Oleibacter. The coexistence of all these microbial taxa with different physiologies in the biofilm makes the whole community nutritionally self-sufficient and adequately aerated, a condition quite suitable for the microbial biodegradation of aquatic pollutant hydrocarbons.