Nitrate and oxygen significantly changed the abundance rather than structure of sulphate-reducing and sulphur-oxidising bacteria in water retrieved from petroleum reservoirs.

Sulphate-reducing bacteria (SRB) are the main culprits of microbiologically influenced corrosion in water-flooding petroleum reservoirs, but some sulphur-oxidising bacteria (SOB) are stimulated when nitrate and oxygen are injected, which control the growth of SRB. This study aimed to determine the distributions of SRB and SOB communities in injection-production systems and to analyse the responses of these bacteria to different treatments involving nitrate and oxygen. Desulfovibrio, Desulfobacca, Desulfobulbus, Sulfuricurvum and Dechloromonas were commonly detected via 16S rRNA gene sequencing. Still, no significant differences were observed for either the SRB or SOB communities between injection and production wells. Three groups of water samples collected from different sampling sites were incubated. Statistical analysis of functional gene (dsrB and soxB) clone libraries and quantitative polymerase chain reaction showed that the SOB community structures were more strongly affected by the nitrate and oxygen levels than SRB clustered according to the sampling site; moreover, both the SRB and SOB community abundances significantly changed. Additionally, the highest SRB inhibitory effect and the lowest dsrB/soxB ratio were obtained under high concentrations of nitrate and oxygen in the three groups, suggesting that the synergistic effect of nitrate and oxygen level was strong on the inhibition of SRB by potential SOB.

Effects of farmland residual mulch film-derived microplastics on the structure and function of soil and earthworm Metaphire guillelmi gut microbiota.

Microplastics derived from polyethylene (PE) mulch films are widely found in farmland soils and present considerable potential threats to agricultural soil ecosystems. However, the influence of microplastics derived from PE mulch films, especially those derived from farmland residual PE mulch films, on soil ecosystems remains unclear. In this study, we analyzed the bacterial communities attached to farmland residual transparent PE mulch film (FRMF) collected from peanut fields and the different ecological effects of unused PE mulch film-derived microplastics (MPs) and FRMF-derived microplastics (MPs-aged) on the soil and earthworm Metaphire guillelmi gut microbiota, functional traits, and co-occurrence patterns. The results showed that the assembly and functional patterns of the bacterial communities attached to the FRMF were clearly distinct from those in the surrounding farmland soil, and the FRMF enriched some potential plastic-degrading and pathogenic bacteria, such as Nocardioidaceae, Clostridiaceae, Micrococcaceae, and Mycobacteriaceae. MPs substantially influenced the assembly and functional traits of soil bacterial communities; however, they only significantly changed the functional traits of earthworm gut bacterial communities. MPs-aged considerably affected the assembly and functional traits of both soil and earthworm gut bacterial communities. Notably, MPs had a more remarkable effect on nitrogen-related functions than the MPs-aged in numbers for both soil and earthworm gut samples. Co-occurrence network analysis revealed that both MPs and MPs-aged enhanced the synergistic interactions among operational taxonomic units (OTUs) of the composition networks for all samples. For community functional networks, MPs and MPs-aged enhanced the antagonistic interactions for soil samples; however, they exhibited contrasting effects for earthworm gut samples, as MPs enhanced the synergistic interactions among the functional contents. These findings broaden and deepen our understanding of the effects of FRMF-derived microplastics on soil ecosystems, suggesting that the harmful effects of aged plastics on the ecological environment should be considered.

Coal-mining subsidence changed distribution of the microbiomes and their functional genes in a farmland.

Land subsidence is a serious geological event, and can trigger severe environmental and ecological issues. In this study, the influences of coal-mining subsidence on distribution of farmland microbiomes and their functional genes were investigated by 16 S ribosomal RNA (rRNA) gene and metagenome sequencing. The results showed the existence of a core microbiome, which determined the community compositions across the subsidence farmland. Subsidence decreased the relative abundances of dominant Streptomyces, Nocardioides, and Rhizophagus, but increased the relative abundances of dominant Bradyrhizobium, Rhizobium, and Trichoderma. Subsidence also decreased the relative abundances of genes related to carbon metabolism, Quorum sensing, aminoacyl-transfer RNA (tRNA) biosynthesis, and oxidative phosphorylation, and increased the relative abundances of genes related to two-component system and bacterial chemotaxis. Furthermore, subsidence weakened the biosynthesis of organic carbons by decreasing the relative abundances of genes encoding glycosyl transferases, and strengthened decomposition of degradable organic carbons of the microbiomes and auxiliary activities by increasing the relative abundances of genes encoding glycoside hydrolases and polysaccharide lyases. The concentrations of total phosphorus, Mg2+ , and Ca2+ at the lower areas were significantly higher than those at the upper areas, indicating an associated loss of soil nutrients. Canonical correspondence analysis showed that soil moisture, pH, and the concentrations of NH4 + and Ca2+ were the main factors affecting the distribution of the microbiomes and their functional genes. Collectively, this study shows that coal-mining subsidence alters soil physicochemical properties and distribution of farmland microbiomes and their functional genes.

Bacillus subtilis HG-15, a Halotolerant Rhizoplane Bacterium, Promotes Growth and Salinity Tolerance in Wheat (Triticum aestivum).

Certain plant growth-promoting bacteria (PGPB) reduce salt stress damage in plants. Bacillus subtilis HG-15 is a halotolerant bacterium (able to withstand NaCl concentrations as high as 30%) isolated from the wheat rhizoplane in the Yellow River delta. A qualitative and quantitative investigation of the plant growth-promoting characteristics of this strain confirmed nitrogen fixation, potassium dissolution, ammonia, plant hormone, ACC deaminase, and proline production abilities. B. subtilis HG-15 colonization of wheat roots, stems, and leaves was examined via scanning electron microscopy, rep-PCR, and double antibiotic screening. After inoculation with the B. subtilis HG-15 strain, the pH (1.08-2.69%), electrical conductivity (3.17-11.48%), and Na+ (12.98-15.55%) concentrations of rhizosphere soil significantly decreased (p < 0.05). Under no-salt stress (0.15% NaCl), low-salt stress (0.25% NaCl), and high-salt stress (0.35% NaCl) conditions, this strain also significantly increased (p < 0.05) the dry weight (17.76%, 24.46%, and 9.31%), fresh weight (12.80%, 20.48%, and 7.43%), plant height (7.79%, 5.86%, and 13.13%), and root length (10.28%, 17.87%, and 48.95%). Our results indicated that B. subtilis HG-15 can effectively improve the growth of wheat and elicit induced systemic tolerance in these plants, thus showing its potential as a microbial inoculant that can protect wheat under salt stress conditions.

Bacterial and Archaeal Community Distribution in Oilfield Water Re-injection Facilities and the Influences from Microorganisms in Injected Water.

Water flooding is widely employed for oil production worldwide. However, there has never been a systematic investigation of the microbial communities occurring in oilfield water re-injection facilities. Here, we investigated the distribution of bacterial and archaeal communities in water re-injection facilities of an oilfield, and illustrated the combined influences of environmental variation and the microorganisms in injected water on the microbial communities. Bacterial communities from the surface injection facilities were dominated by aerobic or facultative anaerobic Betaproteobacteria, Alphaproteobacteria, and Flavobacteria, whereas Clostridia, Deltaproteobacteria, Anaerolineae, and Synergistia predominated in downhole of the injection wells, and Gammaproteobacteria, Betaproteobacteria, and Epsilonproteobacteria predominated in the production wells. Methanosaeta, Methanobacterium, and Methanolinea were dominant archaea in the injection facilities, while Methanosaeta, Methanomethylovorans, and Methanoculleus predominated in the production wells. This study also demonstrated that the microorganisms in injected water could be easily transferred from injection station to wellheads and downhole of injection wells, and environmental variation and diffusion-limited microbial transfer resulted from formation filtration were the main factors determining microbial community assembly in oil-bearing strata. The results provide novel information on the bacterial and archaeal communities and the underlying mechanisms occurring in oilfield water re-injection facilities, and benefit the development of effective microbiologically enhanced oil recovery and microbiologically prevented reservoir souring programs.

Ecotoxicological effects of different size ranges of industrial-grade polyethylene and polypropylene microplastics on earthworms Eisenia fetida.

The effects of microplastics (MPs) on terrestrial organisms remain poorly understood, even though soil is an important MPs sink. In this study, the earthworms Eisenia fetida were exposed to 0.25% (w/w) of industrial-grade high-density polyethylene (HDPE, 28-145, 133-415 and 400-1464 µm) and polypropylene (PP, 8-125, 71-383 and 761-1660 µm) MPs in an agricultural soil for 28 d. The results showed that HDPE and PP MPs with different size ranges can be ingested by E. fetida. Exposure to different size ranges of HDPE and PP MPs altered the activities of superoxide dismutase, catalase and glutathione S-transferase and induced an increase in the 8-hydroxy-2'-deoxyguanosine level in E. fetida, suggesting that MPs-induced oxidative stress occurred in E. fetida. A size and type-dependent toxicity of MPs to E. fetida was demonstrated by the integrated biological response index. In addition, to obtain detailed molecular information on the responses of E. fetida to MPs exposure, transcriptomic analysis was conducted for E. fetida from HDPE (28-145 µm) and PP (8-125 µm) treatment groups. Transcriptomic analysis identified 34,937 and 28,494 differentially expressed genes in the HDPE and PP MPs treatments compared with the control, respectively. And, exposure to HDPE and PP MPs significantly disturbed several pathways closely related to neurodegeneration, oxidative stress and inflammatory responses in E. fetida. This study provides important information for the ecological risk assessment of different size ranges and types of industrial-grade MPs.

The effects of high-density polyethylene and polypropylene microplastics on the soil and earthworm Metaphire guillelmi gut microbiota.

As an emerging pollutant in terrestrial ecosystem, studies on the effects of microplastics on the gut microbiota of terrestrial organisms are relatively little even though gut microbiota is closely related to host health, metabolism and immunity as well as soil decomposition processes. In this study, earthworms Metaphire guillelmi were exposed to soil amended with 0.25% (w/w) high-density polyethylene (HDPE, 25 µm) or polypropylene (PP, 13 µm) microplastics for 28 d. The ingestion of HDPE and PP microplastics by M. guillelmi was clearly demonstrated by Nile Red fluorescence staining method. There were significant differences for the microbiota between the M. guillelmi gut and the surrounding soil, which may result from the influence of specific conditions in the gut habitat. HDPE and PP microplastics exposure did not induce gut microbiota dysbiosis in M. guillelmi. However, PP microplastics exposure significantly reduced bacterial diversity and altered bacterial community structure in the soil. Specifically, the relative abundance of Aeromonadaceae and Pseudomonadaceae significantly increased while the relative abundance of Nitrososphaeraceae and two unidentified families affiliated with Proteobacteria significantly decreased. This study broadens our understanding of the ecotoxicity of microplastics on the soil and gut microbiota of terrestrial organisms.

Compositions and Abundances of Sulfate-Reducing and Sulfur-Oxidizing Microorganisms in Water-Flooded Petroleum Reservoirs with Different Temperatures in China.

Sulfate-reducing bacteria (SRB) have been studied extensively in the petroleum industry due to their role in corrosion, but very little is known about sulfur-oxidizing bacteria (SOB), which drive the oxidization of sulfur-compounds produced by the activity of SRB in petroleum reservoirs. Here, we surveyed the community structure, diversity and abundance of SRB and SOB simultaneously based on 16S rRNA, dsrB and soxB gene sequencing, and quantitative PCR analyses, respectively in petroleum reservoirs with different physicochemical properties. Similar to SRB, SOB were found widely inhabiting the analyzed reservoirs with high diversity and different structures. The dominant SRB belonged to the classes Deltaproteobacteria and Clostridia, and included the Desulfotignum, Desulfotomaculum, Desulfovibrio, Desulfobulbus, and Desulfomicrobium genera. The most frequently detected potential SOB were Sulfurimonas, Thiobacillus, Thioclava, Thiohalomonas and Dechloromonas, and belonged to Betaproteobacteria, Alphaproteobacteria, and Epsilonproteobacteria. Among them, Desulfovibrio, Desulfomicrobium, Thioclava, and Sulfurimonas were highly abundant in the low-temperature reservoirs, while Desulfotomaculum, Desulfotignum, Thiobacillus, and Dechloromonas were more often present in high-temperature reservoirs. The relative abundances of SRB and SOB varied and were present at higher proportions in the relatively high-temperature reservoirs. Canonical correspondence analysis also revealed that the SRB and SOB communities in reservoirs displayed high niche specificity and were closely related to reservoir temperature, pH of the formation brine, and sulfate concentration. In conclusion, this study extends our knowledge about the distribution of SRB and SOB communities in petroleum reservoirs.

An Exogenous Surfactant-Producing Bacillus subtilis Facilitates Indigenous Microbial Enhanced Oil Recovery.

This study used an exogenous lipopeptide-producing Bacillus subtilis to strengthen the indigenous microbial enhanced oil recovery (IMEOR) process in a water-flooded reservoir in the laboratory. The microbial processes and driving mechanisms were investigated in terms of the changes in oil properties and the interplay between the exogenous B. subtilis and indigenous microbial populations. The exogenous B. subtilis is a lipopeptide producer, with a short growth cycle and no oil-degrading ability. The B. subtilis facilitates the IMEOR process through improving oil emulsification and accelerating microbial growth with oil as the carbon source. Microbial community studies using quantitative PCR and high-throughput sequencing revealed that the exogenous B. subtilis could live together with reservoir microbial populations, and did not exert an observable inhibitory effect on the indigenous microbial populations during nutrient stimulation. Core-flooding tests showed that the combined exogenous and indigenous microbial flooding increased oil displacement efficiency by 16.71%, compared with 7.59% in the control where only nutrients were added, demonstrating the application potential in enhanced oil recovery in water-flooded reservoirs, in particular, for reservoirs where IMEOR treatment cannot effectively improve oil recovery.

Spatial isolation and environmental factors drive distinct bacterial and archaeal communities in different types of petroleum reservoirs in China.

To investigate the spatial distribution of microbial communities and their drivers in petroleum reservoir environments, we performed pyrosequencing of microbial partial 16S rRNA, derived from 20 geographically separated water-flooding reservoirs, and two reservoirs that had not been flooded, in China. The results indicated that distinct underground microbial communities inhabited the different reservoirs. Compared with the bacteria, archaeal alpha-diversity was not strongly correlated with the environmental variables. The variation of the bacterial and archaeal community compositions was affected synthetically, by the mining patterns, spatial isolation, reservoir temperature, salinity and pH of the formation brine. The environmental factors explained 64.22% and 78.26% of the total variance for the bacterial and archaeal communities, respectively. Despite the diverse community compositions, shared populations (48 bacterial and 18 archaeal genera) were found and were dominant in most of the oilfields. Potential indigenous microorganisms, including Carboxydibrachium, Thermosinus, and Neptunomonas, were only detected in a reservoir that had not been flooded with water. This study indicates that: 1) the environmental variation drives distinct microbial communities in different reservoirs; 2) compared with the archaea, the bacterial communities were highly heterogeneous within and among the reservoirs; and 3) despite the community variation, some microorganisms are dominant in multiple petroleum reservoirs.

Microbial diversity and abundance in the Xinjiang Luliang long-term water-flooding petroleum reservoir.

Microbial populations associated with microbial enhanced oil recovery (MEOR) and their abundance in the Xinjiang Luliang water-flooding petroleum reservoir were investigated using 16S rRNA, nitrate reductases, dissimilatory sulfate reductase, and methyl coenzyme-M reductase-encoded genes to provide ecological information for the potential application of MEOR. 16S rRNA gene miseq sequencing revealed that this reservoir harbored large amounts of taxa, including 155 bacterial and 7 archeal genera. Among them, Arcobacter, Halomonas, Marinobacterium, Marinobacter, Sphingomonas, Rhodococcus, Pseudomonas, Dietzia, Ochrobactrum, Hyphomonas, Acinetobacter, and Shewanella were dominant, and have the potential to grow using hydrocarbons as carbon sources. Metabolic gene clone libraries indicated that the nitrate-reducing bacteria (NRB) mainly belonged to Pseudomonas, Azospirillum, Bradyrhizobium, Thauera, Magnetospirillum, Sinorhizobium, Azoarcus, and Rhodobacter; the sulfate-reducing bacteria (SRB) were Desulfarculus, Desulfomonile, Desulfosarcina, Desulfotignum, Desulfacinum, Desulfatibacillum, Desulfatibacillum, Desulfomicrobium, and Desulfovibrio; while the methanogens were archaea and belonged to Methanomethylovorans, Methanosaeta, Methanococcus, Methanolobus, and Methanobacterium. Real-time quantitative PCR analysis indicated that the number of bacterial 16S rRNA reached 106 copies/mL, while the metabolic genes of NRB, SRB, and methanogens reached 104 copies/mL. These results show that the Luliang reservoir has abundant microbial populations associated with oil recovery, suggesting that the reservoir has potential for MEOR.

Microbial abundance and community composition influence production performance in a low-temperature petroleum reservoir.

Enhanced oil recovery using indigenous microorganisms has been successfully applied in the petroleum industry, but the role of microorganisms remains poorly understood. Here, we investigated the relationship between microbial population dynamics and oil production performance during a water flooding process coupled with nutrient injection in a low-temperature petroleum reservoir. Samples were collected monthly over a two-year period. The microbial composition of samples was determined using 16S rRNA gene pyrosequencing and real-time quantitative polymerase chain reaction analyses. Our results indicated that the microbial community structure in each production well microhabitat was dramatically altered during flooding with eutrophic water. As well as an increase in the density of microorganisms, biosurfactant producers, such as Pseudomonas, Alcaligenes, Rhodococcus, and Rhizobium, were detected in abundance. Furthermore, the density of these microorganisms was closely related to the incremental oil production. Oil emulsification and changes in the fluid-production profile were also observed. In addition, we found that microbial community structure was strongly correlated with environmental factors, such as water content and total nitrogen. These results suggest that injected nutrients increase the abundance of microorganisms, particularly biosurfactant producers. These bacteria and their metabolic products subsequently emulsify oil and alter fluid-production profiles to enhance oil recovery.

Dynamic processes of indigenous microorganisms from a low-temperature petroleum reservoir during nutrient stimulation.

Compared to medium-high temperature petroleum reservoirs (30°C-73°C), little is known about microbial regulation by nutrients in low-temperature reservoirs. In this study, we report the performance (oil emulsification and biogas production) and community structure of indigenous microorganisms from a low-temperature (22.6°C) petroleum reservoir during nutrient stimulation. Culture-dependent approaches indicated that the number of hydrocarbon-oxidizing bacteria (HOB), nitrate-reducing bacteria (NRB) and methane-producing bacteria (MPB) increased by between 10- and 1000-fold, while sulfate-reducing bacteria (SRB) were observed at low levels during stimulation. Phylogenetic analysis of the 16S rRNA gene indicated that Pseudomonas, Ochrobactrum, Acinetobacter, Halomonas and Marinobacter, which have the capability to produce surfactants, were selectively enriched. Methanoculleus, Methanosaeta, Methanocorpusculum and Methanocalculus showed the largest increase in relative abundance among archaea. Micro-emulsion formed with an average oil droplet diameter of 14.3 µm (ranging between 4.1 µm and 84.2 µm) during stimulation. Gas chromatographic analysis of gas production (186 mL gas/200 mL medium) showed the levels of CO2 and CH4 increased 8.97% and 6.21%, respectively. Similar to medium-high temperature reservoirs, HOB, NRB, SRB and MPB were ubiquitous in the low-temperature reservoir, and oil emulsification and gas production were the main phenomena observed during stimulation. Oil emulsification required a longer duration of time to occur in the low-temperature reservoir.

Genetic analysis of benzothiophene biodesulfurization pathway of Gordonia terrae strain C-6.

Sulfur can be removed from benzothiophene (BT) by some bacteria without breaking carbon-carbon bonds. However, a clear mechanism for BT desulfurization and its genetic components have not been reported in literatures so far. In this study, we used comparative transcriptomics to study differential expression of genes in Gordonia terrae C-6 cultured with BT or sodium sulfate as the sole source of sulfur. We found that 135 genes were up-regulated with BT relative to sodium sulfate as the sole sulfur source. Many of these genes encode flavin-dependent monooxygenases, alkane sulfonate monooxygenases and desulfinase, which perform similar functions to those involved in the 4S pathway of dibenzothiophene (DBT) biodesulfurization. Three of the genes were found to be located in the same operon, designated bdsABC. Cell extracts of pET28a-bdsABC transfected E. coli Rosetta (DE3) converted BT to a phenolic compound, identified as o-hydroxystyrene. These results advance our understanding of enzymes involved in the BT biodesulfurization pathway.