Effect of Ba2+ on the biomineralization of Ca2+ and Mg2+ ions induced by Bacillus licheniformis.

The effect of barium ions on the biomineralization of calcium and magnesium ions is often overlooked when utilizing microbial-induced carbonate precipitation technology for removing barium, calcium, and magnesium ions from oilfield wastewater. In this study, Bacillus licheniformis was used to bio-precipitate calcium, magnesium, and barium ions. The effects of barium ions on the physiological and biochemical characteristics of bacteria, as well as the components of extracellular polymers and mineral characteristics, were also studied in systems containing coexisting barium, calcium, and magnesium ions. The results show that the increasing concentrations of barium ions decreased pH, carbonic anhydrase activity, and concentrations of bicarbonate and carbonate ions, while it increased the contents of humic acids, proteins, polysaccharides, and DNA in extracellular polymers in the systems containing all three types of ions. With increasing concentrations of barium ions, the content of magnesium within magnesium-rich calcite and the size of minerals precipitated decreased, while the full width at half maximum of magnesium-rich calcite, the content of O-C=O and N-C=O, and the diversity of protein secondary structures in the minerals increased in systems containing all three coexisting ions. Barium ions does inhibit the precipitation of calcium and magnesium ions, but the immobilized bacteria can mitigate the inhibitory effect. The precipitation ratios of calcium, magnesium, and barium ions reached 81-94%, 68-82%, and 90-97%. This research provides insights into the formation of barium-enriched carbonate minerals and offers improvements for treating oilfield wastewater.

Mn2+ recycling in hypersaline wastewater: unnoticed intracellular biomineralization and pre-cultivation of immobilized bacteria.

Microbially induced manganese carbonate precipitation has been utilized for the treatment of wastewater containing manganese. In this study, Virgibacillus dokdonensis was used to remove manganese ions from an environment containing 5% NaCl. The results showed a significant decrease in carbonic anhydrase activity and concentrations of carbonate and bicarbonate ions with increasing manganese ion concentrations. However, the levels of humic acid analogues, polysaccharides, proteins, and DNA in EPS were significantly elevated compared to those in a manganese-free environment. The rhodochrosite exhibited a preferred growth orientation, abundant morphological features, organic elements including nitrogen, phosphorus, and sulfur, diverse protein secondary structures, as well as stable carbon isotopes displaying a stronger negative bias. The presence of manganese ions was found to enhance the levels of chemical bonds O-C=O and N-C=O in rhodochrosite. Additionally, manganese in rhodochrosite exhibited both + 2 and + 3 valence states. Rhodochrosite forms not only on the cell surface but also intracellularly. After being treated with free bacteria for 20 days, the removal efficiency of manganese ions ranged from 88.4 to 93.2%, and reached a remarkable 100% on the 10th day when using bacteria immobilized on activated carbon fiber that had been pre-cultured for three days. The removal efficiency of manganese ions was significantly enhanced under the action of pre-cultured immobilized bacteria compared to non-pre-cultured immobilized bacteria. This study contributes to a comprehensive understanding of the mineralization mechanism of rhodochrosite, thereby providing an economically and environmentally sustainable biological approach for treating wastewater containing manganese.

Effects of bioaugmentation in para-nitrophenol-contaminated soil on the abundance and community structure of ammonia-oxidizing bacteria and archaea.

Pseudomonas sp. strain WBC-3 mineralizes the priority pollutant para-nitrophenol (PNP) and releases nitrite (NO2 (-)), which is probably involved in the nitrification. In this study, the rate of PNP removal in soil bioaugmented with strain WBC-3 was more accelerated with more NO2 (-) accumulation than in uninoculated soils. Strain WBC-3 survived well and remained stable throughout the entire period. Real-time polymerase chain reaction (real-time PCR) indicated a higher abundance of ammonia-oxidizing bacteria (AOB) than ammonia-oxidizing archaea (AOA), suggesting that AOB played a greater role in nitrification in the original sampled soil. Real-time PCR and multivariate analysis based on the denaturing gradient gel electrophoresis showed that PNP contamination did not significantly alter the abundance and community structure of ammonia oxidizers except for inhibiting the AOB abundance. Bioaugmentation of PNP-contaminated soil showed a significant effect on AOB populations and community structure as well as AOA populations. In addition, ammonium (NH4 (+)) variation was found to be the primary factor affecting the AOB community structure, as determined by the correlation between the community structures of ammonia oxidizers and environmental factors. It is here proposed that the balance between archaeal and bacterial ammonia oxidation could be influenced significantly by the variation in NH4 (+) levels as caused by bioaugmentation of contaminated soil by a pollutant containing nitrogen.

Co-removal and recycling of Ba2+ and Ca2+ in hypersaline wastewater based on the microbially induced carbonate precipitation technique: Overlooked Ba2+ in extracellular and intracellular vaterite.

This study investigates the co-precipitation of calcium and barium ions in hypersaline wastewater under the action of Bacillus licheniformis using microbially induced carbonate precipitation (MICP) technology, as well as the bactericidal properties of the biomineralized product vaterite. The changes in carbonic anhydrase activity, pH, carbonate and bicarbonate concentrations in different biomineralization systems were negatively correlated with variations in metal ion concentrations, while the changes in polysaccharides and protein contents in bacterial extracellular polymers were positively correlated with variations in barium concentrations. In the mixed calcium and barium systems, the harvested minerals were vaterite containing barium. The increasing concentrations of calcium promoted the incorporation and adsorption of barium onto vaterite. The presence of barium significantly increased the contents of O-CO, N-CO, and Ba-O in vaterite. Calcium promoted barium precipitation, but barium inhibited calcium precipitation. After being treated by immobilized bacteria, the concentrations of calcium and barium ions decreased from 400 and 274 to 1.72 and 0 mg/L (GB/T15454-2009 and GB8978-1996). Intracellular minerals were also vaterite containing barium. Extracellular vaterite exhibited bactericidal properties. This research presents a promising technique for simultaneously removing and recycling hazardous heavy metals and calcium in hypersaline wastewater.

Insights into autotrophic carbon fixation strategies through metagonomics in the sediments of seagrass beds.

Seagrass beds contributes up to 10% ocean carbon storage. Carbon fixation in seagrass bed greatly affect global carbon cycle. Currently, six carbon fixation pathways are widely studied: Calvin, reductive tricarboxylic acid (rTCA), Wood-Ljungdahl (WL), 3-hydroxypropionate (3HP), 3-hydroxypropionate/4-hydroxybutyrate (3HP/4HB) and dicarboxylate/4-hydroxybutyrate (DC/4-HB). Despite the knowledges about carbon fixation increase, the carbon fixation strategies in seagrass bed sediment remain unexplored. We collected seagrass bed sediment samples from three sites with different characteristics in Weihai, a city in Shandong, China. The carbon fixation strategies were investigated through metagenomics. The results exhibited that five pathways were present, of which Calvin and WL were the most dominant. The community structure of microorganisms containing the key genes of these pathways were further analyzed, and those dominant microorganisms with carbon fixing potential were revealed. Phosphorus significantly negatively corelated with those microorganisms. This study provides an insight into the strategies of carbon fixation in seagrass bed sediments.

Calcium ion removal at different sodium chloride concentrations by free and immobilized halophilic bacteria.

Much attention has been paid to Ca2+ ion removal by biomineralization due to the dangers of Ca2+ on industrial processes and human health. However, Ca2+ removal from hypersaline water by biomineralization is quite difficult due to there being few halophilic bacteria tolerating higher salinities. In this study, free and immobilized Virgibacillus massiliensis C halophilic bacteria exhibiting carbonic anhydrase activity were used to remove Ca2+ ions from water at different NaCl concentrations. With increasing NaCl concentrations (10, 50, 100, 150 and 200 g/L), Ca2+ ion concentrations in the presence of free bacteria and in two groups of immobilized bacteria for a period of 6 days sharply decreased from 1200 mg/L to 219-562 mg/L, 71-214 mg/L and 21-159 mg/L, respectively; Ca2+ precipitation ratios were 55%-81%, 82%-94% and 87%-98%, respectively. The humic acid-like substances, protein, DNA and polysaccharide, released by the bacteria, promoted the Ca2+ ion removal. The immobilized bacteria were able to be recycled and precultured, which would save industry costs and increase Ca2+ ion removal efficiency. Biological processes for Ca2+ ion removal include cell surface, intracellular and extracellular biomineralization. The biogenesis of calcium carbonate was proved by SEM-EDS, FTIR, XPS and stable carbon isotope values. This study provides insights into the effective removal of Ca2+ ions by biomineralization in hypersaline water.

Calcium ion biorecovery from industrial wastewater by Bacillus amyloliquefaciens DMS6.

Calcium ions in industrial wastewater needs to be removed to prevent the production of limescale, which can have negative consequences. Biomineralization has become the focus due to its lower costs than traditional methods of remediation. In this study, calcium ions were bio-precipitated under the action of free and immobilized Bacillus amyloliquefaciens DMS6 bacteria, and the calcium ion removal efficiency was also compared. The results show that it only needed 3 days to decrease the calcium ion concentration to an ideal level of 76-116 mg/L under the action of DMS6 bacteria immobilized by activated carbon fiber, with calcium ion removal ratios reaching 99%-95% by the 7th day. DMS6 bacteria immobilized by activated carbon fiber were superior to free bacteria and bacteria immobilized by sodium alginate in calcium ion removal. Calcium ions are biomineralized into calcite, Mg-rich calcite, aragonite and monohydrocalcite with abundant organic functional groups, 4 types of secondary protein structures, amino acids, phospholipids, negative stable carbon isotope δ13CPDB values (-16.68‰ to-17.25‰) and negatively charged biomineral surface. Calcium ions were diffused into cells and took part in the intracellular biomineralization of monohydrocalcite, also facilitating calcium ion removal. The formation of intracellular monohydrocalcite has rarely been reported. This study demonstrates an economic and environmentally friendly method to remove calcium ions from industrial wastewater.

RuBisCO large subunit gene primers for assessing the CO2-assimilating planktonic community structure in Jiaozhou Bay, China.

The protein coding genes (rbcL/cbbL/cbbM) for RuBisCO large subunit, the most abundant protein on earth that drives biological CO2 fixation, were considered as useful marker genes in characterizing CO2-assimilating plankton. However, their community specificity has hindered comprehensive screening of genetic diversity. In this study, six different rbcL/cbbL/cbbM primers were employed to screen clone libraries to identify CO2-assimilating plankton in Jiaozhou Bay. The following community compositions were observed: the community components in Form I A/B rbcL/cbbL clone library mainly comprised Chlorophyta and Proteobacteria, Form ID2 and ID3 libraries consisted of Bacillariophyta, Form II cbbM library consisted of Proteobacteria and Alveolata, and both Form I green and red libraries included Proteobacteria, respectively. At the genus taxonomic level, no overlaps among these clone libraries were observed, except for ID2 and ID3. Overall, the phytoplankton in Jiaozhou Bay mainly consists of Bacillariophyta, Chlorophyta, Cryptophyta, Haptophyceae and Alveolata. The CO2-assimilating prokaryotes mainly consist of Proteobacteria. Considering the high-sequence specificities of these marker genes, we propose that the joint use of multiple primers may be utilized in unveiling the diversity of CO2-assimilating organisms. In addition, designing novel RuBisCO gene primers that generate longer amplicons and have broader phylogenetic coverage may be necessary in the future.

Genome-wide identification and characterization of genes encoding cyclohexylamine degradation in a novel cyclohexylamine-degrading bacterial strain of Pseudomonas plecoglossicida NyZ12.

The Gram-negative strain of Pseudomonas plecoglossicida NyZ12 isolated from soil has the ability to degrade cyclohexylamine (CHAM). The genes encoding CHAM degradation by gram-negative bacteria, however, have not been reported previously. In this study, ORFs predicted to encode CHAM degradation by NyZ12 were identified by bioinformatics analysis. Differential expression of the proposed ORFs was analyzed via RNA-seq and quantitative reverse transcription-PCR (qRT-PCR), using RNA extracted from NyZ12 cultured with or without CHAM addition. One CHAM-inducible ORF, RK21_02867 predicted to encode a cyclohexanone monooxygenase (ChnB) was disrupted, as were five ORFs, RK21_00425, RK21_02631, RK21_04207, RK21_04637 and RK21_05539, that had weak homology to the only known cyclohexylamine oxidase (CHAO encoded by chaA) found in Brevibacterium oxydans IH-35A. We also found that a tandem array of five ORFs (RK21_02866-02870) shared homology with those in an operon responsible for oxidation of cyclohexanone to adipic acid, although the ORFs in strain NyZ12 were arranged in a different order with previously found in cyclohexane, cyclohexanol or cyclohexanone degradation strains. The ORFs in this cluster were all up-regulated when CHAM was supplied as the sole carbon source. When one of these five genes, RK21_02867 encoding cyclohexanone (CHnone) monooxygenase, was knocked out, NyZ12 could not grow on CHAM, but it accumulated equimolar amounts of CHnone. Our results show that strain NyZ12 metabolized CHAM directly to CHnone which was then further metabolized to adipate. Despite clearly identifying genes encoding the steps for metabolism of CHAM metabolites, not every one of the putative chaAs was differentially expressed in the presence of CHAM and deletion of each one individually did not completely eliminate the capacity of NyZ12 to degrade CHAM, though it did reduce its growth in several instances. Our results suggest that there is genetic redundancy encoding the initial step in the oxidation of CHAM to CHnone in NyZ12 and that its CHAOs differ considerably from the ChaA, originally described in Brevibacterium oxydans IH-35A.

Bioaugmentation with a consortium of bacterial nitrophenol-degraders for remediation of soil contaminated with three nitrophenol isomers.

A consortium consisting of para-nitrophenol utilizer Pseudomonas sp. strain WBC-3, meta-nitrophenol utilizer Cupriavidus necator JMP134 and ortho-nitrophenol utilizer Alcaligenes sp. strain NyZ215 was inoculated into soil contaminated with three nitrophenol isomers for bioaugmentation. Accelerated removal of all nitrophenols was achieved in inoculated soils compared to un-inoculated soils, with complete removal of nitrophenols in inoculated soils occurring between 2 and 16 days. Real-time polymerase chain reaction (PCR) targeting nitrophenol-degradation functional genes indicated that the three strains survived and were stable over the course of the incubation period. The abundance of total indigenous bacteria (measured by 16S rRNA gene real-time PCR) was slightly negatively impacted by the nitrophenol contamination. Denaturing gradient gel electrophoresis profiles of total and group-specific indigenous community suggested a dynamic change in species richness occurred during the bioaugmentation process. Furthermore, Pareto-Lorenz curves and Community organization parameters indicated that the bioaugmentation process had little impact on species evenness within the microbial community.