Nitrous oxide (N₂O) is a devastating greenhouse gas and acts as an ozone-depleting agent. Earthworms are a potential source of soil N₂O emissions. Application of biochar can mitigate earthworm-induced N₂O emissions. However, the underlying interactive mechanism between earthworms and biochar in soil N₂O emissions is still unclear. A 35-day laboratory experiment was conducted to examine the soil N₂O emission dynamics for four different treatments, earthworm presence with biochar application (EC), earthworm presence without biochar application (E), earthworm absence with biochar application (C) and earthworm absence without biochar application, and the control. Results indicated a negative impact of biochar on earthworm activity, displaying a significantly (p ≤ 0.05) lower survival rate and biomass of earthworms in treatment EC than E. Compared with the control, earthworm presence significantly (p ≤ 0.05) increased cumulative N₂O emissions, while application of biochar in the presence of earthworms significantly (p ≤ 0.05) decreased cumulative N₂O emissions (485 and 690 μg kg⁻¹ for treatments EC and E, respectively). Treatments E and EC significantly (p ≤ 0.05) increased soil microbial biomass carbon (MBC), ammonium (NH₄⁺-N), nitrate (NO₃⁻N), and dissolved organic carbon (DOC) content and soil pH as compared with the control. The gene copy number of 16 S rRNA, AOA, AOB, nirS, and nosZ increased for all treatments when compared with the control; however, a significant (p ≤ 0.05) difference among the studied genes was only observed for the nosZ gene (2.05 and 2.56 × 10⁶ gene copies g⁻¹ soil for treatments E and EC, respectively). Earthworm-induced soil N₂O emissions were significantly (p ≤ 0.05) reduced by biochar addition. The possible underlying mechanisms may include: (1) short-term negative impacts on earthworm activity; (2) a change of functional gene abundance in earthworm casts; and (3) an increase in soil pH due to addition of biochar.

Aquatic ecosystems and drinking water supply systems worldwide are increasingly affected by taste and odor episodes. In this study, molecular approaches including next-generation sequencing (NGS) and quantitative polymerase chain reaction (qPCR) were used to study the diversity and dynamics of cyanobacteria and 2-methylisoborneol (2-MIB)-producing cyanobacteria in Yuqiao Reservoir, a eutrophicated drinking water reservoir in Tianjin city, northern China. NGS revealed that the entire cyanobacterial community consisted of 16 genera, with Planktothrix (28.8%), Pseudanabaena (18.4%), Cylindrospermosis (7.8%), and Microcystis (7.6%) being the dominant genera, while microscopic examination identified only eight cyanobacterial genera. NGS of the 2-MIB synthesis gene revealed that Pseudanabaena and Planktothricoides were the main 2-MIB producers, with Pseudanabaena being dominant. This finding demonstrated that NGS can identify 2-MIB producers quickly and accurately and it can thus play an important role in the practical monitoring of aquatic ecology. The qPCR test showed 2-MIB synthesis gene with 4.27 × 10⁶ copies/L to 2.24 × 10⁹copies/L occurring at the three sampling sites. The mic gene copy number increased before the 2-MIB concentration increased, indicating that forecasting role in dealing with the 2-MIB concentration by gene copy number. Predicting 2-MIB by qPCR in the field must be verified with additional studies. The combination of NGS and qPCR can be an even more comprehensive method to provide early warning information to managers of reservoirs and water utilities facing taste and odor incidents. This is the first amplicon NGS dataset based on 2-MIB gene to study the diversity and dynamics of 2-MIB-producing cyanobacteria.

Antibiotics can effectively protect livestock from pathogen infection, but residual antibiotics in manure bring risks to ecosystems and public health. Here, we demonstrated that black soldier fly larvae (BSFL) could provide an environmentally friendly manure treatment based on their ability to effectively and rapidly degrade tetracycline (TC). Investigation of the biological mechanisms and degradation pathways of TC by BSFL indicated that nearly 97% of TC was degraded within 12 days in a non-sterile BSFL treatment system, which is up to 1.6-fold faster than that achieved by normal composting. Our results showed that rapid TC-degradation was largely carried out by the intestinal microbiota of the larvae, which doubled the TC-degradation rates compared to those achieved in sterile BSFL systems. This conclusion was further supported by highly-efficient TC-biodegradation both in vivo and in vitro by four larval intestinal isolates. Moreover, detailed microbiome analysis indicated that intestinal bacterial and fungal communities were modified along with significantly increased tet gene copy number in the gut, providing the means to tolerate and degrade TC. Through analysis of TC degradation in vitro, four possible biodegradation products, two hydrolysis products and three conceivable inactivation products were identified, which suggested TC degradation reactions including hydrolysis, oxygenation, deamination, demethylation, ring-cleavage, modification, etc. In conclusion, our studies suggested an estimation of the fate of TC antibiotics in manure treatment by BSFL colonized by gut microbes. These results may provide a strategy for accelerating the degradation of antibiotics by adjusting the intestinal microbiota of BSFL.

Propylea japonica is a very important predator in agricultural ecosystems, which could be exposed to Bt protein. In this study, the bacterial community of P. japonica fed with normal food and food containing Cry2Ab protein was characterized for the first time using qPCR and high-throughput sequencing approaches. Results showed no effect of Cry2Ab on P. japonica development and reproduction. The most abundant bacterial phylum was Firmicutes, and the most abundant genus was Staphylococcus. The total bacteria copy number was not significantly different across four larval stages. Bacteria species composition was gathered more closely in feed on sucrose solution (sucrose-fed) than in larvae only fed on pea aphid (aphid-fed), the diversity indices of some operational taxonomic unit (OTU) were significantly different between sucrose-fed and aphid-fed samples. Different instar larval stages of P. japonica fed with sucrose solution containing Cry2Ab Bt protein and found no effect on microbial community composition and total bacteria copy numbers. However, effects on relative abundance of microbes, copy numbers of Corynebacterium 1 and Glutamicibacter arilaitensis were observed significantly lower in Bt-fed first and fourth larval stages. Low and high concentrations of Cry2Ab protein altered the microbial abundance relative to sucrose-fed P. japonica and copy numbers of G. arilaitensis and Staphylococcus xylosus were significantly lower in Bt-fed samples than control sucrose-fed. Our results are the first report showing that feeding on Cry2Ab protein does not alter microbial species composition in P. japonica, but effects gene copy number of some dominant bacteria. Further investigations are needed to assess the effect of copy number change on P. japonica.

Mine tailings contain toxic metals and can lead to serious pollutions of soil environment. Phytoremediation using legumes has been regarded as an eco-friendly way for the rehabilitation of tailings-laden lands but little is known about the changes of microbial structure during the process. In the present study, we monitored the dynamic change of microbiota in the rhizosphere of Pongamia pinnata during a 2-year on-site remediation of vanadium-titanium magnetite tailings. After remediation, overall soil health conditions were significantly improved as increased available N and P contents and enzyme activities were discovered. There was also an increase of microbial carbon and nitrogen contents. The Illumina sequencing technique revealed that the abundance of taxa under Proteobacteria was increased and rhizobia-related OTUs were preferentially enriched. A significant difference was discovered for sample groups before and after remediation. Rhizobium and Nordella were identified as the keystone taxa at genus rank. Functional predictions indicated that nitrogen fixation was enhanced, corresponding well with qPCR results which showed a significant increase of nifH gene copy numbers by the 2nd year. Our findings for the first time elucidated that legume phytoremediation can effectively cause microbial communities to shift in favour of rhizobia in heavy metal contaminated soil.

Genetically engineered (GE) maize has been thoroughly studied regarding its agro-environmental impact; however, its concerns for the soil environment remain. This work was aimed to decode rhizosphere microbe interactions and potential ecological hazards associated with GE maize. Rhizobacterial communities of field grown transgenic insect-resistant 2A5 maize carrying mcry1Ab and mcry2Ab genes were compared with control Z58 using PacBio sequencing platform. Also full-length 16S rDNA gene sequencing was used to verify the partial (V3–V4) sequencing results obtained in 2017. Measures of α-diversity displayed transgenic 2A5 to be significantly lower in species richness at the flowering stage; however, diversity remained undisturbed. β-diversity was least affected by genetic modifications where similar community profiles were shared by transgenic 2A5 and control Z58. In addition, root exudation patterns were found to drive variations in bacterial assemblages based on developmental stages. For example, genus Massilia successfully colonized the rhizosphere at jointing stage, while Mucilaginobacter showed higher relative abundance in flowering stages of both 2A5 and Z58. These members are known to possess attributes related to plant growth. The impact of dual-transgene insertion on nifH gene abundance was also analyzed where no apparent significant difference in nifH gene copy number was observed. Our results confirmed that full-length 16S rDNA sequencing was sufficient to provide higher taxonomic resolution. Also, results of our 2-year field trials confirmed that there is no significant impact of mcry gene integration on belowground biomasses. Therefore, GE insect-resistant 2A5 maize carrying mcry1Ab and mcry2Ab genes can continue to benefit human populations by increasing crop productivity. In future, further research needs to be catalyzed to analyze the impact of Bt-insertion on microbial community structure across the years for ecosystem sustainability.

Submerged macrophytes and microbial communities are important parts of lake ecosystems. In this study, the bacterial community composition in rhizosphere sediments and water from areas cultivated with (PL) and without (CK) shining pondweed (Potamogeton lucens Linn.) was investigated to determine the effects of P. lucens Linn. on the structure of the bacterial communities in Nansi Lake, China. Molecular techniques, including Illumina MiSeq and qPCR targeting of the 16S rRNA gene, were used to analyze the composition and abundance of the bacterial community. We found that bacterial alpha diversity was higher in PL water than in CK water, and the opposite trend was observed in sediment. In addition, 16S rRNA gene copy number in sediment was lower in PL than in CK. We found 30 (e.g., Desulfatiglans) and 29 (e.g., Limnohabitans) significantly different genera in sediment and water, respectively. P. lucens Linn. can change chemical properties in sediment and water and thereby affect the bacterial community. At the genus level, members of bacterial community clustered according to source (water/sediment) and area (PL/CK). Our study demonstrated that submerged macrophytes can affect the bacterial community composition in both sediment and water, suggesting that submerged macrophytes affect the transportation and cycling of nutrients in lake ecosystems.

Oil spills are events that frequently lead to petroleum pollution. This pollution may cause stress to microbial communities, which require long adaption periods. Soil petroleum pollution is currently considered one of the most serious environmental problems. In the present work, processes occurring in the bacterial communities of three soil samples with different physicochemical characteristics, artificially polluted with 12% of crude oil, were investigated in 120-day laboratory experiment. It was found that the total petroleum hydrocarbon content did not decrease during this time; however, the proportion of petroleum fractions was altered. Petroleum pollution led to a short-term decrease in the bacterial 16S rRNA gene copy number. On the basis of amplicon sequencing analysis, it was concluded that bacterial community successions were similar in the three soils investigated. Thus, the phyla Actinobacteria and Proteobacteria and candidate TM7 phylum (Saccaribacteria) were predominant with relative abundances ranging from 35 to 58%, 25 to 30%, and 15 to 35% in different samples, respectively. The predominant operational taxonomic units (OTUs) after pollution belonged to the genera Rhodococcus and Mycobacterium, families Nocardioidaceae and Sinobacteraceae, and candidate class ТМ7-3. Genes from the alkIII group encoding monoxygenases were the most abundant compared with other catabolic genes from the alkI, alkII, GN-PAH, and GP-PAH groups, and their copy number significantly increased after pollution. The copy numbers of expressed genes involved in the horizontal transfer of catabolic genes, FlgC, TraG, and OmpF, also increased after pollution by 11–33, 16–63, and 11–71 times, respectively. The bacterial community structure after a high level of petroleum pollution changed because of proliferation of the cells that initially were able to decompose hydrocarbons, and in the second place, because proliferation of the cells that received these catabolic genes through horizontal transfer.

This study analyzed the application of three microorganism inoculums, including Bacillus subtilis, Bacillus cereus, and commercial effective microorganism (EM) solution in order to determine cadmium (Cd) reduction in rice (Oryza sativa L.) and rice growth promotion. Rice was grown in Cd-contaminated soil (120 mg/kg) and selected microorganisms were inoculated. Cd concentration and rice weight were measured at 45 and 120 days of the experiment. The result showed that B. subtilis inoculation into rice can highly reduce Cd accumulation in every part of rice roots and shoots (45 days), and grains (120 days). This species can effectively absorb Cd compared to other inoculums, which might be the main mechanism to reduce Cd transportation in rice plants. Interestingly, plants that were inoculated with bacterial species individually harbored higher calcium (Ca) and magnesium (Mg) accumulation; B. subtilis-inoculated plants had the highest levels of Ca and Mg compared to other inoculated ones. Moreover, inoculating rice plants with these microorganisms could increase the dry weight of the plant and protect them from Cd stress because all the inoculums presented the ability to solubilize phosphate, produce indole-3-acetic acid (IAA), and control ethylene levels by 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. After 120 days, quantification of each inoculum by quantitative polymerase chain reaction (qPCR) confirmed the root colonization of bacterial species, with B. subtilis showing higher 16S rRNA gene copy numbers than the other species. B. subtilis was classified as a non-human pathogenic strain, reassuring the safe application of this plant growth-promoting bacterium as a crop inoculum.

The problem of producing strong greenhouse gas of nitrous oxide (N₂O) from biological nitrogen removal (BNR) process in wastewater treatment plants (WWTP) has elicited great concern from various sectors. In this study, three laboratory-scale wastewater treatment systems, with influent C/N ratios of 3.4, 5.4, and 7.5, were set up to study the effect of influent C/N ratio on N₂O generation in anaerobic/anoxic/oxic (A²O) process. Results showed, with the increased influent C/N ratio, N₂O generation from both nitrification and denitrification process was decreased, and the N₂O-N conversion ratio of the process was obviously reduced from 2.23 to 0.05%. Nitrification rate in oxic section was reduced, while denitrification rate in anaerobic and anoxic section was elevated and the removal efficiency of COD, NH₄ ⁺-N, TN, and TP was enhanced in different extent. As the C/N ratio increased from 3.4 to 7.5, activities of three key denitrifying enzymes of nitrate reductase, nitrite reductase, and nitrous oxide reductase were increased. Moreover, microorganism analysis indicated that the relative abundance of ammonium-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) were positively correlated with N₂O generation, which was reduced from (8.42 ± 3.65) to (3.61 ± 1.66)% and (10.38 ± 4.12) to (4.67 ± 1.62)%, respectively. NosZ gene copy numbers of the A²O system were increased from (1.19 ± 0.49) × 10⁷ to (2.84 ± 0.54) × 10⁸ copies/g MLSS with the influent C/N ratio elevated from 3.4 to 7.5. Hence, appropriate influent C/N condition of A²O process could optimize the microbial community structure that simultaneously improve treatment efficiency and decrease the N₂O generation.