Anaerobic Nitrate Research Articles

A New Real-Time Simple Method to Measure the Endogenous Nitrate Reductase Activity (Nar) in Paracoccus denitrificans and Other Denitrifying Bacteria.

The transmembrane nitrate reductase (Nar) is the first enzyme in the dissimilatory alternate anaerobic nitrate respiratory chain in denitrifying bacteria. To date, there has been no real-time method to determine its specific activity embedded in its native membrane; here, we describe such a new method, which is useful with the inside-out membranes of Paracoccus denitrificans and other denitrifying bacteria. This new method takes advantage of the native coupling of the endogenous NADH dehydrogenase or Complex I with the reduction of nitrate by Nar through the quinone pool of the inner membranes of P. denitrificans. This is achieved under previously reached anaerobic conditions. Inner controls confirming the specific Nar activity determined by this new method were made by the total inhibition of the Nar enzyme by sodium azide and cyanide, well-known Nar inhibitors. The estimation of the Michaelis-Menten affinity of Nar for NO3- using this so-called Nar-JJ assay gave a Km of 70.4 μM, similar to previously determined values. This new Nar-JJ assay is a suitable, low-cost, and reproducible method to determine in real-time the endogenous Nar activity not only in P. denitrificans, but in other denitrifying bacteria such as Brucella canis, and potentially in other entero-pathogenic bacteria.

Feasibility and mechanism exploration of enhancing short chain fatty acid production assisted by nitrate photolysis during sludge fermentation

Seeking for the eco-friendly and low-cost pretreatment technology has become the prerequisite for the efficient release of the embedded organics in waste activated sludge (WAS), further for the higher production of short chain fatty acids (SCFAs) via anaerobic fermentation. Nitrate, which can produce reactive oxygen species (ROS) and reactive nitrogen species (RNS) during photolysis, may have the positive effect on WAS disintegration and organics degradation, while its performance and mechanism for WAS treatment is still unknown. This study investigated the feasibility of the nitrate photolysis on WAS disintegration and SCFAs production, and elucidated the underlying mechanism. Results showed that the highest acidification rate (26.6 h−1) and short-chain fatty acid (SCFAs) production (5370.6 mg COD/L) were achieved at 8 d in the nitrate photolysis group (Nitrate-UV) during the anaerobic fermentation, significantly promoted by 2.3 and 12.7 folds than that of Control (raw WAS). Acetic acid (HAc) and propionic acid (HPr) accounted for 89.1 % and 90.1 % in Nitrate-UV and Nitrate group (sole nitrate addition), which were much higher than that obtained in UV and Control. HO• and O2•− were the main contributors compared with NO2•, ONOO– and NO• on the release and bioconversion of the soluble organics. Microbial community and metagenomics revealed the promotion of nitrate photolysis on anaerobic fermentation bacteria (AFB) and nitrate reducing bacteria (NRB), as well as the gene abundance related to the metabolic pathways of glycolysis, amino acid, acetate and nitrogen metabolism, which jointly devoted to the SCFAs enhancement during the anaerobic fermentation.

Complete genome sequence of iron-oxidizing Stutzerimonas stutzeri strain FeN3W isolated from Catalina Harbor sediment.

Stutzerimonas stutzeri strain FeN3W is an iron-oxidizing bacterium isolated from marine sediment. FeN3W's 5.9 Mb genome encodes complete pathways for glycolysis, gluconeogenesis, TCA cycle, pentose phosphate pathway, and aerobic and anaerobic (nitrate) respiration. The genome contains 32 putative heme-binding proteins predicted to localize to the cell envelope.

Construction and characterization of a modA gene mutant strain of Klebsiella pneumoniae

To construct a mutant strain of Klebsiella pneumoniae NTUH- K2044 with modA gene deletion and its complementary strain and explore the role of modA gene in modulating anaerobic nitrate respiratory growth and phenotypes of K. pneumoniae. The modA deletion mutant K. pneumoniae strain was constructed by homologous recombination using the suicide vector pKO3-Km. To obtain the complementary strain C-modA, the whole sequence fragment containing the promoter, open reading frame and terminator regions of modA was cloned into pGEM-T-easy and electrically transformed into the modA deletion mutant. The NTUH-K2044 wild-type strain, modA gene deletion mutant and complementary strain were compared by measuring in vitro anaerobic nitrate respiration growth, competitiveness index, biofilm quantification, mucoviscosity assay and morphological measurement using Image J. The modA deletion mutant strain ΔmodA and the complementary strain C-modA were successfully constructed. The modA gene knockout strain showed inhibited anaerobic nitrate respiratory growth compared with the wild- type and C-modA strains with significantly weakened competitiveness, reduced capacity of biofilm synthesis during anaerobiosis, and lowered mucoviscosity under anaerobic conditions. The ΔmodA strain showed a spherical morphology in anaerobic conditions as compared with the normal short rod-like morphology of K. pneumoniae, with also distinctly shorter length than the wild-type and C-modA strains. The molybdate transport system encoding gene modA is associated with the pathogenic capacity of K. pneumoniae by modulating its anaerobic nitrate respiration, competitiveness, biofilm formation, hypermucoviscous phenotype and morphology.

L-arginine supplementation abrogates hypoxia-induced virulence of Staphylococcus aureus in a murine diabetic pressure wound model.

Diabetic foot ulcers (DFUs) are the most common complications of diabetes resulting from hyperglycemia leading to ischemic hypoxic tissue and nerve damage. Staphylococcus aureus is the most frequently isolated bacteria from DFUs and causes severe necrotic infections leading to amputations with a poor 5-year survival rate. However, very little is known about the mechanisms by which S. aureus dominantly colonizes and causes severe disease in DFUs. Herein, we utilized a pressure wound model in diabetic TALLYHO/JngJ mice to reproduce ischemic hypoxic tissue damage seen in DFUs and demonstrated that anaerobic fermentative growth of S. aureus significantly increased the virulence and the severity of disease by activating two-component regulatory systems leading to expression of virulence factors. Our in vitro studies showed that supplementation of nitrate as a terminal electron acceptor promotes anaerobic respiration and suppresses the expression of S. aureus virulence factors through inactivation of two-component regulatory systems, suggesting potential therapeutic benefits by promoting anaerobic nitrate respiration. Our in vivo studies revealed that dietary supplementation of L-arginine (L-Arg) significantly attenuated the severity of disease caused by S. aureus in the pressure wound model by providing nitrate. Collectively, these findings highlight the importance of anaerobic fermentative growth in S. aureus pathogenesis and the potential of dietary L-Arg supplementation as a therapeutic to prevent severe S. aureus infection in DFUs.IMPORTANCES. aureus is the most common cause of infection in DFUs, often resulting in lower-extremity amputation with a distressingly poor 5-year survival rate. Treatment for S. aureus infections has largely remained unchanged for decades and involves tissue debridement with antibiotic therapy. With high levels of conservative treatment failure, recurrence of ulcers, and antibiotic resistance, a new approach is necessary to prevent lower-extremity amputations. Nutritional aspects of DFU treatment have largely been overlooked as there has been contradictory clinical trial evidence, but very few in vitro and in vivo modelings of nutritional treatment studies have been performed. Here we demonstrate that dietary supplementation of L-Arg in a diabetic mouse model significantly reduced duration and severity of disease caused by S. aureus. These findings suggest that L-Arg supplementation could be useful as a potential preventive measure against severe S. aureus infections in DFUs.

Acetylation of K188 and K192 inhibits the DNA-binding ability of NarL to regulate Salmonella virulence.

Salmonella infection significantly increases nitrate levels in the intestine, immune cells, and immune organs of the host, and it can exploit nitrate as an electron acceptor to enhance its growth. In the presence of nitrate or nitrite, NarL, a regulatory protein of the Nar two-component system, is activated and regulates a number of genes involved in nitrate metabolism. However, research on NarL at the post-translational level is limited. In this study, we demonstrate that the DNA-binding sites K188 and 192 of NarL can be acetylated by bacterial metabolite acetyl phosphate and that the degree of acetylation has a considerable influence on the regulatory function of NarL. Specifically, acetylation of NarL negatively regulates the transcription of narG, narK, and napF, which affects the utilization of nitrate in Salmonella. Besides, both cell and mouse models show that acetylated K188 and K192 result in attenuated replication in RAW 264.7 cells, as well as impaired virulence in mouse model. Together, this research identifies a novel NarL acetylation mechanism that regulates Salmonella virulence, providing a new insight and target for salmonellosis treatment.IMPORTANCESalmonella is an important intracellular pathogen that can cause limited gastroenteritis and self-limiting gastroenteritis in immunocompetent humans. Nitrate, the highest oxidation state form of nitrogen, is critical in the formation of systemic infection in Salmonella. It functions as a signaling molecule that influences Salmonella chemotaxis, in addition to acting as a reduced external electron acceptor for Salmonella anaerobic respiration. NarL is an essential regulatory protein involved in nitrate metabolism in Salmonella, and comprehending its regulatory mechanism is necessary. Previous research has linked NarL phosphorylation to the formation of its dimer, which is required for NarL to perform its regulatory functions. Our research demonstrated that acetylation also affects the regulatory function of NarL. We found that acetylation affects Salmonella pathogenicity by weakening the ability of NarL to bind to the target sequence, further refining the mechanism of the anaerobic nitrate respiration pathway.

Relationships between environmental factors and N-cycling microbes reveal the indirect effect of further eutrophication on denitrification and DNRA in shallow lakes

Traditional views indicate that eutrophication and subsequent algal blooms favor denitrification and dissimilatory nitrate reduction to ammonium (DNRA) in lake ecosystems. However, lakes tend to show an increasing propensity for inorganic nitrogen (N) limitation as they become more eutrophic. Thus, the influence of further eutrophication on denitrification and DNRA in eutrophic lakes are unclear due to the uncertainty of N availability. To fill this gap, we investigated the genes abundance (AOA, AOB, nirS, nirK and nrfA) and the composition of N-cycling microbes through quantitative PCR and 16S rRNA sequencing analysis, respectively, in 15 shallow eutrophic lakes of the Yangtze-Huaihe River basin, China. The results indicated that denitrification and DNRA rates could be modulated mainly by their functional gene abundances (nirS, nirK and nrfA), followed by the environmental factors (sediment total organic carbon and nitrogen). Denitrification rates significantly increased from slightly to highly eutrophic lakes, but DNRA rates were not. An explanation is that nitrification provided ample nitrate for denitrification, and this cooperative interaction was indicated by the positive correlation of their gene abundances. In addition, Pseudomonas and Anaeromyxobacter was the dominant genus mediated denitrification and DNRA, showing the potential to perform facultative anaerobic and strict anaerobic nitrate reduction, respectively. High level of dissolved oxygen might favor the facultatively aerobic denitrifiers over the obligately anaerobic fermentative DNRA bacteria in these shallow lakes. Chlorophyll a had a weak but positive effect on the gene abundances for nitrification (AOA and AOB). Further eutrophication had an indirect effect on denitrification and DNRA rates through modulating the genes abundances of N-cycling microbes.

Targeting Anaerobic Respiration in Pseudomonas aeruginosa with Chlorate Improves Healing of Chronic Wounds.

Objective: Pseudomonas aeruginosa is an opportunistic pathogen that can establish chronic infections and form biofilm in wounds. Because the wound environment is largely devoid of oxygen, P. aeruginosa may rely on anaerobic metabolism, such as nitrate respiration, to survive in wounds. While nitrate reductase (Nar) typically reduces nitrate to nitrite, it can also reduce chlorate to chlorite, which is a toxic oxidizing agent. Therefore, chlorate can act as a prodrug to specifically eradicate hypoxic/anoxic, nitrate-respiring P. aeruginosa populations, which are often tolerant to conventional antibiotic treatments. Approach: Using a diabetic mouse model for chronic wounds, we tested the role that anaerobic nitrate respiration plays in supporting chronic P. aeruginosa infections. Results: P. aeruginosa forms biofilm deep within the wound where the environment is anoxic. Daily treatment of P. aeruginosa-infected wounds with chlorate supported wound healing. Chlorate treatment was as effective as a treatment with ciprofloxacin (a conventional antibiotic that targets both oxic and hypoxic/anoxic P. aeruginosa populations). Chlorate-treated wounds showed markers of good-quality wound healing, including well-formed granulation tissue, reepithelialization and microvessel development. Loss- and gain-of-function experiments showed that P. aeruginosa requires nitrate respiration to establish a chronic wound infection and form biofilms. Innovation: We show that the small molecule chlorate, kills the opportunistic pathogen, P. aeruginosa, by targeting a form of anaerobic metabolism called nitrate respiration. Conclusion: Chlorate holds promise as a treatment to combat diverse bacterial infections where oxygen is limiting and/or where pathogens grow as biofilms because many other pathogens possess Nar and survive using anaerobic metabolism.

The response of C/N/S cycling functional microbial communities to redox conditions in shallow aquifers using in-situ sediment as bio-trap matrix

ABSTRACT Microbial communities are fundamental components driving critical biogeochemical carbon (C), nitrogen (N) and sulfur (S) cycles in groundwater ecosystems. The reduction–oxidation (redox) potential is one important environmental factor influencing the microbial community composition. Here, we developed a bio-trap method using in-situ sediment as a matrix to collect aquifer sediment samples and evaluate the response of microbial composition and C/N/S cycling functions to redox variations created by providing sole O2, joint O2 and H2, and sole H2 to three wells. Illumina sequencing analyses showed that the microbial communities in the bio-trap sediment could respond quickly to redox changes in the wells, demonstrating that this bio-trap method is promising for detecting microbial variation in the aquifer sediment. The microbial metabolic functions related to C, N and S cyclings and organic pollutants degradation were predicted by the Kyoto Encyclopedia of Genes and Genomes (KEGG) approach. It was found that the joint O2 and H2 injection produced medium oxidation–reduction potential (ORP −346 and −614 mV) and enhanced more microbial functions than sole O2 or H2, which mainly include oxidative phosphorylation, most carbon source metabolism, various pollutants degradation, and nitrogen and sulfur metabolism. Moreover, the functional genes encoding phenol monooxygenase, dioxygenase, nitrogen fixation, nitrification, aerobic and anaerobic nitrate reductase, nitrite reductase, nitric oxide reductase, and sulfur oxidation increased. These findings tell us the contaminant bioremediation and N, S metabolism can be promoted by adjusting ORP realised by injecting joint O2 and H2.

The role of anthraquinone-2-sulfonate on intra/extracellular electron transfer of anaerobic nitrate reduction

To improve the electron (e−) transfer efficiency, exogenous redox mediators (RMs) were usually employed to enhance the denitrification efficiency due to the electron shuttling. Previous studies were mainly focused on how to improve the extracellular electron transfer (EET) by exogenous RMs. However, the intracellular electron transfer (IET), another crucial e− transfer pathway, of biological denitrification was scarcely reported, especially for the relationship between the denitrification and IET. In this study, Coenzyme Q, Complexes I, II and III were determined as the core components in the IET chain of denitrification by using four specific respiration chain inhibitors (RCIs). Anthraquinone-2-sulfonate (AQS) partially recovered the IET of denitrification from NO3−-N to N2 gas when the RCIs were added. Specifically, the generations of N2 gas were improved by 9.68%–18.25% in the experiments with RCIs and AQS, comparing to that with RCIs. nrfA gene was not detected by reverse transcription-polymerase chain reaction, suggesting that Klebsiella oxytoca strain could not conduct dissimilatory nitrate reduction to ammonium. Nitrate assimilation was considered as the main NH4+-N formation way of K. oxytoca strain. The two e− transfer pathways of denitrification were constructed and the roles of AQS on the IET and EET of denitrification were specifically discussed. The results of this study provided a better understanding of the e− transfer pathways of denitrification, and suggested a potential practical use of exogenous RM on bio-treatment of nitrate-containing wastewater.

Composition of Prokaryotic Communities in Anaerobic Enrichment Cultures of Crude Oil from Romashkino Oilfield (Russia)

Despite the many papers published on oilfield ecosystems, there is a lack of data about microbial community structure and functioning in many crude oils or oilfields and wells. One of such poorly studied oilfields is supergiant Romashkino situated in Tatarstan (Russia). In the present study, prokaryotic community composition and abundance of functional genes encoding sulfate reduction and rhamnolipid production, was investigated in enrichment cultures of four samples from Romashkino oilfield production wells with different oil/water ratios. The enrichment was conducted under anaerobic conditions, sulfate and nitrate were used as electron acceptors, and acetate – as an electron donor. In enrichment culture A and enrichment culture B, bacteria of the Pseudomonadaceae family predominated with 99% and 93% relative abundance, respectively. In enrichment culture C (obtained on the basis of initial sample contained oil with medium water content) and D (obtained on the basis of initial sample contained oil with very high water content), Pseudomonadaceae were one of the two predominating families, along with Micrococcaceae (enrichment culture C) and Dietziaceae (enrichment culture D). The proportion of dsrA and apsA genes encoding sulfate reduction to 16S rRNA gene copy numbers was quite low and ranged between 0.003% (enrichment cultures A, B) and 0.027% (enrichment culture D). The proportion of rhlB gene encoding rhamnolipid production from the total 16S rRNA gene copy number was 1.10% in sample A and 0.06% in sample B. In enrichment cultures C and D the rhlB gene was not found.

Acetylation of NarL K188 and K192 is involved in regulating Escherichia coli anaerobic nitrate respiration.

As a facultative anaerobe, Escherichia coli can activate various respiratory chains during anaerobic growth, among which the mode of anaerobic respiration with nitrate allows good energy conservation. NarL is one of the regulatory proteins in the Nar two-component system that regulates anaerobic respiration in E. coli. Previous studies have shown that NarL activates downstream gene regulation through phosphorylation. However, there are few studies on other protein translational modifications that influence the regulatory function of NarL. Herein, we demonstrate that acetylation modification exists on K188 and K192, the two lysine residues involved in contacting to DNA, and the degree of acetylation has significant effects on DNA-binding abilities, thus affecting the anaerobic growth of E. coli. In addition, NarL is mainly regulated by acetyl phosphate, but not by peptidyl-lysine N-acetyltransferase. These results indicate that non-enzymatic acetylation of NarL by AcP is one of the important mechanisms for the nitrate anaerobic respiratory pathway in response to environmental changes, which extends the idea of the mechanism underlying the response of intestinal flora to changes in the intestinal environment. KEY POINTS: • Acetylation was found in NarL, which was mainly mediated by AcP. • Non-enzymatic acetylation at K188 and K192 affects NarL binding ability. • Acetylation of NarL K188 and K192 regulates anaerobic nitrate growth of E. coli.

Effects of frequency and amount of stover mulching on soil nitrogen and the microbial functional guilds of the endosphere and rhizosphere.

Stover mulching as a conservation and sustainable agricultural practice is beneficial for maintaining soil nitrogen (N) requirements and plant health. The microbial functional guilds of the root and rhizosphere are important factors in the soil nitrogen cycle. However, it is unclear how the frequency and amount of stover mulching influence microbial functional guilds in the root and rhizosphere. Therefore, we investigated the responses of the microbial functional guilds in the endosphere and rhizosphere to maize stover mulching amounts (0, 1/3, 2/3, and total stover mulching every year) and frequencies (once every 3 years and twice every 3 years) under 10-year no-till management. The bacterial functional guilds of nitrogen fixation, nitrification, and anaerobic nitrate oxidation displayed the significantly correlation with C/N, total nitrogen, NO3−, and NH4+. The fungal functional guilds of plant pathogens and saprotrophs showed significantly correlations with C/N, total nitrogen, and NO3−. Moreover, we found that bacterial guilds play a pivotal role in maintaining N requirements at the jointing stage, whereas root endophytic fungal guilds play a more important role than bacterial guilds in regulating plant health at the mature stage. The frequency and amount of stover mulching had significant effects on the microbial functional guilds in the root and rhizosphere. Our data suggest that stover mulch application twice every 3 years is the optimal mulching frequency because it yielded the lowest abundance of nitrifying and anaerobic nitrate-oxidising bacteria and the highest abundance of nitrogen-fixing bacteria at the jointing stage, as well as the lowest abundance of fungal plant pathogens in roots at the mature stage.

SRNA-controlled iron sparing response in Staphylococci.

Staphylococcus aureus, a human opportunist pathogen, adjusts its metabolism to cope with iron deprivation within the host. We investigated the potential role of small non-coding RNAs (sRNAs) in dictating this process. A single sRNA, named here IsrR, emerged from a competition assay with tagged-mutant libraries as being required during iron starvation. IsrR is iron-repressed and predicted to target mRNAs expressing iron-containing enzymes. Among them, we demonstrated that IsrR down-regulates the translation of mRNAs of enzymes that catalyze anaerobic nitrate respiration. The IsrR sequence reveals three single-stranded C-rich regions (CRRs). Mutational and structural analysis indicated a differential contribution of these CRRs according to targets. We also report that IsrR is required for full lethality of S. aureus in a mouse septicemia model, underscoring its role as a major contributor to the iron-sparing response for bacterial survival during infection. IsrR is conserved among staphylococci, but it is not ortholog to the proteobacterial sRNA RyhB, nor to other characterized sRNAs down-regulating mRNAs of iron-containing enzymes. Remarkably, these distinct sRNAs regulate common targets, illustrating that RNA-based regulation provides optimal evolutionary solutions to improve bacterial fitness when iron is scarce.

Subtle sequence differences between two interacting σ54 -dependent regulators lead to different activation mechanisms.

In the strictly anaerobic nitrate reducing bacterium Aromatoleum anaerobium, degradation of 1,3-dihydroxybenzene (1,3-DHB, resorcinol) is controlled by two bacterial enhancer-binding proteins (bEBPs), RedR1 and RedR2, which regulate the transcription of three σ54 -dependent promoters controlling expression of the pathway. RedR1 and RedR2 are identical over their length except for their N-terminal tail which differ in sequence and length (six and eight residues, respectively), a single change in their N-terminal domain (NTD), and nine non-identical residues in their C-terminal domain (CTD). Their NTD is composed of a GAF and a PAS domain connected by a linker helix. We show that each regulator is controlled by a different mechanism: whilst RedR1 responds to the classical NTD-mediated negative regulation that is released by the presence of its effector, RedR2 activity is constitutive and controlled through interaction with BtdS, an integral membrane subunit of hydroxyhydroquinone dehydrogenase carrying out the second step in 1,3-DHB degradation. BtdS sequesters the RedR2 regulator to the membrane through its NTD, where a four-Ile track in the PAS domain, interrupted by a Thr in RedR1, and the N-terminal tail are involved. The presence of 1,3-DHB, which is metabolized to hydroxybenzoquinone, releases RedR2 from the membrane. Most bEBPs assemble into homohexamers to activate transcription; we show that hetero-oligomer formation between RedR1 and RedR2 is favoured over homo-oligomers. However, either an NTD-truncated version of RedR1 or a full-length RedR2 are capable of promoter activation on their own, suggesting they should assemble into homohexamers in vivo. We show that promoter DNA behaves as an allosteric effector through binding the CTD to control ΔNTD-RedR1 multimerization and activity. Overall, the regulation of the 1,3-DHB anaerobic degradation pathway can be described as a novel mode of bEBP activation and assembly.

Bioaugmentation of polycyclic aromatic hydrocarbon (PAH)-contaminated soil with the nitrate-reducing bacterium PheN7 under anaerobic condition

The remediation of polycyclic aromatic hydrocarbon (PAH)-contaminated soil under anaerobic condition is still a huge challenge. In this study, an anaerobic Bacillus firmus strain named PheN7 was firstly isolated from mixture of contaminated soil and sludge samples with phenanthrene as the sole carbon resource under nitrate reducing environment. The anaerobic strain was then inoculated combining with nitrate into the phenanthrene-spiked PAH-contaminated soil to investigate the remediation efficiency by anaerobic bioaugmentation (BA). Results showed that the synergy between PheN7 and indigenous degrading bacteria promoted the remediation efficiency of soil. The average removal efficiencies of phenanthrene in 56 days were 1.73 mg/kg soil·d in BA group, much higher than biostimulation group (sole nitrate addition) and natural degradation which achieved 1.48 mg/kg soil·d and 1.24 mg/kg soil·d of degradation rate, respectively. The outstanding adaptability of PheN7 made it become the dominant species in soil in the terminal period, but the invasion of PheN7 also resulted in the decline of diversity of the indigenous microbial community. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States 2 (PICRUSt 2) results showed that a series of functional genes encoding anaerobic phenanthrene degradation and nitrate reductase enzymes in soil were remarkably strengthened with the addition of PheN7. This study confirmed the contribution of PheN7 as the anaerobic inoculum in PAH-contaminated soil remediation, further evaluating the practical applicability of anaerobic bioaugmentation technology in on-site remediation of real PAH-contaminated sites.

Exogenous spermidine regulates the anaerobic enzyme system through hormone concentrations and related-gene expression in Phyllostachys praecox roots under flooding stress

PurposeAcclimation to hypoxia and anoxia is important in various ecological systems, especially flooded soil. Phyllostachys pracecox is sensitive to flooding, and therefore, it is important to explore ways of alleviating hypoxia stress in the roots. In this study, we investigated the regulatory effect of spermidine (Spd) on flooded P. praecox seedlings. MethodsA batch experiment was carried out in roots treated with Spd under flooding for eight days. The following factors were subsequently measured: growth, survival rate, root respiratory activity, soluble protein and anaerobic respiration enzyme contents (pyruvate decarboxylase, PDC; alcohol dehydrogenase, ADH; lactate dehydrogenase, LDH; alanine aminotransferase, AlaAT), S-adenosylmethionine decarboxylase (SAMDC), nitrate reductase (NR), ACC oxidase (ACO) and ACC synthetase (ACS) activities, free Spd, spermine (Spm) and the diamine precursor putrescine (Put) content, indole-3-acetic acid (IAA) and abscisic acid (ABA) content, ethylene emissions and expression of hormone-related genes. ResultsApplication of Spd promoted root growth (root length, volume, surface and dry weight) and root respiratory inhibition, improved the soluble protein content, and reduced the O2·- production rate, H2O2 and MDA content to alleviate the damage of roots under flooding. A significant increase in SAMDC activity, and ABA and IAA contents were also observed, along with a reduction in ethylene emissions, NR, ACO and ACS activities (p < 0.05). Exogenous Spd increased the free Spd and Spm contents in the P. praecox roots, but decreased the free Put content. Taken together, these findings suggest that hypoxia stress was alleviated. Moreover, exogenous Spd up-regulated the expression of auxin-related genes ARF1, AUX1, AUX2, AUX3 and AUX4, and down-regulated the expression of ethylene-related ACO and ACS genes during flooding. In addition, correlation and RDA analysis showed that ARF1, ACO and ACS significantly promoted the expression of auxin, ACO and ACS enzyme activities, respectively (p < 0.05), while ADH, NR, AlaAT, ethylene emissions, Put, Spd, ACS and ACO were significantly correlated with ACS, ACO, and auxin-related gene expression (p < 0.05). Overall, ethylene emissions, ACS and ACO were identified as the main drivers of ethylene and auxin-related gene structure. ConclusionsThese results suggest that Spd regulated hormone concentrations, the content of Spd, Spm and Put, and related gene expression, in turn regulating physiological changes such as anaerobic enzyme activity, mitigating flooding stress in the roots and improving overall growth. Spd therefore has the potential to improve the adaptability of P. praecox to flooding stress.

Microbial community changes during anaerobic nitrate reduction and Fe(II) oxidation of a coastal saline paddy soil under alkaline pH

Microbial community changes during anaerobic nitrate reduction and Fe(II) oxidation of a coastal saline paddy soil under alkaline pH

The intrinsic relevance of nitrogen removal pathway to varying nitrate loading rate in a polycaprolactone-supported denitrification system.

Up to date, the intrinsic association of nitrate loading rate (NLR) with treatment performance of solid-phase denitrification (SPD) systems is still ambiguous. To address this issue, three continuous up-flow bioreactors were configured. They were packed with polycaprolactone (PCL) under a filling ratio of 30%, 60% or 90% and were operated under a varying NLR of 0.34 ± 0.01-3.99 ± 0.12 gN/(L·d). Results showed that the denitrification efficiency was high (RE > 96%) and stable except the case with the highest NLR, which was mainly attributed to the lack of available carbon sources. At the phylum or genus level, most of the detected dominant bacterial taxa were either associated with organics degradation or nitrogen metabolism. The difference in bacterial community structure among the three stages was mainly caused by NLR rather than the filling ratio. Moreover, as the NLR got higher, the Bray-Curtis distance between samples from the same stage became shorter. By the results of gene or enzyme prediction performed in PICRUSt2, the main nitrogen metabolism pathways in these reactors were denitrification, dissimilatory nitrate reduction to ammonium (DNRA), assimilatory nitrate reduction to ammonium (ANRA) and nitrogen fixation. Moreover, aerobic and anaerobic nitrate dissimilation coexisted in the systems with the latter playing a dominant role. Finally, denitrification and DNRA occurred under both high and low NLR conditions while nitrogen fixation and ANRA preferred to occur under low NLR environments. These findings might help guide practical applications.

Use of Digestate as an Alternative to Mineral Fertilizer: Effects on Soil Mineral Nitrogen and Winter Wheat Nitrogen Accumulation in Clay Loam

Knowledge of the mineralisation and nutrient release of organic fertilisers is essential to ensure plant nutrient demand and availability, to increase N use efficiency and to minimise environmental risks. In 2018–2020, two similar field experiments were carried out on clay loam Cambisol with winter wheat (Triticum aestivum L.) grown without N application and applying liquid anaerobic digestate (LD), pig slurry (PS) and ammonium nitrate (AN) fertilizer with and without additional fertilization (N120 and N120+50). The aim of the research was to compare the effect of organic and mineral fertilizers on the variation of soil mineral nitrogen forms in the 0–30, 30–60 cm soil layers and N accumulation in wheat yield. Fertilizers applied during the previous growing season increased the nitrate and ammonium nitrogen (N-NO3 and N-NH4) content after the resumption of winter wheat vegetation. The dry period in spring (2019) had a negative impact on winter wheat N uptake. In a year of normal moisture content (2020), PS and LD fertilizers and the fertilizer application of the previous year (2019) significantly increased the N-NO3 content in the topsoil, while all applied fertilizers increased it in the deeper soil layer (by a factor of between 3.6 and 12.3), compared to unfertilized soil.