Microbial diversity and keystone species drive soil nutrient cycling and multifunctionality following mangrove restoration.

Vegetation restoration exerts transformative effects on nutrient cycling, microbial communities, and ecosystem functions. While extensive research has been conducted on the significance of mangroves and their restoration efforts, the effectiveness of mangrove restoration in enhancing soil multifunctionality in degraded coastal wetlands remains unclear. Herein, we carried out a field experiment to explore the impacts of mangrove restoration and its chronosequence on soil microbial communities, keystone species, and soil multifunctionality, using unrestored aquaculture ponds as controls. The results revealed that mangrove restoration enhanced soil multifunctionality, with these positive effects progressively amplifying over the restoration chronosequence. Furthermore, mangrove restoration led to a substantial increase in microbial diversity and a reshaping of microbial community composition, increasing the relative abundance of dominant phyla such as Nitrospirae, Deferribacteres, and Fusobacteria. Soil multifunctionality exhibited positive correlations with microbial diversity, suggesting a link between variations in microbial diversity and soil multifunctionality. Metagenomic screening demonstrated that mangrove restoration resulted in a simultaneous increase in the abundance of nitrogen (N) related genes, such as N fixation (nirD/H/K), nitrification (pmoA-amoA/B/C), and denitrification (nirK, norB/C, narG/H, napA/B), as well as phosphorus (P)-related genes, including organic P mineralization (phnX/W, phoA/D/G, phnJ/N/P), inorganic P solubilization (gcd, ppx-gppA), and transporters (phnC/D/E, pstA/B/C/S)). The relationship between the abundance of keystone species (such as phnC/D/E) and restoration-induced changes in soil multifunctionality indicates that mangrove restoration enhances soil multifunctionality through an increase in the abundance of keystone species associated with N and P cycles. Additionally, it was observed that changes in microbial community and multifunctionality were largely associated with shifts in soil salinity. These findings demonstrate that mangrove restoration positively influences soil multifunctionality and shapes nutrient dynamics, microbial communities, and overall ecosystem resilience. As global efforts continue to focus on ecosystem restoration, understanding the complexity of mangrove-soil interactions is critical for effective nutrient management and mangrove conservation.

Moderate salinity improves the availability of soil P by regulating P-cycling microbial communities in coastal wetlands.

Accelerated sea-level rise is expected to cause the salinization of freshwater wetlands, but the responses to salinity of the availability of soil phosphorus (P) and of microbial genes involved in the cycling of P remain unexplored. We conducted a field experiment to investigate the effects of salinity on P cycling by soil microbial communities and their regulatory roles on P availability in coastal freshwater and brackish wetlands. Salinity was positively correlated with P availability, with higher concentrations of labile P but lower concentrations of moderately labile P in the brackish wetland. The diversity and richness of microbial communities involved in P cycling were higher in the brackish wetland than the freshwater wetland. Salinity substantially altered the composition of the P-cycling microbial community, in which those of the brackish wetland were separated from those of the freshwater wetland. Metagenomic sequence analysis indicated that functional genes involved in the solubilization of inorganic P and the subsequent transport and regulation of P were more abundant in coastal soils. The relative abundances of most of the target genes differed between the wetlands, with higher abundances of P-solubilization (gcd and ppa) and -mineralization (phoD, phy, and ugpQ) genes and lower abundances of P-transport genes (pstB, ugpA, ugpB, ugpE, and pit) in the brackish wetland. A significant positive correlation between the concentration of labile P and the abundances of the target genes suggested that salinity may, at least in part, improve P availability by regulating the P-cycling microbial community. Our results suggest that the P-cycling microbial community abundance and P availability respond positively to moderate increases in salinity by promoting the microbial solubilization and mineralization of soil P. Changes in microbial communities and microbially mediated P cycling may represent microbial strategies to adapt to moderate salinity levels, which in turn control soil function and nutrient balance.

Reclamation of tidal flats to paddy soils reshuffles the soil microbiomes along a 53-year reclamation chronosequence: Evidence from assembly processes, co-occurrence patterns and multifunctionality.

Coastal soil microbiomes play a key role in coastal ecosystem functioning and are intensely threatened by land reclamation. However, the impacts of coastal reclamation on soil microbial communities, particularly on their assembly processes, co-occurrence patterns, and the multiple soil functions they support, remain poorly understood. This impedes our capability to comprehensively evaluate the impacts of coastal reclamation on soil microbiomes and to restore coastal ecosystem functions degraded by reclamation. Here, we investigated the temporal dynamics of bacterial and fungal communities, community assembly processes, co-occurrence patterns, and ecosystem multifunctionality along a 53-year chronosequence of paddy soil following reclamation from tidal flats. Reclamation of tidal flats to paddy soils resulted in decreased ß-diversity, increased homogeneous selection, and decreased network complexity and robustness of both bacterial and fungal communities, but caused contrasting α-diversity response patterns of them. Reclamation of tidal flats to paddy soils also decreased the multifunctionality of coastal ecosystems, which was largely associated with the fungal network complexity and α-diversity. Collectively, this work demonstrates that coastal reclamation strongly reshaped the soil microbiomes at the level of assembly mechanisms, interaction patterns, and functionality level, and highlights that soil fungal community complexity should be considered as a key factor in restoring coastal ecosystem functions deteriorated by land reclamation.

Effects of microplastics on denitrification and associated N2O emission in estuarine and coastal sediments: insights from interactions between sulfate reducers and denitrifiers.

Global estuarine and coastal zones are facing severe microplastics (MPs) pollution. Sulfate reducers (SRB) and denitrifiers (DNB) are two key functional microorganisms in these zones, exhibiting intricate interactions. However, whether and how MPs modulate the interactions between SRB and DNB, with implications for denitrification and associated N2O emissions, remains poorly understood. Here, we simultaneously investigated the spatial response patterns of SRB-DNB interactions and denitrification and associated N2O emissions to different MPs exposure along an estuarine gradient in the Yangtze Estuary. Spatial responses of denitrification to polyvinyl chloride (PVC) and polyadipate/butylene terephthalate (PBAT) MPs exposure were heterogeneous, while those of N2O emissions were not. Gradient-boosted regression tree and multiple regression model analyses showed that sulfide, followed by nitrate (NO3-), controlled the response patterns of denitrification to MPs exposure. Further mechanistic investigation revealed that exposure to MPs resulted in a competitive and toxic (sulfide accumulation) inhibition of SRB on DNB, ultimately inhibiting denitrification at upstream zones with high sulfide but low NO3- levels. Conversely, MPs exposure induced a competitive inhibition of DNB on SRB, generally promoting denitrification at downstream zones with low sulfide but high NO3- levels. These findings advance the current understanding of the impacts of MPs on nitrogen cycle in estuarine and coastal zones, and provide a novel insight for future studies exploring the response of biogeochemical cycles to MPs in various ecosystems.

Benzimidazole fungicide biotransformation by comammox Nitrospira bacteria: Transformation pathways and associated proteomic responses.

Benzimidazole fungicides are frequently detected in aquatic environments and pose a serious health risk. Here, we investigated the metabolic capacity of the recently discovered complete ammonia-oxidizing (comammox) Nitrospira inopinata and kreftii to transform a representative set of benzimidazole fungicides (i.e., benzimidazole, albendazole, carbendazim, fuberidazole, and thiabendazole). Ammonia-oxidizing bacteria and archaea, as well as the canonical nitrite-oxidizing Nitrospira exhibited no or minor biotransformation activity towards all the five benzimidazole fungicides. In contrast, the investigated comammox bacteria actively transformed all the five benzimidazole fungicides, except for thiabendazole. The identified transformation products indicated hydroxylation, S-oxidation, and glycosylation as the major biotransformation pathways of benzimidazole fungicides. We speculated that these reactions were catalyzed by comammox-specific ammonia monooxygenase, cytochrome P450 monooxygenases, and glycosylases, respectively. Interestingly, the exposure to albendazole enhanced the expression of the antibiotic resistance gene acrB of Nitrospira inopinata, suggesting that some benzimidazole fungicides could act as environmental stressors that trigger cellular defense mechanisms. Altogether, this study demonstrated the distinct substrate specificity of comammox bacteria towards benzimidazole fungicides and implies their significant roles in the biotransformation of these fungicides in nitrifying environments.

Invasive Spartina alterniflora accelerates the increase in microbial nitrogen fixation over nitrogen removal in coastal wetlands of China.

Salt marsh plants play a vital role in mediating nitrogen (N) biogeochemical cycle in estuarine and coastal ecosystems. However, the effects of invasive Spartina alterniflora on N fixation and removal, as well as how these two processes balance to determine the N budget, remain unclear. Here, simultaneous quantifications of N fixation and removal via 15N tracing experiment with native Phragmites australis, invasive S. alterniflora, and bare flats as well as corresponding functional gene abundance by qPCR were carried out to explore the response of N dynamics to S. alterniflora invasion. Our results showed that N fixation and removal rates ranged from 0.77 ± 0.08 to 16.12 ± 1.13 nmol/(g·h) and from 1.42 ± 0.14 to 16.35 ± 1.10 nmol/(g·h), respectively, and invasive S. alterniflora generally facilitated the two processes rates. Based on the difference between N removal and fixation rates, net N2 fluxes were estimated in the range of -0.39 ± 0.14 to 8.24 ± 2.23 nmol/(g·h). Estimated net N2 fluxes in S. alterniflora stands were lower than those in bare flats and P. australis stands, indicating that the increase in N removal caused by S. alterniflora invasion may be more than offset by N fixation process. Random forest analysis revealed that functional microorganisms were the most important factor associated with the corresponding N transformation process. Overall, our results highlight the importance of N fixation in evaluating N budget of estuarine and coastal wetlands, providing valuable insights into the ecological effect of S. alterniflora invasion.

Antibiotics sulfamethoxazole alter nitrous oxide production and pathways in estuarine sediments: Evidenced by the N15-O18 isotopes tracing.

Estuarine antibiotic residues are profoundly impacting microbial nitrogen (N) cycling and associated N2O production, but the response of N2O production pathways to antibiotics remains poorly understood. Here, 15N-18O labeling technique combined with molecular methods were used to investigate the impacts of sulfamethoxazole on the contribution of ammonia oxidation (nitrifier nitrification, nitrifier denitrification, and nitrification-coupled denitrification) and heterotrophic denitrification (HD) to N2O production in estuarine sediments. Results showed that environmental concentration of sulfamethoxazole (4 ng/g) promoted the total N2O production by 17.1% through nitrifier denitrification. Environmentally relevant (40-4000 ng/g) and irrelevant (40,000 ng/g) concentration of sulfamethoxazole drove nitrification denitrification to gradually lose the dominant role in total N2O production and ammonia oxidation-derived N2O, replaced by HD and nitrifier nitrification, while total N2O production were inhibited. Furthermore, when HD dominated the total N2O production, the HD-derived N2O increased by 63.6% with sulfamethoxazole concentration reaching 40,000 ng/g. The mechanistic investigation further showed that nitrifying bacteria were more susceptible to sulfamethoxazole than nitrifying archaea and denitrifiers. The increased expression of nirS gene carried by non-dominant denitrifiers improved the ratio of nirS:nosZ and hence increased HD-derived N2O under high sulfamethoxazole stresses. Overall, our results provide a comprehensive view into how antibiotics regulate N2O production and its pathways in estuarine sediments.

Salinity gradients shape the nitrifier community composition in Nanliu River Estuary sediments and the ecophysiology of comammox Nitrospira inopinata.

The recent discovery of complete ammonia oxidizers (comammox), which convert ammonia to nitrate in a single organism, revolutionized the conventional understanding that two types of nitrifying microorganisms have to be involved in the nitrification process for more than 100 years. However, how different types of nitrifiers in response to salinity change remains largely unclear. This study not only investigated nitrifier community (including ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB), comammox and nitrite-oxidizing Nitrospira) in the Nanliu estuary to find the ecological relationship between salinity and functional communities and also studied the physiology of a typical comammox Nitrospira inopinata in response to a salinity gradient. Based on sequences retrieved with four sets of functional gene primes, comammox Nitrospira was in general, mainly composed of clade A, with a clear separation of clade A1 subgroup in all samples and clade A2 subgroup in low salinity ones. As expected, group I.1b and group I.1a AOA dominated the AOA community in low- and high-salinity samples, respectively. Nitrosomonas-AOB were detected in all samples while Nitrosospira-AOB were mainly found in relatively high-salinity samples. Regarding general Nitrospira, lineages II and IV were the major groups in most of the samples, while lineage I Nitrospira was only detected in low-salinity samples. Furthermore, the comammox pure culture of N. inopinata showed an optimal salinity at 0.5‰ and ceased to grow at 12.8‰ for ammonia oxidation, but remained active for nitrite oxidation. These results show new evidence regarding niche specificity of different nitrifying microorganisms modulated mainly by salinity, and also a clear response by comammox N. inopinata to a wide range of simulated salinity levels.

Corrigendum: Distribution and Diversity of Comammox Nitrospira in Coastal Wetlands of China.

[This corrects the article DOI: 10.3389/fmicb.2020.589268.].

N2O and NOy production by the comammox bacterium Nitrospira inopinata in comparison with canonical ammonia oxidizers.

Nitrous oxide (N2O) and NOy (nitrous acid (HONO) + nitric oxide (NO) + nitrogen dioxide (NO2)) are released as byproducts or obligate intermediates during aerobic ammonia oxidation, and further influence global warming and atmospheric chemistry. The ammonia oxidation process is catalyzed by groups of globally distributed ammonia-oxidizing microorganisms, which are playing a major role in atmospheric N2O and NOy emissions. Yet, little is known about HONO and NO2 production by the recently discovered, widely distributed complete ammonia oxidizers (comammox), able to individually perform the oxidation of ammonia to nitrate via nitrite. Here, we examined the N2O and NOy production patterns by comammox bacterium Nitrospira inopinata during aerobic ammonia oxidation, in comparison to its canonical ammonia-converting counterparts, representatives of the ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB). Our findings, i) show low yield NOy production by the comammox bacterium compared to AOB; ii) highlight the role of the NO reductase in the biological formation of N2O based on results from NH2OH inhibition assays and its stimulation during archaeal and bacterial ammonia oxidations; iii) postulate that the lack of hydroxylamine (NH2OH) and NO transformation enzymatic activities may lead to a buildup of NH2OH/NO which can abiotically react to N2O ; iv) collectively confirm restrained N2O and NOy emission by comammox bacteria, an unneglectable consortium of microbes in global atmospheric emission of reactive nitrogen gases.

Biotransformation of lincomycin and fluoroquinolone antibiotics by the ammonia oxidizers AOA, AOB and comammox: A comparison of removal, pathways, and mechanisms.

In this study, we evaluated the biotransformation mechanisms of lincomycin (LIN) and three fluoroquinolone antibiotics (FQs), ciprofloxacin (CFX), norfloxacin (NFX), and ofloxacin (OFX), which regularly enter aquatic environments through human activities, by different ammonia-oxidizing microorganisms (AOM). The organisms included a pure culture of the complete ammonia oxidizer (comammox) Nitrospira inopinata, an ammonia oxidizing archaeon (AOA) Nitrososphaera gargensis, and an ammonia-oxidizing bacterium (AOB) Nitrosomonas nitrosa Nm90. The removal of these antibiotics by the pure microbial cultures and the protein-normalized biotransformation rate constants indicated that LIN was significantly co-metabolically biotransformed by AOA and comammox, but not by AOB. CFX and NFX were significantly co-metabolized by AOA and AOB, but not by comammox. None of the tested cultures transformed OFX effectively. Generally, AOA showed the best biotransformation capability for LIN and FQs, followed by comammox and AOB. The transformation products and their related biotransformation mechanisms were also elucidated. i) The AOA performed hydroxylation, S-oxidation, and demethylation of LIN, as well as nitrosation and cleavage of the piperazine moiety of CFX and NFX; ii) the AOB utilized nitrosation to biotransform CFX and NFX; and iii) the comammox carried out hydroxylation, demethylation, and demethylthioation of LIN. Hydroxylamine, an intermediate of ammonia oxidation, chemically reacted with LIN and the selected FQs, with removals exceeding 90%. Collectively, these findings provide important fundamental insights into the roles of different ammonia oxidizers and their intermediates on LIN and FQ biotransformation in nitrifying environments including wastewater treatment systems.

Distribution and Diversity of Comammox Nitrospira in Coastal Wetlands of China.

Complete ammonia oxidizers (comammox), able to individually oxidize ammonia to nitrate, are considered to play a Complete ammonia oxidizers (comammox), able to individually oxidize ammonia to nitrate, are considered to play a significant role in the global nitrogen cycle. However, the distribution of comammox Nitrospira in estuarine tidal flat wetland and the environmental drivers affecting their abundance and diversity remain unknown. Here, we present a large-scale investigation on the geographical distribution of comammox Nitrospira along the estuarine tidal flat wetlands of China, where comammox Nitrospira were successfully detected in 9 of the 16 sampling sites. The abundance of comammox Nitrospira ranged from 4.15 × 105 to 6.67 × 106 copies/g, 2.21- to 5.44-folds lower than canonical ammonia oxidizers: ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA). Phylogenetic analysis based on the alpha subunit of the ammonia monooxygenase encoding gene (amoA) revealed that comammox Nitrospira Clade A, mainly originating from upstream river inputs, accounts for more than 80% of the detected comammox Nitrospira, whereas comammox Nitrospira clade B were rarely detected. Comammox Nitrospira abundance and dominant comammox Nitrospira OTUs varied within the estuarine samples, showing a geographical pattern. Salinity and pH were the most important environmental drivers affecting the distribution of comammox Nitrospira in estuarine tidal flat wetlands. The abundance of comammox Nitrospira was further negatively correlated with high ammonia and nitrite concentrations. Altogether, this study revealed the existence, abundance and distribution of comammox Nitrospira and the driving environmental factors in estuarine ecosystems, thus providing insights into the ecological niches of this recently discovered nitrifying consortium and their contributions to nitrification in global estuarine environments.

[Parallel Factor Analysis of Fluorescence Excitation Emission Matrix Spectroscopy of CDOM from the Mid-culture Period of Shrimp Ponds in a Subtropical Estuary].

Components of chromophoric dissolved organic matter (CDOM) from eight coastal land-based shrimp ponds in the Minjiang, Mulan, and Jiulong rivers of subtropical southeastern China were determined by fluorescence excitation emission matrix spectroscopy combined with parallel factor analysis (EEMs-PARAFAC). Four separate fluorescence components, including two protein-like components (C1, C4) and two humic-like components (C2, C3), were identified as the dominant components of the CDOM in these shrimp ponds. The fluorescence indices (FI, BIX, and HIX) suggest that the CDOM of shrimp ponds shows low humification and is mainly derived from spontaneous sources. The protein-like and humic-like components have similar sources and exhibit a similar geochemical behavior. The salinity is negatively correlated with all CDOM fractions, whereas the DOC concentrations show only a positive correlation with the humic fractions of the CDOM. This study provides a scientific basis for the photochemical properties of CDOM in shrimp ponds.

[Short-term Effects of Nitrogen Deposition on CO2 and CH4 Fluxes from Wetlands in the Minjiang River Estuary].

In order to evaluate the effects of nitrogen deposition on CO2 and CH4 fluxes from a Cyperus malaccensis marsh in the Min River estuary at Daoqingzhou, Fujian, static chamber-gas chromatography (GC) techniques were used. CO2 and CH4 fluxes at a control (CK)[0 g·(m2·a)-1], N1[24 g·(m2·a)-1], N2[48 g·(m2·a)-1], and N3[96 g·(m2·a)-1] were measured and analyzed and the related environmental factors were recorded. The results show that,① compared to the control treatment, the effects of nitrogen deposition on the CO2 fluxes for N1 and N3 increased by 20.30% and 4.06%, respectively, whereas the CO2 fluxes for N2 reduced by 10.05%, furthermore, the CO2 fluxes under different treatments were not significantly different (P>0.05), except for the N2 and N3 treatments in December (P<0.05). ② Compared to the control treatment, the effects of nitrogen deposition on CH4 fluxes at N1, N2, and N3 were increased by 64.51%, 30.23%, and 80.57%, respectively, whereas the CH4 fluxes under different treatments were not significantly different (P>0.05). ③ There was significant positive correlation between CO2 and CH4 fluxes and soil temperature (P<0.05), however, the CO2 and CH4 fluxes were not significant with regards to soil pH and soil EC (P>0.05).