Response of bacterial and micro-eukaryotic communities to spatio-temporal fluctuations of wastewater in full scale constructed wetlands.

How microbial communities respond to wastewater fluctuations is poorly understood. Full-scale surface flow constructed wetlands (SFCWs) were constructed for investigating microbial communities. Results showed that influent wastewater changed sediment bacterial community composition seasonally, indicating that a single bacterial taxonomic group had low resistance (especially, Actinobacteriota and Gammaproteobacteria). However, copy numbers of 16S rRNA, ammonia oxidizing archaea, ammonia oxidizing bacteria, nirS and nirK in the first stage SFCWs were 2.49 × 1010, 3.48 × 109, 5.76 × 106, 8.77 × 108 and 9.06 × 108 g-1 dry sediment, respectively, which remained stable between seasons. Moreover, decreases in the nitrogen concentration in wastewater, changed microbial system state from heterotrophic to autotrophic. Micro-eukaryotic communities were more sensitive to wastewater fluctuations than bacterial communities. Overall, results revealed that microbial communities responded to spatio-temporal fluctuations in wastewater through state changes and species asynchrony. This highlighted complex processes of wastewater treatment by microbial components in SFCWs.

Human-induced homogenization of microbial taxa and function in a subtropical river and its impacts on community stability.

Combination of taxa and function can provide a more comprehensive picture on human-induced microbial homogenization. Here, we obtained 2.58 billion high-throughput sequencing reads and 479 high-quality metagenome-assembled genomes (MAGs) of planktonic microbial communities in a subtropical river for 5 years. We found the microbial taxa homogenization and functional homogenization were uncoupled. Although human activities in downstream sites significantly decreased the taxonomic diversity of non-abundant ASV communities (16S rRNA gene amplicon sequence variants), they did not significantly decrease the taxonomic diversity of abundant ASV and total observed MAG communities. However, the total observed MAG communities in downstream sites tended to homogenize into some specific taxa which encode human-activity-related functional genes, such as nutrient cycles, greenhouse gas emission, antibiotic and arsenic resistance. Those specific MAGs with high taxonomic diversity caused the weak heterogenization of total observed MAG communities in downstream sites. Moreover, functional homogenization promoted the synchrony among downstream MAGs, and these MAGs constructed some specific network modules might to synergistically execute or resist the human-activity-related functions. High synchrony also led to the tandem effects among MAGs and thus decreased community stability. Overall, our findings revealed the links of microbial taxa, functions and stability under human activity impacts, and provided a strong evidence to encourage us re-thinking biotic homogenization based on microbial taxa and their functional attributes.

Biogeographic patterns of micro-eukaryotic generalists and specialists and their effects on regional α-diversity at inter-oceanic scale.

Inter-oceanic scale studies allow us to understand the global spread of micro-organisms in marine ecosystems. In this study, micro-eukaryotic communities in marine surface sediment were collected from tropical to Arctic sites. We found that micro-eukaryotic generalists had much higher intraspecific variation than specialists which allow them to distribute more widely through higher spatiotemporal asynchrony and complementary niche preferences among conspecific taxa. Moreover, comparing to the host-associated protozoa and small metazoa, the algae and free-living protozoa with higher intraspecific variation allow them to have wider distribution ranges. Species abundance also played an important role in driving the distribution ranges of generalists and specialists. The generalists had important effects on regional α-diversity even at an inter-oceanic scale which led to the micro-eukaryotic species richness in polar sites to be mainly influenced by the regional generalists but not the local specialists. In particular, more than 97% of algal species in polar sites were shared with the tropical and subtropical sites (including toxic dinoflagellate). Overall, our study suggests that the effects of global change and human activities on the vulnerable high latitude habitats may lead to biotic homogenization for the whole microbial community (not only the dispersal of some harmful algae) through the potential long-distance spread of generalists.

Response of microbial communities to nutrient removal in coastal sediment by using ecological concrete.

Ecological concrete (eco-concrete) is a kind of environment-friendly material with porous characteristics. In this study, the eco-concrete was used to remove the total nitrogen (TN), total phosphorus (TP), and total organic carbon (TOC) in marine coastal sediment. The bacterial communities in sediment and on eco-concrete surface were also investigated by using high-throughput sequencing and quantitative PCR of 16S rRNA gene. We found that the mean removal efficiencies of TN, TP, and TOC in treatment group were 8.3%, 8.4%, and 12.3% after 28 days. The bacterial community composition in the treatment group was significantly different from that in the control group on day 28. In addition, the bacterial community composition on eco-concrete surface was slightly different from that in sediment, and the copy numbers of 16S rRNA gene were higher on eco-concrete surface than in sediment. The types of eco-concrete aggregates (gravel, pebble, and zeolite) also had effects on the bacterial community composition and 16S rRNA gene copy numbers. Furthermore, we found the abundant genus Sulfurovum increased significantly on eco-concrete surface in the treatment group after 28 days. Bacteria belonging to this genus were found having denitrification ability and were commonly detected in bioreactors for nitrate removal. Overall, our study expands the application scopes of eco-concrete and suggests that the bacterial communities in eco-concrete can potentially enhance the removal efficiency of nutrients in coastal sediment.

Microbial one­carbon and nitrogen metabolisms are beneficial to the reservoir recovery after cyanobacterial bloom.

Cyanobacterial blooms have profound effects on the structure and function of plankton communities in inland waters, but few studies have focused on the effects of microbial-based processes in one­carbon and nitrogen cycling on water quality improvement following the bloom. Here, we compared the structure and function of the bacterial community, focusing on microbial one­carbon and nitrogen metabolisms during and after a cyanobacterial Microcystis bloom in a deep subtropical reservoir. Our data showed that microbial one­carbon and nitrogen cycles were closely related to different periods of the bloom, and the changes of functional genes in microbial carbon and nitrogen cycling showed the same consistent trend as that of Methylomonas sp. With the receding of the bloom, the abundance of Methylomonas as well as the functional genes of microbial one­carbon and nitrogen cycling reached the peak and then recovered. Our results indicate that microbial one­carbon and nitrogen metabolisms were beneficial to the recovery of water quality from the cyanobacterial bloom. This study lays a foundation for a deep understanding of the cyanobacterial decomposition mediated by microbes in one­carbon and nitrogen cycles in inland freshwaters.

A Novel Algicidal Bacterium and Its Effects against the Toxic Dinoflagellate Karenia mikimotoi (Dinophyceae).

The toxic dinoflagellate Karenia mikimotoi is a harmful algal bloom-forming species in coastal areas around the world. It produces ichthyotoxins and hemolytic toxins, with deleterious effects on marine ecosystems. In this study, the bacterium Pseudoalteromonas sp. FDHY-MZ2, with high algicidal efficiency against K. mikimotoi, was isolated from a bloom event. Based on the results, it completely lysed K. mikimotoi cells within 24 h 0.5% (vol/vol), with the algicidal activity of the supernatant of the bacterium culture. Algal cell wall fragmentation occurred, leading to cell death. There was a marked decline in various photochemical traits. When treated with the supernatant, cellulase, pheophorbide a oxygenase (PAO) and cyclin B genes were significantly increased, suggesting induced cell wall deterioration, chloroplast degradation and cell cycle regulation of K. mikimotoi cells. In addition, the expression levels of reactive oxygen species (ROS) scavenging gene was significantly inhibited, indicating that the ROS removal system was damaged. The bacterial culture was dried to obtain the spray-dried powder, which showed algicidal activity rates of 92.2 and 100% against a laboratory K. mikimotoi culture and a field microcosm of Karlodinium sp. bloom within 24 h with the addition of 0.04% mass fraction powder. Our results demonstrate that FDHY-MZ2 is a suitable strain for K. mikimotoi and Karlodinium sp. blooms management. In addition, this study provides a new strategy for the anthropogenic control of harmful algal bloom-forming species in situ. IMPORTANCEK. mikimotoi is a noxious algal bloom-forming species that cause damaging of the aquaculture industry and great financial losses. Bacterium with algicidal activity is an ideal agency to inhibit the growth of harmful algae. In this approach application, the bacterium with high algicidal activity is required and the final management material is ideal for easy-to-use. The algicidal characteristics are also needed to understand the effects of the bacterium for managing strategy exploration. In this study, we isolated a novel algicidal bacterium with extremely high lysis efficiency for K. mikimotoi. The algicidal characteristics of the bacterium as well as the chemical and molecular response of K. mikimotoi with the strain challenge were examined. Finally, the algicidal powder was explored for application. The results demonstrate that FDHY-MZ2 is suitable for K. mikimotoi and Karlodinium sp. blooms controlling, and this study provides a new strategy for algicidal bacterium application.

Nutrient Removal in Eutrophic Water Promotes Stability of Planktonic Bacterial and Protist Communities.

Nutrient (nitrogen and phosphorus) removal by using bioremediation technologies in eutrophic water alters bacterial and protist community structure and function, but how it changes the stability of community remains unclear. To fill this gap, in this study, bacterial and protist communities were investigated using 16S and 18S rRNA gene high-throughput sequencing during the nutrient removal by using ecological floating beds of Canna indica L. Our results showed that both bacterial and protist community compositions in the treatment group were similar to those in the control group at the beginning of the experiment (day 1 to day 11), but then bacterial and protist community compositions became more stable with the removal of nutrients in the treatment group than those in the control group (day 12 to day 18). We further explored the mechanisms for this increased stability and found that the contribution of the stochastic process to bacterial and protist community variations was higher in the control group than that in the treatment group. This suggests that the high nutrient concentration in the control group might increase the random colonization or extinction, and therefore resulted in the high temporal variability (i.e., unstable) of bacterial and protist communities. Our findings suggest that bioremediation for eutrophication can promote the stability of aquatic communities, and therefore potentially maintain aquatic ecosystem functions and services to humanity.

Anthropogenic activities change the relationship between microbial community taxonomic composition and functional attributes.

Functional redundancy is considered common in microbial systems, but recent studies have challenged this idea. The mechanism for this contradictory result is not clear. However, in this study, we hypothesize that strong environmental filtering which links to the anthropogenic activities is able to weaken microbial functional redundancy. We used metagenome and 16S rRNA gene high-throughput sequencing to investigate planktonic microbial communities in a subtropical river. We found that the weak anthropogenic activities might result in a low selection pressure in the river upstream area. Therefore, the microbial community functional attributes were stable although the community composition changed with the water temperature and NO3 -N in upstream area (this indicates functional redundancy). However, the strong anthropogenic activities in river downstream area selected pollutant-degraded functions (e.g. nitrogen metabolism, toluene, xylenes and ethylbenzene degradation) and potentially pollutant-degraded (tolerant) microbes, and therefore caused the microbial community composition synchronously changed with the variation of community functional attributes. Our results reveal that strong environmental filtering which associates with the anthropogenic activities not only has effects on microbial community composition and community functional attributes but also on their relationships. These results provide a new insight to refine the functional redundancy idea.

Transformations from specialists to generalists cause bacterial communities are more stable than micro-eukaryotic communities under anthropogenic activity disturbance.

Different microbial components have different responses to environmental disturbances. Here, we found that the planktonic bacterial and micro-eukaryotic communities had different responses to anthropogenic activity disturbance in a subtropical river, because they had different survival strategies (generalist and specialist). We used nutrients (nitrogen and phosphorus) as indicators of anthropogenic activities. We found that river stretch 1 showed low nutrient concentrations from October 2018 to September 2019. However, a nutrient disturbance was observed in river stretch 2. The nutrient concentrations increased largely in December and January but recovered to low values in June. Bacterial communities had higher resilience under this disturbance than micro-eukaryotic communities in river stretch 2. The bacterial community composition were quite different between the two river stretches in December and January but were similar in June and July. However, the differences of micro-eukaryotic community composition between the two river stretches were always high during the study period. The bacterial communities in river stretch 2 contained more generalists and nutrient tolerant specialists. The bacterial nutrient tolerant specialists rapidly decreased in the low nutrient months and were replaced by the generalists. Bacteria which were involved in this shifts accounted for 29.3% of the total abundance. However, the micro-eukaryotic communities in river stretch 2 contained more moderate generalists. These moderate generalists were insensitive to the variation of nutrients and only 19.56% of the micro-eukaryotes had significant responses to the disturbance. The survival strategies caused bacterial communities had higher adaptability than eukaryotes to environmental fluctuation.

Transcriptome responses of the dinoflagellate Karenia mikimotoi driven by nitrogen deficiency.

The availability of ambient N nutrient is often correlated with the occurrences of harmful algal bloom formed by certain dinoflagellates, making it important to understand how these species might be responding to such conditions. Here, transcriptome sequencing of Karenia mikimotoi was conducted to understand the underlying molecular mechanisms by which this dinoflagellate copes with nitrogen (N) deficiency. Transcriptomic analysis revealed 8802 unigenes (3.56%) that were differentially expressed with ≥ 2-fold change. Under N-depleted conditions, genes involved in glycolysis, fatty acid metabolism, and the tricarboxylic acid (TCA) cycle as well as lipid accumulation were significantly upregulated. The elevated expression of enzymes used in protein degradation and turnover suggests possible metabolic reconfiguration towards accelerated N recycling. Moreover, a significant increase in urea transporter was observed, indicating increased assimilation of organic nitrogen resources as an alternative in N-depleted cultures of K. mikimotoi. The down-regulated glutamate synthase genes were also identified under N deficiency, suggesting suppression of primary amino acid synthesis to save N resource. Taken together, results of this study show enhanced multiple N resource acquisition and reuse of multiple N resources constitute a comprehensive strategy to cope with N deficiency in a dinoflagellate.

Use of ecological concrete for nutrient removal in coastal sediment and its effects on sediment microbial communities.

Ecological concrete (eco-concrete) can reduce excess nutrients (nitrogen and phosphorus) in water, but its effectiveness in removing nutrients in marine coastal sediments and the response of sediment microbial communities to its use are largely unknown. In this study, eco-concrete planted with Bruguiera gymnorrhiza was used to remove nutrients in marine coastal sediment. We found that the mean removal efficiencies of sediment total nitrogen and total phosphorus by using planted eco-concrete were 11.50% and 30.31% on day 60, and were higher than those obtained by only using B. gymnorrhiza (7.14% and 7.36%). the Diatoms and bacterial genera Fusibacter and Anoxynatronum (which belong to Firmicutes) increased and became the abundant microbes by day 60 when using planted eco-concrete, indicating their potential roles in nutrient removal. Moreover, the eco-concrete did not endanger the core microbes in sediment suggesting its environment-friendly character. Our results suggest a potential method to control marine coastal eutrophication.

Enhancement of co-production of lutein and protein in Chlorella sorokiniana FZU60 using different bioprocess operation strategies.

Co-production of multiple compounds is an efficient approach to enhance the economic feasibility of microalgae-based metabolites production. In this study, Chlorella sorokiniana FZU60 was cultivated under different bioprocess strategies to enhance the co-production of lutein and protein. Results showed that both lutein and protein content (7.72 and 538.06 mg/g, respectively) were highest at the onset of nitrogen deficiency under batch cultivation. Semi-batch III strategy, with 75% microalgal culture replacement by fresh medium, obtained similar content, productivity, and yield of lutein and protein as batch cultivation, demonstrating that it can be used for stable and continuous production. Fed-batch II strategy, feeding with 1/3 modified BG11 medium, achieved super-high lutein and protein yield (28.81 and 1592.77 mg/L, respectively), thus can be used for high-output production. Besides, two-stage strategy, combining light intensity shift and semi-batch cultivation, gained extremely high lutein and protein productivity (15.31 and 1080.41 mg/L/day, respectively), thereby is a good option for high-efficiency production. Moreover, the fed-batch II and two-stage strategy achieved high-quality lutein and protein, thus are promising for the co-production of lutein and protein in C. sorokiniana FZU60 for commercial application.

Comprehensive Utilization of Marine Microalgae for Enhanced Co-Production of Multiple Compounds.

Marine microalgae are regarded as potential feedstock because of their multiple valuable compounds, including lipids, pigments, carbohydrates, and proteins. Some of these compounds exhibit attractive bioactivities, such as carotenoids, ω-3 polyunsaturated fatty acids, polysaccharides, and peptides. However, the production cost of bioactive compounds is quite high, due to the low contents in marine microalgae. Comprehensive utilization of marine microalgae for multiple compounds production instead of the sole product can be an efficient way to increase the economic feasibility of bioactive compounds production and improve the production efficiency. This paper discusses the metabolic network of marine microalgal compounds, and indicates their interaction in biosynthesis pathways. Furthermore, potential applications of co-production of multiple compounds under various cultivation conditions by shifting metabolic flux are discussed, and cultivation strategies based on environmental and/or nutrient conditions are proposed to improve the co-production. Moreover, biorefinery techniques for the integral use of microalgal biomass are summarized. These techniques include the co-extraction of multiple bioactive compounds from marine microalgae by conventional methods, super/subcritical fluids, and ionic liquids, as well as direct utilization and biochemical or thermochemical conversion of microalgal residues. Overall, this review sheds light on the potential of the comprehensive utilization of marine microalgae for improving bioeconomy in practical industrial application.

Temperature elevation and acidification damage microstructure of abalone via expression change of crystal induction genes.

Ocean warming and acidification caused by global climate change interferes with the shell growth of mollusks. In abalone Haliotis discus hannai, the microstructural changes in the shell under stress are unclear, and the effect of thermal stress on biomineralization is unknown. The lack of gene information has also hampered the study of abalone biomineralization mechanisms. In this study, the microstructure of reconstructed shell in H. discus hannai was observed to determine the effects of thermal and acidification stress on shell growth. Three nacre protein genes, Hdh-AP7, Hdh-AP24, and Hdh-perlustrin, were characterized, and their expression pattern during shell repair was measured under thermal and acidification stress and compared with those of two known biomineralization-related genes, Hdh-AP-1 and Hdh-defensin. The stress resulted in aragonite plates with corroded or irregular microstructures. The gene expression of two nacre proteins (Hdh-AP7 and Hdh-AP24), which directly induce crystal formation, were more sensitive to thermal stress than to acidification, but the expression of the regulatory nacre protein (Hdh-perlustrin) and the two known genes (Hdh-AP-1 and Hdh-defensin), which are also related to immunity, showed an interlinked, complex pattern change. We concluded that high temperature and acidification damages the shell microstructure by disturbing the expression pattern of biomineralization-related genes.

Pilot-scale cultivation of Chlorella sorokiniana FZU60 with a mixotrophy/photoautotrophy two-stage strategy for efficient lutein production.

Chlorella sorokiniana FZU60, a lutein-enriching microalga, was cultivated in 50 L column photobioreactor to evaluate its potential for lutein production. Initial cell concentration, phosphate concentration and aeration rate were optimized, and results showed that optimal conditions of these three parameters were 0.10 g/L, 0.06 g/L and 0.02 vvm (2.5% CO2), respectively. In addition, a novel two-stage strategy was successfully developed, in which algae were firstly cultivated under fed-batch mixotrophic condition to achieve high biomass concentration, and then shifted to photoautotrophic condition for enhancing lutein accumulation. Moreover, dissolved oxygen was found to be an efficient indicator of acetate depletion in fed-batch stage. The obtained lutein content, production and productivity reached 9.51 mg/g, 33.55 mg/L and 4.67 mg/L/d, respectively, which were greater than those reported in other pilot-scale studies. This proposed strategy provided a cost-effective approach for high-efficient microalgae-based lutein production at pilot-scale, indicating great potential for commercial production.

Characteristics of microbial eukaryotic community recovery in eutrophic water by using ecological floating beds.

Ecological floating beds can rapidly remove nutrients (nitrogen and phosphorus) from eutrophic water, but we still know little about whether this process can simultaneously recover microbial eukaryotic communities. To fill this gap, planktonic microbial eukaryotic communities were investigated using 18S rRNA high-throughput gene sequencing during nutrient removal by floating beds of Canna indica L. We found that nutrient concentrations were high in both the control and treatment groups during period 1 (days 0-5) but rapidly decreased in the treatment group during period 2 (days 6-9) and period 3 (days 10-18). However, the microbial eukaryotic species richness and community compositions were similar between the control and treatment groups during periods 1 and 2 but showed small differences during period 3. The microbial eukaryotic co-occurrence networks between the control and treatment groups also showed similar degree centrality and interconnected eukaryotic members. We found that some abundant fungi species significantly responded to nutrient variations, but a large number of abundant ciliates were insensitive to nutrient removal. Our findings suggest that ecological floating beds can rapidly remove nutrients in eutrophic waters but that it is difficult to quickly and simultaneously improve microbial eukaryotic communities. This result reveals the critical influence of nutrient pollution on aquatic ecosystems and therefore on long-term and comprehensive aquatic habitat restoration, as aquatic macrophyte recoveries should be conducted after nutrient controls have been implemented.

Seasonal Variability of Conditionally Rare Taxa in the Water Column Bacterioplankton Community of Subtropical Reservoirs in China.

Conditionally rare bacteria are ubiquitous and perhaps the most diverse of microbial lifeforms, but their temporal dynamics remain largely unknown. High-throughput and deep sequencing of the 16S rRNA gene has allowed us to identify and compare the conditionally rare taxa with other bacterioplankton subcommunities. In this study, we examined the effect of season, water depth, and ecological processes on the fluctuations of bacterial subcommunities (including abundant, conditionally rare, moderate, and rare taxa) from three subtropical reservoirs in China. We discovered that the conditionally rare taxa (CRT) made up 49.7 to 71.8% of the bacterioplankton community richness, and they accounted for 70.6 to 84.4% of the temporal changes in the community composition. Beta-diversity analysis revealed strong seasonal succession patterns among all bacterioplankton subcommunities, suggesting abundant, conditionally rare, moderate, and rare taxa subcommunities have comparable environmental sensitivity. The dominant phyla of CRT were Proteobacteria, Actinobacteria, and Bacteroidetes, whose variations were strongly correlated with environmental variables. Both deterministic and stochastic processes showed strong effect on bacterioplankton community assembly, with deterministic patterns more pronounced for CRT subcommunity. The difference in bacterial community composition was strongly linked with seasonal change rather than water depth. The seasonal patterns of CRT expand our understanding of underlying mechanisms for bacterial community structure and composition. This implies their importance in the function and stability of freshwater ecosystem after environmental disturbance.

Hysteretic response of Microbial Eukaryotic Communities to Gradually Decreased Nutrient Concentrations in Eutrophic Water.

External environments to microbial eukaryotic communities often change gradually with time. However, whether the responses of microbial eukaryotic communities to these gradually changed environments are continuous or hysteretic and the mechanisms underlying these responses are largely unknown. Here, we used a microcosm to investigate the temporal variation of microbial eukaryotic communities with the gradually decreased nutrient concentrations (nitrogen and phosphorus). We found the differences of microbial eukaryotic community composition and species richness between the control and treatment groups were low during the days 0 to 12, although the nutrient concentrations decreased rapidly during this period in treatment group. However, these differences were clear during the days 14 to 18, although the nutrient concentrations decreased slowly during this period in treatment group. The mechanisms for these results are that the strong homogenous selection (perhaps due to the biotic factors) during the days 8 to 10 in treatment group might enhance the stability of microbial eukaryotic communities. However, the continuously decreased nutrient concentrations weakened the homogenous selection and promoted the strength of environmental filtering, and therefore resulted in the distinct change of microbial eukaryotic communities during the days 14 to 18 in treatment group. Fungi, Chlorophyta and Chrysophyta which associated with the nutrient removal played important roles in this hysteretic change of microbial eukaryotic communities. Overall, our findings suggest that disentangling the non-linear response of communities to gradual environmental changes is essential for understanding ecosystem restoration and degradation in future.

Cloning and characterization of a novel Lustrin A gene from Haliotis discus hannai.

Lustrin A is the first nacre protein with specific structure and amino acid residue content that was identified in abalone; since its identification, homologs have been found in several abalone species. In this study, we isolated and cloned the complete cDNA of Lustrin A from Haliotis discus hannai, which was named Hdh-Lustrin A. Hdh-Lustrin A has characteristic cysteine- and proline-rich domains, glycine- and serine-rich domains, and a whey acidic protein (WAP)-like C-terminus. The cysteine- and proline-rich domains showed internal similarity repeats that arrayed in gene coding region, and the phylogenetic tree of these repeats indicated that the similarity of structural repetitive unit components in different abalone species, reflecting their evolutionary distance. A tissue distribution analysis showed that the mRNA level of Hdh-Lustrin A has tissue-specific expression in mantle. Under lipopolysaccharide (LPS) challenge, Hdh-Lustrin A showed a significantly increase, while it showed a more complex pattern with two peaks in the process of shell regeneration. Moreover, acidification and warming raised the expression level of Hdh-Lustrin A in shell regeneration in two different manners; acidification raised the gene expression in quick response, in contrast the long run in warming treatment. Similar pattern also has been detected in immune reaction and the thermal treatments. These results suggest that the Hdh-Lustrin A is a nacre protein, which can be distinguished by its cysteine- and proline-rich domain. It involves in shell regeneration and innate immunity in abalone, and its expression pattern during shell regeneration can be disrupted by physicochemical properties of the environment.

The impacts of different high-throughput profiling approaches on the understanding of bacterial antibiotic resistance genes in a freshwater reservoir.

Bacterial antibiotic resistance genes (ARGs), a kind of emerging environmental pollutants, greatly threat human health through pathogenic bacteria. High-throughput quantitative PCR (HT-qPCR) and metagenomic approaches are two popular tools applied in aquatic environmental ARGs monitoring. However, current poor knowledge of different ARG profiling approaches' impacts on the understanding of the ARGs in aquatic environments greatly limit the further field application of these two approaches. For filling such knowledge gap, this study simultaneously employed these two approaches to examine and compare the ARGs in a freshwater reservoir across space and time. We found metagenomic approach detected more ARG subtypes and much higher bacitracin resistance genes' abundances than HT-qPCR. In general, HT-qPCR and metagenomics analyses both revealed similar ARG dynamic patterns and co-occurrence patterns between ARGs and bacterial taxa as well as the relationships between ARGs and environmental factors. Our results indicated the impacts of different ARG profiling approaches on the understanding of bacterial ARGs might be minor or negligible. HT-qPCR approach has the superiorities of time-saving, absolute quantification, low requirement for bioinformatics skills but also has some drawbacks including higher PCR amplification & primer bias, higher primer dependency and relative lower ARG subtype quantification capability compared to metagenomic approach. We suggest HT-qPCR approach can be employed for routine aquatic environmental monitoring, and metagenomic approach could be applied in comprehensive surveys for getting more ARG subtype information. Our data can be a useful reference for choosing right ARG profiling approaches for bacterial ARGs monitoring and risk assessment in aquatic environments.