Effects of Nutrient Level and Growth Rate on the Conjugation Process That Transfers Mobile Antibiotic Resistance Genes in Continuous Cultures.

Bacteria in the effluent of wastewater treatment plants (WWTPs) can transfer antibiotic resistance genes (ARGs) to the bacteria in receiving water through conjugation; however, there is a lack of quantitative assessment of this phenomenon in continuous cultures. Our objective was to determine the effects of background nutrient levels in river water column and growth rates of bacteria on the conjugation frequency of ARGs from effluent bacteria to river bacteria, as well as on the resulting resistance level (i.e., MICs) of the river bacteria. Chemostats were employed to simulate the discharge points of WWTPs into rivers, where effluent bacteria (donor cells) meet river bacteria (recipient cells). Both donor and recipient cells were Escherichia coli cells, and the donor cells were constructed by filter mating with bacteria in the effluent of a local WWTP. Results showed that higher bacterial growth rate (0.45 h-1 versus 0.15 h-1) led to higher conjugation frequencies (10-4 versus 10-6 transconjugant per recipient). The nutrient level also significantly affected the conjugation frequency, albeit to a lesser extent than the growth rate. The MIC against tetracycline increased from 2 mg/L in the recipient to 64 to 128 mg/L in transconjugants. In comparison, the MIC only increased to as high as 8 mg/L in mutants. Whole-genome sequencing showed that the tet-containing plasmid in both the donor and the transconjugant cells also occur in other fecal bacterial genera. The quantitative information obtained from this study can inform hazard identification related to the proliferation of wastewater-associated ARGs in surface water. IMPORTANCE WWTPs have been regarded as an important hot spot of ARGs. The discharge point of WWTP effluent, where ARGs may be horizontally transferred from bacteria of treated wastewater to bacteria of receiving water, is an important interface between the human-dominated ecosystem and the natural environment. The use of batch cultures in previous studies cannot adequately simulate the nutrient conditions and growth rates in receiving water. In this study, chemostats were employed to simulate the continuous growth of bacteria in receiving water. Furthermore, the experimental setup allowed for separate investigations on the effects of nutrient levels (i.e., simulating background nutrients in river water) and bacterial growth rates on conjugation frequencies and resulting resistance levels. The study generates statistically sound ecological data that can be used to estimate the risk of wastewater-originated ARGs as part of the One Health framework.

Hyperspectral Image Data and Waveband Indexing Methods to Estimate Nutrient Concentration on Lettuce (Lactuca sativa L.) Cultivars.

Lettuce is an important vegetable in the human diet and is commonly consumed for salad. It is a source of vitamin A, which plays a vital role in human health. Improvements in lettuce production will be needed to ensure a stable and economically available supply in the future. The influence of nitrogen (N), phosphorus (P), and potassium (K) compounds on the growth dynamics of four hydroponically grown lettuce (Lactuca sativa L.) cultivars (Black Seeded Simpson, Parris Island, Rex RZ, and Tacitus) in tubs and in a nutrient film technique (NFT) system were studied. Hyperspectral images (HSI) were captured at plant harvest. Models developed from the HSI data were used to estimate nutrient levels of leaf tissues by employing principal component analysis (PCA), partial least squares regression (PLSR), multivariate regression, and variable importance projection (VIP) methods. The optimal wavebands were found in six regions, including 390.57-438.02, 497-550, 551-600, 681.34-774, 802-821, and 822-838 nm for tub-grown lettuces and four regions, namely 390.57-438.02, 497-550, 551-600, and 681.34-774 nm for NFT-system-grown lettuces. These fitted models' levels showed high accuracy (R2=0.85-0.99) in estimating the growth dynamics of the studied lettuce cultivars in terms of nutrient content. HSI data of the lettuce leaves and applied N solutions demonstrated a direct positive correlation with an accuracy of 0.82-0.99 for blue and green regions in 400-575 nm wavebands. The results proved that, in most of the tested multivariate regression models, HSI data of freshly cut leaves correlated well with laboratory-measured data.

Coastal Sediment Nutrient Enrichment Alters Seagrass Blue Carbon Sink Capacity.

The seagrass ecosystem is among the most efficient natural carbon sinks that can contribute to climate change mitigation. However, little is known about the effects of coastal nutrient enrichment caused by anthropogenic activities and/or climate change on the capacity of the seagrass blue carbon sink. Our experimental manipulations of sediment nutrient enrichment shifted the blue carbon sink capabilities of seagrass meadows. Sediment nutrient enrichment significantly increased the nutrient content of seagrass litter, stimulating the decomposition of rhizome + root litter by ∼10% while retarding the decomposition of leaf litter by ∼5%. Sediment N + P enrichment increased seagrass growth and litter production, while enrichment of N or P alone did not. Organic carbon (Corg) stocks in the surface sediments (0-5 cm) were 34% higher than those in the control with N + P enrichment due to high litter production and the low decomposition rate of nutrient-enriched leaf litter. However, Corg stocks in the subsurface sediments (5-20 cm) did not increase with sediment nutrient enrichment, which is likely due to accelerated decomposition of rhizome + root litter. Our findings suggest that nutrient loading in coastal sediments alters the blue carbon sink and storage capacities in seagrass meadows by changing the rates of carbon sequestration and decomposition.

Nutrient Level Determines Biofilm Characteristics and Subsequent Impact on Microbial Corrosion and Biocide Effectiveness.

The impact that nutrient level has on biofilm characteristics, biocide effectiveness, and the associated risk of microbiologically influenced corrosion (MIC) was assessed using multispecies biofilms from two different oilfield consortia. A range of microbiological, microscopy, and corrosion methods demonstrated that the continuous flow of nutrients for the microbial growth resulted in higher activity, thickness, and robustness of the biofilms formed on carbon steel, which induced greater localized corrosion compared to biofilms formed under batch, nutrient-depleted conditions. Despite of the differences in biofilm characteristics, biofilms displayed comparable susceptibilities to glutaraldehyde biocide, with similar log10 reductions and percent reductions of microorganisms under both nutrient conditions. Nevertheless, nutrient replenishment impacted the effectiveness of the biocide in controlling microbial populations; a higher concentration of cells survived the biocide treatment in biofilms formed under a continuous flow of nutrients. Complementary DNA-/RNA-based amplicon sequencing and bioinformatics analysis were used to discriminate the active within the total populations in biofilms established at the different nutrient conditions and allowed the identification of the microbial species that remained active despite nutrient depletion and biocide treatment. Detection of persistent active microorganisms after exposure to glutaraldehyde, regardless of biofilm structure, suggested the presence of microorganisms less susceptible to this biocide and highlighted the importance of monitoring active microbial species for the early detection of biocide resistance in oil production facilities.IMPORTANCE Microbiologically influenced corrosion (MIC) is a complex process that generates economic losses to the industry every year. Corrosion must be managed to prevent a loss of containment of produced fluids to the external environment. MIC management includes the identification of assets with higher MIC risk, which could be influenced by nutrient levels in the system. Assessing biofilms under different nutrient conditions is essential for understanding the impact of flow regime on microbial communities and the subsequent impact on microbial corrosion and on the effectiveness of biocide treatment. This investigation simulates closely oil production systems, which contain piping sections exposed to continuous flow and sections that remain stagnant for long periods. Therefore, the results reported here are useful for MIC management and prevention. Moreover, the complementary methodological approach applied in this investigation highlighted the importance of implementing RNA-based methods for better identification of active microorganisms that survive stress conditions in oil systems.

Response of the common reed (Phragmites australis) to nutrient enrichment depends on the growth stage and degree of enrichment: A mesocosm experiment.

Human-induced nutrient enrichment is a major stressor in aquatic ecosystems that has resulted in the alteration of ecosystem structures and functions. However, to date, relatively few studies have explored the temporal dynamics of reed biomass and morphological and biochemical traits under different nutrient levels, as well as the phenological pattern. Based on a mesocosm experiment, we monitored the aboveground and underground biomass of reed at the different plant growth stages, along with plant height, ramet and leaf number, leaf length and width, and carbohydrate and nutrient contents in different organs. We found that the significantly different ratio of aboveground to underground biomass was only observed at the late flowering stage between the slight enrichment (S-E) and heavy enrichment (H-E) groups. The start of the fast-growth phase of the aboveground part and underground part was delayed in the higher nutrient enrichment groups. The length of the fast-growth phase of the aboveground part was the same in the medium enrichment (M-E) and H-E groups and longer than that in the S-E group. For the underground part, the longest fast-growth phase was found in the S-E group (105 days), followed by the H-E and M-E groups (46 and 41 days, respectively). As the nutrient level increased, both increased and decreased values were observed for the 29 monitored morphological and biochemical traits, and the magnitude changed with the different growth stages. Moreover, different degrees of nutrient enrichment could differentially enhance or weaken the relationships among the groups between total biomass and the integrated morphological trait, between structural carbohydrate (SC) and total nitrogen (TN) contents, between total organic carbon (TOC) and TN, between total phosphorus (TP) contents, between TOC and SC contents. Our findings highlight a crucial contribution of ambient nutrient supply to temporal variation in plant biomass and phenological, morphological and biochemical traits.

Denitrifier abundance and community composition linked to denitrification potential in river sediments.

Denitrification in river sediments plays a very important role in removing nitrogen in aquatic ecosystem. To gain insight into the key factors driving denitrification at large spatial scales, a total of 135 sediment samples were collected from Huaihe River and its branches located in the northern of Anhui province. Bacterial community composition and denitrifying functional genes (nirS, nirK, and nosZ) were measured by high-throughput sequencing and real-time PCR approaches. Potential denitrification rate (PDR) was measured by acetylene inhibition method, which varied from 0.01 to 15.69 µg N g-1 h-1. The sequencing results based on 16S rRNA gene found that the main denitrification bacterial taxa included Bacillus, Thiobacillus, Acinetobacter, Halomonas, Denitratisoma, Pseudomonas, Rhodanobacter, and Thauera. Therein, Thiobacillus might play key roles in the denitrification. Total nitrogen and N:P ratio were the only chemical factors related with all denitrification genes. Furthermore, nirS gene abundance could be more susceptible to environmental parameters compared with nirK and nosZ genes. Canonical correspondence analysis indicated that NO3-, NO2-, NH4+ and IP had the significant impacts on the nirS-encoding bacterial community and spatial distributions. There was a significantly positive correlation between Thiobacillus and nirS gene. We considered that higher numbers of nosZ appeared in nutrient rich sediments. More strikingly, PDR was positively correlated with the abundance of three functional genes. Random forest analysis showed that NH4+ was the most powerful predictor of PDR. These findings can yield practical and important reference for the bioremediation or evaluation of wetland systems.

Effects of enzyme-treated soy protein on performance, digestive enzyme activity and mRNA expression of nutrient transporters of laying hens fed different nutrient density diets.

It has been shown that enzyme-treated plant protein can increase performance and promote intestinal health, and save dietary protein. However, our understanding of the effects of enzyme-treated soy protein on performance and intestine function in laying hens, and its rational use, remains limited. The experiment was conducted to study the effect of enzyme-treated soy protein (ETSP) in different nutrient density diets on performance, egg quality, digestive enzyme activity and mRNA expression of amino acid transporters of laying hens. A total of 1 200 Lohmann laying hens (52 wk of age) was randomly divided into a 3 × 2 factorial design that included three nutrient levels: [positive control (PC), metabolisable energy (ME): 2 680 kcal/kg, CP: 15.5%; negative control 1 (NC1), ME: 2 630 kcal/kg, CP: 15%; negative control 2 (NC2), ME:2 580 kcal/kg, CP: 14.5%] and 2 ETSP levels (0 and 0.5%) for 12 weeks. Each treatment had 10 replicates with 20 birds. With the decrease of dietary nutrition density, egg production rate (P = 0.07) and feed conversion ratio (FCR) (P = 0.06) were reduced. Yolk colour was decreased, and yolk index was increased. Supplemented ETSP improved FCR (P = 0.05) and qualified egg rate (P < 0.05). The mass loss rate of egg was decreased after storage for 30 days (P < 0.05). An interaction between nutrient density and ETSP was observed on albumen height and Haugh unit (P < 0.05), and the effects were most noticeable in hens fed 0.5% ETSP in NC2 group. An increase in the activity of trypsin in duodenum (P < 0.05) and the relative expressions of jejunum peptide transporter 1 (PepT1) (P < 0.05) and B0 system neutral amino acid transport carrier (B0AT) mRNA (P < 0.01) was observed during ETSP supplementation. The nutrient density and ETSP supplementation had no significant effect on microbiota in the cecal contents. Overall, the results in this study indicated that the ME decreased 100 kcal/kg and CP decreased 1% in diet of laying hens had a decreasing trend on production performance, no effects on enzyme activity, amino acid transporter mRNA, and gut microbiota, whereas 0.5% ETSP can increase activity of trypsin, PepT1 and B0AT mRNA relative expressions, and improve FCR, qualified egg rate.

Insight into nitrogen and phosphorus coupling effects on mixotrophic Chlorella vulgaris growth under stably controlled nutrient conditions.

In water environment, nitrogen (N) and phosphorus (P) are biochemically dependent nutrients following the co-limitation concept for algae growth under mixotrophic mode. From a practical viewpoint, algae growth may not bring about significant change of the background nutrient concentration of an actual waterbody in contrast to a conventional batch system. In order to better understand the growth pattern of microalgae in aquatic environments, a series of experiments were conducted under stably controlled N-P levels for studying the N-P coupling effect on mixotrophic Chlorella vulgaris growth process, with attention paid to the physiological and biochemical characteristics. It was found that within the concentration range of N = 1-8 mg·L-1 and P = 0.1-1.0 mg·L-1, the variation of the N-P level slightly affected the specific growth rate, but significantly influenced nutrients uptake, biomass dry weight, chlorophyll contents of the grown C. vulgaris. The biochemical and elemental composition of the microalgae tended to be more sensitive to the N-P concentrations and ratios in the lower nutrient range (1-2 mg N·L-1, 0.1-0.4 mg P·L-1) in which the highest N and P conversion rates were gained as 90.18 ± 1.23% and 60.47 ± 1.59%, respectively. The P assimilation and conversion efficiencies were much affected by both N and P supplies, while the P supply showed little influence on N assimilation and conversion efficiencies. It was also noticed that the N level greatly affected the metabolic pathway involving nutrient assimilation, carbohydrate fixation and monosaccharide profile, resulting in conversion of the dominant fraction of protein at N ≤ 2 mg·L-1 into other biochemical compositions including lipids at N ≥ 3 mg·L-1. The fatty acid methyl esters (FAMEs) composition tended to differ with varied nutrient levels. These findings may deepen our understanding of algal growth in aquatic environment and provide perspective for eutrophication control.

Transcriptome profiling of genes involved in nutrient uptake regulated by phosphate-solubilizing bacteria in pepper (Capsicum annuum L.).

Improving nutrient absorption in pepper has become a vital prerequisite for growth to produce a sustainable yield. In this study, transcriptome gene expression in pepper inoculated with two types of phosphate-solubilizing bacteria (PSB) and grown under low and high nutrient levels (LN and HN) was analyzed. Results showed that the root length increased when pepper was grown under LN; however, the root structure was intensively tight under HN. Our data revealed that the roots preferred horizontal growth than longitudinal growth under HN. PSB strains 'M01' and 'N3' significantly (P < 0.01) increased the P uptake by 70.44% and 98.20%, respectively, but decreased the Ca2+ content by 8.96% and 9.13%, respectively, compared with the control (L1). Although no remarkable difference was detected in the chlorophyll content, inoculation with the two PSB strains decreased the Fe3+ content in pepper under HN. The total clean sequenced data from samples ranged between 5,923,659,118 and 9,955,045,953 bp. Transcriptome profiling revealed 320 upregulated and 449 downregulated genes in L3 versus L1 and 468 upregulated and 532 downregulated genes in L4 versus L1. Gene ontology analysis revealed that the biological processes, including response to stress and secondary metabolic process, were involved. Several pathways were subordinate to glycosphingolipid biosynthesis and linoleic acid and nitrogen metabolisms. Analysis of the eukaryotic orthologous group function revealed that most differential genes were attributed to RNA processing and modification, transcription, and signal transduction. Our results provided new insights into the molecular mechanism related to nutrient uptake in pepper inoculated with PSBs.

Effects of exogenous lanthanum(III) exposure on the positive interaction between mutually beneficial populations.

Rare earth elements (REEs) are widely used in various fields, and their accumulation has been reported to pose environmental risks. Most studies confirmed the damage of excessive REE exposure to individual plants; however, little attention has been given to their effects on plant populations. A positive interaction indicates a mutually beneficial relationship between two populations, which is beneficial to the survival and growth of the populations. However, it remains unknown whether exogenous REEs affect the positive interactions between populations. This study investigated the effects of exogenous lanthanum(III) [La(III)] exposure on the positive interaction between soybean (Glycine max L.) and wheat (Triticum aestivum L.) populations by their modules. At normal nutrient level (½-strength Hoagland), the inhibition of excessive La(III) on population modules decreased with increasing population density. Decreases of 39.26 to 1.05% for soybean and 41.45 to 2.41% for wheat indicated the inhibition of La(III) on the positive interaction of both populations weakened with increasing population density. At low nutrient level (»-strength Hoagland), the inhibition of excessive La(III) on population modules increased with increasing population density. Decreases of 5.82-57.14% for soybean and 4.22-59.04% for wheat indicated the inhibition of La(III) on the positive interaction of both population was strengthened with increasing population density. In summary, the inhibitory effects of exogenous La(III) exposure on the positive interaction between populations vary with both nutrient level and population density. This is a new factor that needs to be considered when evaluating the safety risks of REEs in the environment.

Temperature Response of Planktonic Microbiota in Remote Alpine Lakes.

Alpine lakes are considered pristine freshwater ecosystems and sensitive to direct and indirect changes in water temperature as induced by climate change. The bacterial plankton constitutes a key component in the water column and bacterial metabolic activity has direct consequences for water quality. In order to understand bacterial response to global temperature rise in five alpine lakes located in the Austrian Alps (1700-2188 m a.S.L.) water temperature was compared within a decadal period. Depth-integrated samples were characterized in community composition by 16S rDNA deep-amplicon sequencing early [56 ± 16 (SD) days after ice break up] and later (88 ± 16 days) in the growing season. Within the 10 years period, temperature rise was observed through reduced ice cover duration and increased average water temperature. During the early growing season, the average water temperature recorded between circulation in spring until sampling date (WAS), and the day of autumn circulation, as well as chemical composition including dissolved organic carbon influenced bacterial community composition. In contrast, only nutrients (such as nitrate) were found influential later in the growing season. Metabolic theory of ecology (MTE) was applied to explain the dependence of taxonomic richness on WAS in mathematical terms. The calculated activation energy exceeded the frequently reported prediction emphasizing the role of WAS during early growing season. Accordingly, the relative abundance of predicted metabolism related genes increased with WAS. Thus, the dominant influence of temperature after ice break up could be explained by overall climate change effects, such as a more intense warming in spring and an overall higher amplitude of temperature variation.

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Shallow lakes, sensitive to environmental changes due to low environmental carrying capacity, generally experience decreases in ecosystem function and even regime shifts after over-exploitation. Surface sediments of 18 shallow lakes in Yunnan were collected and analyzed to identify the spatial pattern and the influencing factors of cladoceran communities. The results showed that there was significant heterogeneity in the spatial distribution of cladocera in those lakes. For example, the cladoceran community was generally dominated by benthic taxa in alpine lakes, while there was a clear replacement of benthic cladocerans by planktonic ones with increasing nutrient levels across lakes from Southeast Yunnan. Altitude and total phosphorus were the main driving factors, which independently explained 22.0% and 7.7% of the change in cladocera communities, respectively. The altitude gradient and related changes in climate and catchment features were the main factors in structuring cladocerans for shallow lakes of Yunnan, while the anthropogenic impacts on cladoceran distribution was significant via the processes such as catchment development, pollutant input and macrophyte changes. Meanwhile, the interaction between elevation and total phosphorus explained 26.3% of the total variance in cladoceran community shift, indicating that the increase in human activity intensity in lakes at lower altitudes would have stronger impact on cladocerans through anthropogenic nutrient inputs.

Temporal and spatial distribution of Microcystis biomass and genotype in bloom areas of Lake Taihu.

Cyanobacterial blooms as a global environmental issue are of public health concern. In this study, we investigated the spatial (10 sites) and temporal (June, August and October) variations in: 1) their biomass based on chlorophyll-a (chl-a) concentration, 2) their toxic genotype based on gene copy ratio of mcyJ to cpcBA, and 3) their cpcBA genotype composition of Microcystis during cyanobacterial bloom in Lake Taihu. While spatial-temporal variations were found in chl-a and mcyJ/cpcBA ratio, only spatial variation was observed in cpcBA genotype composition. Samples from northwestern part had a higher chl-a, but mcyJ/cpcBA ratio didn't vary among the sites. High chl-a was observed in August, while mcyJ/cpcBA ratio and genotypic richness increased with time. The spatial variations in chl-a and mcyJ/cpcBA ratio and temporal variation in cpcBA genotype were correlated negatively with dissolved N and positively with dissolved P. Spatial distribution of Microcystis biomass was positively correlated with nitrite and P excluding October, but no correlation was found for spatial distribution of mcyJ/cpcBA ratio and cpcBA genotype. Spatial distribution of toxic and cpcBA genotypes may result from horizontal transport of Microcystis colonies, while spatial variation in Microcystis biomass was probably controlled by both nutrient-mediated growth and horizontal transport of Microcystis. The temporal variation in Microcystis biomass, toxic genotype and cpcBA genotype composition were related to nutrient levels, but cause-and-effect relationships require further study.