Duplex Real-Time Fluorescent Quantitative PCR Assays for the Detection of Toxigenic Microcystis Genotypes Based on SNP/InDel Variation in mcy Gene Cluster.

In this study, the toxigenic characteristics of 14 strains of Microcystis were analyzed, and single nucleotide polymorphism (SNP) and insertion/deletion (InDel) loci in microcystin synthetase (mcy) gene clusters were screened. Based on SNP and InDel loci associated with the toxigenic characteristics, primers and TaqMan or Cycling fluorescent probes were designed to develop duplex real-time fluorescent quantitative PCR (FQ-PCR) assays. After evaluating specificity and sensitivity, these assays were applied to detect the toxigenic Microcystis genotypes in a shrimp pond where Microcystis blooms occurred. The results showed a total of 2155 SNP loci and 66 InDel loci were obtained, of which 12 SNP loci and 5 InDel loci were associated with the toxigenic characteristics. Three duplex real-time FQ-PCR assays were developed, each of which could quantify two genotypes of toxigenic Microcystis. These FQ-PCR assays were highly specific, and two Cycling assays were more sensitive than TaqMan assay. In the shrimp pond, six genotypes of toxigenic Microcystis were detected using the developed FQ-PCR assays, indicating that above genotyping assays have the potential for quantitative analysis of the toxigenic Microcystis genotypes in natural water.

Control of cyanobacterial growth with potassium; implications for bloom control in nuclear storage ponds.

Microbial blooms have been reported in the First Generation Magnox Storage Pond at the Sellafield Nuclear Facility. The pond is kept alkaline with NaOH to minimise fuel rod corrosion, however alkali-tolerant microbial blooms dominated by the cyanobacterium Pseudanabaena catenata are able to thrive in this hostile environment. This study assessed the impact of alternative alkali-dosing regimens (KOH versus NaOH treatment) on biomass accumulation, using a P. catenata dominated mixed culture, which is representative of the pond environment. Optical density was reduced by 40-67 % with KOH treatment over the 3-month chemostat experiment. Microbial community analysis and proteomics demonstrated that the KOH-dependent inhibition of cell growth was mostly specific to P. catenata. The addition of KOH to nuclear storage ponds may therefore help control growth of this pioneer photosynthetic organism due to its sensitivity to potassium, while maintaining the high pH needed to inhibit the corrosion of stored nuclear fuel.

Nitrogen Removal Performance and Microbial Community Structure of IMTA Ponds (Apostistius japonicus-Penaeus japonicus-Ulva).

Denitrification and anaerobic ammonium oxidation (anammox) are key processes for nitrogen removal in aquaculture, reducing the accumulated nitrogen nutrients to nitrogen gas or nitrous oxide gas. Complete removal of nitrogen from aquaculture systems is an important measure to solve environmental pollution. In order to evaluate the nitrogen removal potential of marine aquaculture ponds, this study investigated the denitrification and anammox rates, the flux of nitrous oxide (N2O) at the water-air interface, the sediment microbial community structure, and the gene expression associated with the nitrogen removal process in integrated multi-trophic aquaculture (IMTA) ponds (Apostistius japonicus-Penaeus japonicus-Ulva) with different culture periods. The results showed that the denitrification and anammox rates in sediments increased with the increase of cultivation periods and depth, and there was no significant difference in nitrous oxide gas flux at the water-air interface between different cultivation periods (p > 0.05). At the genus and phylum levels, the abundance of microorganisms related to nitrogen removal reactions in sediments changed significantly with the increase of cultivation period and depth, and was most significantly affected by the concentration of particulate organic nitrogen (PON) in sediments. The expression of denitrification gene (narG, nirS, nosZ) in surface sediments was significantly higher than that in deep sediments (p < 0.05), and was negatively correlated with denitrification rate. All samples had a certain anammox capacity, but no known anammox bacteria were found in the microbial diversity detection, and the expression of gene (hzsB) related to the anammox process was extremely low, which may indicate the existence of an unknown anammox bacterium. The data of this study showed that the IMTA culture pond had a certain potential for nitrogen removal, and whether it could make a contribution to reducing the pollution of culture wastewater still needed additional practice and evaluation, and also provided a theoretical basis for the nitrogen removal research of coastal mariculture ponds.

Roseateles subflavus sp. nov. and Roseateles aquae sp. nov., isolated from artificial pond water and Roseateles violae sp. nov., isolated from a Viola mandshurica root.

Two novel strains, designated APW6T and APW11T, were isolated from artificial pond water, and one novel strain, designated PFR6T, was isolated from a Viola mandshurica root. These strains were found to be Gram-negative, rod-shaped, motile by means of flagella, and oxidase-positive. Growth conditions of the type strains were as follows: APW6T, 15-43 °C (optimum, 28 °C), pH 6.0-12.0 (optimum, pH 7.0), with no salinity; APW11T, 4-35 °C (optimum, 25 °C), pH 6.0-11.0 (optimum, pH 9.0), with 0-1 % NaCl (w/v, optimum 0 %); PFR6T, 10-38 °C (optimum 28 °C), pH 6.0-12.0 (optimum, pH 7.0), with 0-2 % NaCl (w/v; optimum, 0 %). Strains APW6T, APW11T, and PFR6T belonged to the genus Roseateles, having the most 16S rRNA gene sequence similarity to Roseateles saccharophilus DSM 654T (98.1 %), Roseateles oligotrophus CHU3T (98.7 %), and Roseateles puraquae CCUG 52769T (98.1 %). The estimated genome sizes of APW6T, APW11T, and PFR6T were 50 50 473, 56 70 008, and 52 16 869 bp, respectively and the G+C contents were 69.5, 66, and 68.5 mol%. The digital DNA-DNA hybridization, average amino acid identity, and average nucleotide identity values among the novel strains and related taxa were all lower than 22.4, 74.7, and 78.9 %, respectively. The predominant cellular fatty acids (>10 %) of all strains were summed feature 3 (comprising C16 : 1 ω6c and/or C16 : 1 ω7c) and C16 : 0. PFR6T also had summed feature 8 (comprising C18 :  1 ω7c and/or C18 :  1 ω6c) as a major fatty acid. The polar lipid profile of all strains contained phosphatidylethanolamine, phosphoaminoglycolipid, and phosphoglycolipid. The distinct phylogenetic, physiological, and chemotaxonomic features reported in this study indicate that strains APW6T, APW11T, and PFR6T represent novel species within the genus Roseateles, for which the names Roseateles subflavus sp. nov., with the type strain APW6T (=KACC 22877T=TBRC 16606T), Roseateles aquae sp. nov., with the type strain APW11T (=KACC 22878T=TBRC 16607T), and Roseateles violae sp. nov (=KACC 23257T=TBRC 17653T) are respectively proposed.

Gemmobacter denitrificans sp. nov., a denitrifying bacterium, isolated from pond water for Litopenaeus vannamei.

A novel Gram-stain-negative strain, designated JM10B15T, was isolated from pond water for Litopenaeus vannamei collected from Jiangmen City, Guangdong province, south PR China. Cells of the strain were aerobic, rod-shaped, and motile by lateral flagella. JM10B15T could grow at 15-40 °C, pH 6.0-9.5, and in 0-3.0 % NaCl, with optimal growth at 25-35 °C, pH 7.5-8.5, and in 0 % NaCl, respectively. Furthermore, this strain grew well on Reasoner's 2A agar but not on nutrient broth agar or Luria-Bertani agar. JM10B15T was a denitrifying bacterium capable of removing nitrites and nitrates, and three key functional genes, nasA, nirS, and nosZ, were identified in its genome. The results of phylogenetic analyses based on the 16S rRNA gene and genome sequences indicated that JM10B15T belonged to the genus Gemmobacter. JM10B15T showed the highest 16S rRNA sequence similarity to Gemmobacter lutimaris YJ-T1-11T (98.8 %), followed by Gemmobacter aquatilis IFAM 1031T (98.6 %) and Gemmobacter serpentinus HB-1T (98.1 %). The average nucleotide identity and digital DNA-DNA hybridization values between JM10B15T and the other type strains of genus Gemmobacter were 78.1-82.1 % and 18.4-22.1 %, respectively. The major fatty acids of strain JM10B15T were summed feature 8 (C18 : 1 ω6c and/or C18 : 1 ω7c) and C18 : 1 ω7c 11-methyl. In addition, the major respiratory quinone of this novel strain was Q-10, and the predominant polar lipids were phosphatidylglycerol, phosphatidylethanolamine, four unidentified phospholipids, three unidentified lipids, and an unidentified aminophospholipid. Results of analyses of the phylogenetic, genomic, physiological, and biochemical characteristics indicated that JM10B15T represents a novel species of the genus Gemmobacter, for which the name Gemmobacter denitrificans sp. nov. is proposed. The type strain is JM10B15T (=GDMCC 1.4148T=KCTC 8140T).

Global biochemical profiling of fast-growing Antarctic bacteria isolated from meltwater ponds by high-throughput FTIR spectroscopy.

Fourier transform infrared (FTIR) spectroscopy is a biophysical technique used for non-destructive biochemical profiling of biological samples. It can provide comprehensive information about the total cellular biochemical profile of microbial cells. In this study, FTIR spectroscopy was used to perform biochemical characterization of twenty-nine bacterial strains isolated from the Antarctic meltwater ponds. The bacteria were grown on two forms of brain heart infusion (BHI) medium: agar at six different temperatures (4, 10, 18, 25, 30, and 37°C) and on broth at 18°C. Multivariate data analysis approaches such as principal component analysis (PCA) and correlation analysis were used to study the difference in biochemical profiles induced by the cultivation conditions. The observed results indicated a strong correlation between FTIR spectra and the phylogenetic relationships among the studied bacteria. The most accurate taxonomy-aligned clustering was achieved with bacteria cultivated on agar. Cultivation on two forms of BHI medium provided biochemically different bacterial biomass. The impact of temperature on the total cellular biochemical profile of the studied bacteria was species-specific, however, similarly for all bacteria, lipid spectral region was the least affected while polysaccharide region was the most affected by different temperatures. The biggest temperature-triggered changes of the cell chemistry were detected for bacteria with a wide temperature tolerance such Pseudomonas lundensis strains and Acinetobacter lwoffii BIM B-1558.

Identification and Composition of Cyanobacteria in Ecuadorian Shrimp Farming Ponds-Possible Risk to Human Health.

Toxic cyanobacterial blooms in various water bodies have been given much attention nowadays as they release hazardous substances in the surrounding areas. These toxic planktonic cyanobacteria in shrimp ponds greatly affect the survival of shrimps. Ecuador is the second highest shrimp producing country in the Americas after Brazil; and the shrimp-based economy is under threat due to toxic cyanobacterial blooms in Ecuador shrimp ponds. This study investigated the abundance of different cyanobacteria in the shrimp ponds at the Chone and Jama rivers (in Manabi province) at Ecuadorian pacific coast, focusing on different environmental factors, such as temperature, pH, salinity, and light. Temperature and pH were identified as key factors in influencing the abundance of cyanobacteria, with a significant positive correlation between Raphidiopsis raciborskii and pH. The highest and lowest abundance of cyanobacteria found during the dry season in the shrimp ponds near the Chone and Jama rivers were > 3 × 106 and 1 × 106 Cell.m-3, respectively. The Shannon-Wiener Diversity Index fluctuated between 0.41-1.15 and 0.31-1.15 for shrimp ponds of Chone and Jama rivers, respectively. This variation was linked to changes in salinity and the presence of harmful algal blooms, highlighting the importance of continuous monitoring. Additionally, the study areas showed eutrophic conditions with low diversity, underlining the need for additional spatiotemporal studies and expanded research in both rivers, to better understand these complex phenomena. The findings underscore the importance of continuous monitoring and expanded research in cyanobacteria ecology, with implications for public health and aquatic resource management.

The roles of rare and abundant microbial species in the primary succession of biological soil crusts are differentiated in metal tailings ponds with different states.

Tailings ponds formed by long-term accumulation of mineral processing waste have become a global environmental problem. Even worse, tailings ponds are often simply abandoned or landfilled after they cease to be used. This allows pollution to persist and continue to spread in the environment. The significance of primary succession mediated by biological soil crusts for tailings pond remediation has been illustrated by previous studies. However, the process of primary succession may not be the same at different stages during the lifetime of tailings ponds. Therefore, we investigated the environmental differences and the successional characteristics of microbial communities in the primary successional stage of tailings ponds at three different states. The results showed that the primary succession process positively changed the environment of tailings ponds in any state of tailings ponds. The primary successional stage determined the environmental quality more than the state of the tailings pond. In the recently abandoned tailings ponds, abundant species were more subjected to heavy metal stress, while rare species were mainly limited by nutrient content. We found that as the succession progressed, rare species gradually acquired their own community space and became more responsive to environmental stresses. Rare species played an important role in microbial keystone species groups.

Prevalence of opportunistic pathogens and anti-microbial resistance in urban aquaculture ponds.

Bacterial antimicrobial resistance (AMR) has emerged as a significant concern worldwide. The microbial community profile and potential AMR level in aquaculture ponds are often undervalued and attract less attention than other aquatic environments. We used amplicon and metagenomic shotgun sequencing to study microbial communities and AMR in six freshwater polyculture ponds in rural and urban areas of Bangladesh. Amplicon sequencing revealed different community structures between rural and urban ponds, with urban ponds having a higher bacterial diversity and opportunistic pathogens including Streptococcus, Staphylococcus, and Corynebacterium. Despite proteobacterial dominance, Firmicutes was the most interactive in the community network, especially in the urban ponds. Metagenomes showed that drug resistance was the most common type of AMR found, while metal resistance was only observed in urban ponds. AMR and metal resistance genes were found mainly in beta and gamma-proteobacteria in urban ponds, while AMR was found primarily in alpha-proteobacteria in rural ponds. We identified potential pathogens with a high profile of AMR and metal resistance in urban aquaculture ponds. As these ponds provide a significant source of protein for humans, our results raise significant concerns for the environmental sustainability of this food source and the dissemination of AMR into the food chain.

Lysobacter stagni sp. nov. and Limnohabitans lacus sp. nov., isolated from a pond.

Two Gram-negative bacteria, designated as strains LF1T and HM2-2T, were isolated from an artificial pond in a honey farm at Hoengseong-gun, Gangwon-do, Republic of Korea. The 16S rRNA sequence analysis results revealed that strain LF1T belonged to the genus Lysobacter and had the highest sequence similarity to Lysobacter niastensis GH41-7T (99.0 %), Lysobacter panacisoli CJ29T (98.9 %), and Lysobacter prati SYSU H10001T (98.2 %). Its growth occurred at 20-37 °C, at pH 5.0-12.0, and in the presence of 0-2% NaCl. The major fatty acids were iso-C15 : 0, iso-C16 : 0, and summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 0 10-methyl). The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, and diphosphatidylglycerol. The DNA G+C content was 67.5 mol%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain LF1T and species of the genus Lysobacter were 79.1-84.4% and 22.0-27.5 %, respectively. The 16S rRNA sequence analysis results revealed that strain HM2-2T belonged to the genus Limnohabitans and was most closely related to Limnohabitans planktonicus II-D5T (98.9 %), Limnohabitans radicicola JUR4T (98.4%), and Limnohabitans parvus II-B4T (98.4 %). Its growth occurred at 10-35 °C, at pH 5.0-11.0, and in the presence of 0-2% NaCl. The major fatty acids were C16 : 0 and summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c). The major polar lipid was phosphatidylethanolamine. The DNA G+C content was 59.9 mol%. The ANI and dDDH values between strain HM2-2T and its closely related strains were 75.1-83.0% and 20.4-26.4 %, respectively. Phenotypic, genomic, and phylogenetic data revealed that strains LF1T and HM2-2T represent novel species in the genera Lysobacter and Limnohabitans, for which the names Lysobacter stagni sp. nov. and Limnohabitans lacus sp. nov. are proposed, respectively. The type strain of Lys. stagni is LF1T (=KACC 23251T=TBRC 17648T), and that of Lim. lacus is HM2-2T (=KACC 23250T=TBRC 17649T).

Fe(II)Cl2 amendment suppresses pond methane emissions by stimulating iron-dependent anaerobic oxidation of methane.

Aquatic ecosystems are large contributors to global methane (CH4) emissions. Eutrophication significantly enhances CH4-production as it stimulates methanogenesis. Mitigation measures aimed at reducing eutrophication, such as the addition of metal salts to immobilize phosphate (PO43-), are now common practice. However, the effects of such remedies on methanogenic and methanotrophic communities-and therefore on CH4-cycling-remain largely unexplored. Here, we demonstrate that Fe(II)Cl2 addition, used as PO43- binder, differentially affected microbial CH4 cycling-processes in field experiments and batch incubations. In the field experiments, carried out in enclosures in a eutrophic pond, Fe(II)Cl2 application lowered in-situ CH4 emissions by lowering net CH4-production, while sediment aerobic CH4-oxidation rates-as found in batch incubations of sediment from the enclosures-did not differ from control. In Fe(II)Cl2-treated sediments, a decrease in net CH4-production rates could be attributed to the stimulation of iron-dependent anaerobic CH4-oxidation (Fe-AOM). In batch incubations, anaerobic CH4-oxidation and Fe(II)-production started immediately after CH4 addition, indicating Fe-AOM, likely enabled by favorable indigenous iron cycling conditions and the present methanotroph community in the pond sediment. 16S rRNA sequencing data confirmed the presence of anaerobic CH4-oxidizing archaea and both iron-reducing and iron-oxidizing bacteria in the tested sediments. Thus, besides combatting eutrophication, Fe(II)Cl2 application can mitigate CH4 emissions by reducing microbial net CH4-production and stimulating Fe-AOM.

Paucibacter sediminis sp. nov., isolated from sediment in a freshwater pond.

A Gram-stain-negative, aerobic, oxidase-positive, weakly catalase-positive, motile by means of a single polar flagellum, rod-shaped bacterium designated as strain S2-9T was isolated from sediment sampled in Wiyang pond, Republic of Korea. Growth of this strain was observed at 10-40 °C (optimum, 35 °C) and pH 5.5-9.5 (optimum, pH 7.0-8.0) and in the presence of 0-0.5 % NaCl in Reasoner's 2A broth. The major fatty acids (>10 %) of strain S2-9T were C16 : 0 and summed feature 3 (comprising a mixture of C16 : 1 ω7c and/or C16 : 1 ω6c). Ubiquinone-8 was detected as the respiratory quinone. The major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. Strain S2-9T showed the highest 16S rRNA gene sequence similarity to Paucibacter oligotrophus CHU3T (98.7 %), followed by 'Paucibacter aquatile' CR182 (98.4 %), all type strains of Pelomonas species (98.1-98.3 %), Mitsuaria chitosanitabida NBRC 102408T (97.9 %), Kinneretia asaccharophila KIN192T (97.8 %), Mitsuaria chitinivorans HWN-4T (97.4 %), and Paucibacter toxinivorans 2C20T (97.4 %). Phylogenetic trees based on the 16S rRNA gene and whole-genome sequences showed that strain S2-9T formed a tight phylogenetic lineage with Paucibacter species (CHU3T, CR182, and 2C20T). Average nucleotide identity and digital DNA-DNA hybridization values between strain S2-9T and Paucibacter strains were 76.6-79.3% and 19.5-21.5 %, respectively. The genomic DNA G+C content of strain S2-9T was 68.3 mol%. Notably, genes responsible for both sulphur oxidation and reduction and denitrification were found in the genome of strain S2-9T, suggesting that strain S2-9T is involved in the nitrogen and sulphur cycles in pond ecosystems. Based on the polyphasic taxonomic results, strain S2-9T represents a novel species of the genus Paucibacter, for which the name Paucibacter sediminis sp. nov. is proposed. The type strain is S2-9T (= KACC 22267T= JCM 34541T).

Fine-scale spatiotemporal variations in bacterial community diversity in agricultural pond water.

Microbial communities in surface waters are affected by environmental conditions and can influence changes in water quality. To explore the hypothesis that the microbiome in agricultural waters associates with spatiotemporal variations in overall water quality and, in turn, has implications for resource monitoring and management, we characterized the relationships between the microbiota and physicochemical properties in a model irrigation pond as a factor of sampling time (i.e., 9:00, 12:00, 15:00) and location within the pond (i.e., bank vs. interior sites and cross-sectional depths at 0, 1, and 2 m). The microbial communities, which were defined by 16S rRNA gene sequencing analysis, significantly varied based on all sampling factors (PERMANOVA P < 0.05 for each). While the relative abundances of dominant phyla (e.g., Proteobacteria and Bacteroidetes) were relatively stable throughout the pond, subtle yet significant increases in α-diversity were observed as the day progressed (ANOVA P < 0.001). Key water quality properties that also increased between the morning and afternoon (i.e., pH, dissolved oxygen, and temperature) positively associated with relative abundances of Cyanobacteria, though were inversely proportional to Verrucomicrobia. These properties, among additional parameters such as bioavailable nutrients (e.g., NH3, NO3, PO4), chlorophyll, phycocyanin, conductivity, and colored dissolved organic matter, exhibited significant relationships with relative abundances of various bacterial genera as well. Further investigation of the microbiota in underlying sediments revealed significant differences between the bank and interior sites of the pond (P < 0.05 for α- and ß-diversity). Overall, our findings emphasize the importance of accounting for time of day and water sampling location and depth when surveying the microbiomes of irrigation ponds and other small freshwater sources.

Exploring the dynamics of algae-associated microbiome during the scale-up process of Tetraselmis sp. microalgae: A metagenomics approach.

Microalgae have become a key source of valuable compounds, promoting commercial scale applications. However, biological contamination is one of the most critical problems associated with large scale algal production, especially in open systems such as raceway ponds. The current research is the first to assess the effectiveness of open raceway ponds in maintaining a pure culture of Tetraselmis sp., starting from 20 L culture up to 10,000 L culture. Microbial profiling of each successive stage revealed lower abundance of eukaryotic organisms, whereas bacterial abundance increased notably resulting in a significant decrease in Tetraselmis sp. abundance. Furthermore, several bacteria with algae growth-promoting properties were found throughout the various culture stages including Balneola, Roseovarius, and Marinobacter. However, some algae-suppressive bacteria were evidenced at later stages such as Ulvibacter, Aestuariicoccus, and Defluviimonas. Overall, due to the increasing bacterial concentration, considerations limiting bacterial contamination need to be taken.

Microbiome analysis reveals Microcystis blooms endogenously seeded from benthos within wastewater maturation ponds.

Toxigenic Microcystis blooms periodically disrupt the stabilization ponds of wastewater treatment plants (WWTPs). Dense proliferations of Microcystis cells within the surface waters (SWs) impede the water treatment process by reducing the treatment efficacy of the latent WWTP microbiome. Further, water quality is reduced when conventional treatment leads to Microcystis cell lysis and the release of intracellular microcystins into the water column. Recurrent seasonal Microcystis blooms cause significant financial burdens for the water industry and predicting their source is vital for bloom management strategies. We investigated the source of recurrent toxigenic Microcystis blooms at Australia's largest lagoon-based municipal WWTP in both sediment core (SC) and SW samples between 2018 and 2020. Bacterial community composition of the SC and SW samples according to 16S rRNA gene amplicon sequencing showed that Microcystis sp. was dominant within SW samples throughout the period and reached peak relative abundances (32%) during the summer. The same Microcystis Amplicon sequence variants were present within the SC and SW samples indicating a potential migratory population that transitions between the sediment water and SWs during bloom formation events. To investigate the potential of the sediment to act as a repository of viable Microcystis cells for recurrent bloom formation, a novel in-vitro bloom model was established featuring sediments and sterilized SW collected from the WWTP. Microcystin-producing Microcystis blooms were established through passive resuspension after 12 weeks of incubation. These results demonstrate the capacity of Microcystis to transition between the sediments and SWs in WWTPs, acting as a perennial inoculum for recurrent blooms.IMPORTANCECyanobacterial blooms are prevalent to wastewater treatment facilities owing to the stable, eutrophic conditions. Cyanobacterial proliferations can disrupt operational procedures through the blocking of filtration apparatus or altering the wastewater treatment plant (WWTP) microbiome, reducing treatment efficiency. Conventional wastewater treatment often results in the lysis of cyanobacterial cells and the release of intracellular toxins which pose a health risk to end users. This research identifies a potential seeding source of recurrent toxigenic cyanobacterial blooms within wastewater treatment facilities. Our results demonstrate the capacity of Microcystis to transition between the sediments and surface waters (SWs) of wastewater treatment ponds enabling water utilities to develop adequate monitoring and management strategies. Further, we developed a novel model to demonstrate benthic recruitment of toxigenic Microcystis under laboratory conditions facilitating future research into the genetic mechanisms behind bloom development.

Sustainable treatment of primary and secondary effluent by algal-bacterial flocculent biomass in raceway ponds.

Two 5.5-L raceway open ponds were used to evaluate the removal of organic material and nutrients from wastewater. Algal-bacterial flocs were placed in the ponds to treat primary and secondary effluent. The organic loading rate ranged from 29 to 95 and 9 to 38 g sCOD m-3 d-1 for the reactor fed with primary and secondary effluent, respectively. The hydraulic retention time (HRT) gradually decreased in both reactors from 5.5 to 2.2 d during a period of 21 days, and after that, both reactors operated at an HRT of 1.1 d. A high biomass concentration of around 2.2 g L-1 was sustained using primary and secondary effluent after 130 days. The biomass, developed with both substrates was very active and completely removed organic material and nutrients in less than 12 h. The algal-bacteria biomass had excellent settling properties and could settle in less than 10 min.

Two machine learning approaches for predicting cyanobacteria abundance in aquaculture ponds.

Cyanobacteria blooms in aquaculture ponds harm the harvesting of aquatic animals and threaten human health. Therefore, it is crucial to identify key drivers and develop methods to predict cyanobacteria blooms in aquaculture water management. In this study, we analyzed monitoring data from 331 aquaculture ponds in central China and developed two machine learning models - the least absolute shrinkage and selection operator (LASSO) regression model and the random forest (RF) model - to predict cyanobacterial abundance by identifying the key drivers. Simulation results demonstrated that both machine learning models are feasible for predicting cyanobacterial abundance in aquaculture ponds. The LASSO model (R2 = 0.918, MSE = 0.354) outperformed the RF model (R2 = 0.798, MSE = 0.875) in predicting cyanobacteria abundance. Farmers with well-equipped aquaculture ponds that have abundant water monitoring data can use the nine environmental variables identified by the LASSO model as an operational solution to accurately predict cyanobacteria abundance. For crude ponds with limited monitoring data, the three environmental variables identified by the RF model provide a convenient solution for useful cyanobacteria prediction. Our findings revealed that chemical oxygen demand (COD) and total organic carbon (TOC) were the two most important predictors in both models, indicating that organic carbon concentration had a close relationship with cyanobacteria growth and should be considered a key metric in water monitoring and pond management of these aquaculture ponds. We suggest that monitoring of organic carbon coupled with phosphorus reduction in feed usage can be an effective management approach for cyanobacteria prevention and to maintain a healthy ecological state in aquaculture ponds.

Responses of bacterial and three sub-microeukaryote communities in the water of white shrimp Penaeus vannamei aquaculture ponds in two polyculture models.

Polyculture operations in freshwater aquaculture ponds can disrupt microbial communities. High-throughput sequencing was used to assess the impact of polyculture operations on bacterial and three sub-microeukaryote communities (fungi, zooplankton, and eukaryotic phytoplankton) in Penaeus vannamei aquaculture ponds containing oriental river prawns and giant freshwater prawns, respectively. The results showed that the bacterial community was less sensitive than the microeukaryote communities to both the polyculture activity and environmental variations. The polyculture of giant freshwater prawns rather than oriental river prawns was the primary factor affecting the beta diversity of the three sub-microeukaryote communities. This may be due to the larger biomass of the polyculture varieties of giant freshwater prawns compared with oriental river prawns. The polyculture activity of giant freshwater prawns with a higher density and that of oriental river prawns with a lower density increased the stochasticity of the community assembly of the three sub-microeukaryote communities. It also affected the topological properties of the microbial communities, including greater correlations between ecosystem elements, and reducing the correlations among zooplanktons. The eukaryotic phytoplankton was the only microbial community that could also be explained by nutrient variation (mainly the total nitrogen). This highlights the potential role of the eukaryotic phytoplankton as a suitable indicator of the effects of nutrient input into ecosystems.

Spatial Versus Nonspatial Variance in Fecal Indicator Bacteria Differs Within and Between Ponds.

Surface water environments are inherently heterogenous, and little is known about variation in microbial water quality between locations. This study sought to understand how microbial water quality differs within and between Virginia ponds. Grab samples were collected twice per week from 30 sampling sites across nine Virginia ponds (n = 600). Samples (100 mL) were enumerated for total coliform (TC) and Escherichia coli (EC) levels, and physicochemical, weather, and environmental data were collected. Bayesian models of coregionalization were used to quantify the variance in TC and EC levels attributable to spatial (e.g., site, pond) versus nonspatial (e.g., date, pH) sources. Mixed-effects Bayesian regressions and conditional inference trees were used to characterize relationships between data and TC or EC levels. Analyses were performed separately for each pond with ≥3 sampling sites (5 intrapond) while one interpond model was developed using data from all sampling sites and all ponds. More variance in TC levels were attributable to spatial opposed to nonspatial sources for the interpond model (variance ratio [VR] = 1.55) while intrapond models were pond dependent (VR: 0.65-18.89). For EC levels, more variance was attributable to spatial sources in the interpond model (VR = 1.62), compared to all intrapond models (VR < 1.0) suggesting that more variance is attributable to nonspatial factors within individual ponds and spatial factors when multiple ponds are considered. Within each pond, TC and EC levels were spatially independent for sites 56-87 m apart, indicating that different sites within the same pond represent different water quality for risk management. Rainfall was positively and pH negatively associated with TC and EC levels in both inter- and intrapond models. For all other factors, the direction and strength of associations varied. Factors driving microbial dynamics in ponds appear to be pond-specific and differ depending on the spatial scale considered.

Strategy for the formation of microalgae-bacteria aggregates in high-rate algal ponds.

This work studied the formation of aggregates used for wastewater treatment in high-rate algal ponds (HRAP). For this, the establishment of microalgae-bacteria aggregates in these systems was evaluated, considering strategies for the inoculation and start-up. Two HRAP were operated in parallel, at first in batch mode and then in continuous flow. The wastewater treatment was efficient, with removal rates around 80% for COD and N-ammoniacal. Volatile suspended solids and chlorophyll for the culture grew continuously reached a concentration of 548 ± 11 mg L-1 and 7.8 mg L-1, respectively. Larger photogranules were observed when the system was placed in a continuous regime. The protein fraction of extracellular polymeric substances was identified as a determinant in photogranules formation. During the continuous regime, more than 50% of the biomass was higher than 0.2 mm, flocculation efficiency of 78 ± 6%, and the volumetric sludge index of 32 ± 5 mL g-1. The genetic sequencing showed the growth of cyanobacteria in the aggregate and the presence of microalgae from the chlorophytes and diatoms groups in the final biomass.