Alleviation of H2O2 toxicity by extracellular catalases in the phycosphere of Microcystis aeruginosa.

High levels of environmental H2O2 represent a threat to many freshwater bacterial species, including toxic-bloom-forming Microcystis aeruginosa, particularly under high-intensity light conditions. The highest extracellular catalase activity-possessing Pseudoduganella aquatica HC52 was chosen among 36 culturable symbiotic isolates from the phycosphere in freshly collected M. aeruginosa cells. A zymogram for catalase activity revealed the presence of only one extracellular catalase despite the four putative catalase genes (katA1, katA2, katE, and srpA) identified in the newly sequenced genome (∼6.8 Mb) of P. aquatica HC52. Analysis of secreted catalase using liquid chromatography-tandem mass spectrometry was identified as KatA1, which lacks a typical signal peptide, although the underlying mechanism for its secretion is unknown. The expression of secreted KatA1 appeared to be induced in the presence of H2O2. Proteomic analysis also confirmed the presence of KatA1 inside the outer membrane vesicles secreted by P. aquatica HC52 following exposure to H2O2. High light intensities (> 100 µmol m-2 s-1) are known to kill catalase-less axenic M. aeruginosa cells, but the present study found that the presence of P. aquatica cells supported the growth of M. aeruginosa, while the extracellular catalases in supernatant or purified form also sustained the growth of M. aeruginosa under the same conditions. Our results suggest that the extracellular catalase secreted by P. aquatica HC52 enhances the tolerance of M. aeruginosa to H2O2, thus promoting the formation of M. aeruginosa blooms under high light intensities.

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.

Seasonal dynamics of toxigenic Microcystis in a large, shallow Lake Peipsi (Estonia) using microcystin mcyE gene abundance.

Large and temperate Lake Peipsi is the fourth largest lake in Europe, where the massive cyanobacterial blooms are composed mostly of Microcystis spp., which have been common for several decades now. The seasonal dynamics of potentially toxic Microcystis were studied using microscopy and quantitative polymerase chain reaction (qPCR) by assessing the microcystin-encoding microcystin synthetase gene E (mcyE) abundances. Water samples were analyzed over the lake areas, varying in depth, trophic level, and cyanobacterial composition during the growing period of 2021. The Microcystis mcyE genes were detected through the growing period (May-October), forming peak abundances in September with decreasing temperatures (8.9-11.1 °C). Total phosphorus (TP) and nitrate (NO3-) were the most relevant environmental variables influencing the Microcystis biomass as well as mcyE abundances. Comparison with previous years (2011, 2012) indicated that the abundance and seasonal dynamics of toxigenic Microcystis can be highly variable between the years and lake areas, varying also in dominant Microcystis species. Contrary to expectations, based on mcyE abundances, the increased risk of toxin-producing Microcystis can occur in Peipsi through the growing period, independently of the water temperature and biomasses of Microcystis.

Metabologenomics reveals strain-level genetic and chemical diversity of Microcystis secondary metabolism.

Microcystis spp. are renowned for producing the hepatotoxin microcystin in freshwater cyanobacterial harmful algal blooms around the world, threatening drinking water supplies and public and environmental health. However, Microcystis genomes also harbor numerous biosynthetic gene clusters (BGCs) encoding the biosynthesis of other secondary metabolites, including many with toxic properties. Most of these BGCs are uncharacterized and currently lack links to biosynthesis products. However, recent field studies show that many of these BGCs are abundant and transcriptionally active in natural communities, suggesting potentially important yet unknown roles in bloom ecology and water quality. Here, we analyzed 21 xenic Microcystis cultures isolated from western Lake Erie to investigate the diversity of the biosynthetic potential of this genus. Through metabologenomic and in silico approaches, we show that these Microcystis strains contain variable BGCs, previously observed in natural populations, and encode distinct metabolomes across cultures. Additionally, we find that the majority of metabolites and gene clusters are uncharacterized, highlighting our limited understanding of the chemical repertoire of Microcystis spp. Due to the complex metabolomes observed in culture, which contain a wealth of diverse congeners as well as unknown metabolites, these results underscore the need to deeply explore and identify secondary metabolites produced by Microcystis beyond microcystins to assess their impacts on human and environmental health.IMPORTANCEThe genus Microcystis forms dense cyanobacterial harmful algal blooms (cyanoHABs) and can produce the toxin microcystin, which has been responsible for drinking water crises around the world. While microcystins are of great concern, Microcystis also produces an abundance of other secondary metabolites that may be of interest due to their potential for toxicity, ecological importance, or pharmaceutical applications. In this study, we combine genomic and metabolomic approaches to study the genes responsible for the biosynthesis of secondary metabolites as well as the chemical diversity of produced metabolites in Microcystis strains from the Western Lake Erie Culture Collection. This unique collection comprises Microcystis strains that were directly isolated from western Lake Erie, which experiences substantial cyanoHAB events annually and has had negative impacts on drinking water, tourism, and industry.

Algae blooms with resistance in fresh water: Potential interplay between Microcystis and antibiotic resistance genes.

Microcystis, a type of cyanobacteria known for producing microcystins (MCs), is experiencing a global increase in blooms. They have been recently recognized as potential contributors to the widespread of antibiotic resistance genes (ARGs). By reviewing approximately 150 pieces of recent studies, a hypothesis has been formulated suggesting that significant fluctuations in MCs concentrations and microbial community structure during Microcystis blooms could influence the dynamics of waterborne ARGs. Among all MCs, microcystin-LR (MC-LR) is the most widely distributed worldwide, notably abundant in reservoirs during summer. MCs inhibit protein phosphatases or increase reactive oxygen species (ROS), inducing oxidative stresses, enhancing membrane permeability, and causing DNA damage. This further enhances selective pressures and horizontal gene transfer (HGT) chances of ARGs. The mechanisms by which Microcystis regulates ARG dissemination have been systematically organized for the first time, focusing on the secretion of MCs and the alterations of bacterial community structure. However, several knowledge gaps remain, particularly concerning how MCs interfere with the electron transport chain and how Microcystis facilitates HGT of ARGs. Concurrently, the predominance of Microcystis forming the algal microbial aggregates is considered a hotspot for preserving and transferring ARGs. Yet, Microcystis can deplete the nutrients from other taxa within these aggregates, thereby reducing the density of ARG-carrying bacteria. Therefore, further studies are needed to explore the 'symbiotic - competitive' relationships between Microcystis and ARG-hosting bacteria under varied nutrient conditions. Addressing these knowledge gaps is crucial to understand the impacts of the algal aggregates on dynamics of waterborne antibiotic resistome, and underscores the need for effective control of Microcystis to curb the spread of antibiotic resistance. Constructed wetlands and photocatalysis represent advantageous strategies for halting the spread of ARGs from the perspective of Microcystis blooms, as they can effectively control Microcystis and MCs while maintaining the stability of aquatic ecosystem.

Toxic mechanisms of the antiviral drug arbidol on microalgae in algal bloom water at transcriptomic level.

Increasing antivirals in surface water caused by their excessive consumption pose serious threats to aquatic organisms. Our recent research found that the input of antiviral drug arbidol to algal bloom water can induce acute toxicity to the growth and metabolism of Microcystis aeruginosa, resulting in growth inhibition, as well as decrease in chlorophyll and ATP contents. However, the toxic mechanisms involved remained obscure, which were further investigated through transcriptomic analysis in this study. The results indicated that 885-1248 genes in algae were differentially expressed after exposure to 0.01-10.0 mg/L of arbidol, with the majority being down-regulated. Analysis of commonly down-regulated genes found that the cellular response to oxidative stress and damaged DNA bonding were affected, implying that the stress defense system and DNA repair function of algae might be damaged. The down-regulation of genes in porphyrin metabolism, photosynthesis, carbon fixation, glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation might inhibit chlorophyll synthesis, photosynthesis, and ATP supply, thereby hindering the growth and metabolism of algae. Moreover, the down-regulation of genes related to nucleotide metabolism and DNA replication might influence the reproduction of algae. These findings provided effective strategies to elucidate toxic mechanisms of contaminants on algae in algal bloom water.

Profiling the interplay and coevolution of Microcystis aeruginosa and cyanosiphophage Mic1.

The cyanosiphophage Mic1 specifically infects the bloom-forming Microcystis aeruginosa FACHB 1339 from Lake Chaohu, China. Previous genomic analysis showed that its 92,627 bp double-stranded DNA genome consists of 98 putative open reading frames, 63% of which are of unknown function. Here, we investigated the transcriptome dynamics of Mic1 and its host using RNA sequencing. In the early, middle, and late phases of the 10 h lytic cycle, the Mic1 genes are sequentially expressed and could be further temporally grouped into two distinct clusters in each phase. Notably, six early genes, including gp49 that encodes a TnpB-like transposase, immediately reach the highest transcriptional level in half an hour, representing a pioneer cluster that rapidly regulates and redirects host metabolism toward the phage. An in-depth analysis of the host transcriptomic profile in response to Mic1 infection revealed significant upregulation of a polyketide synthase pathway and a type III-B CRISPR system, accompanied by moderate downregulation of the photosynthesis and key metabolism pathways. The constant increase of phage transcripts and relatively low replacement rate over the host transcripts indicated that Mic1 utilizes a unique strategy to gradually take over a small portion of host metabolism pathways after infection. In addition, genomic analysis of a less-infective Mic1 and a Mic1-resistant host strain further confirmed their dynamic interplay and coevolution via the frequent horizontal gene transfer. These findings provide insights into the mutual benefit and symbiosis of the highly polymorphic cyanobacteria M. aeruginosa and cyanophages. IMPORTANCE: The highly polymorphic Microcystis aeruginosa is one of the predominant bloom-forming cyanobacteria in eutrophic freshwater bodies and is infected by diverse and abundant cyanophages. The presence of a large number of defense systems in M. aeruginosa genome suggests a dynamic interplay and coevolution with the cyanophages. In this study, we investigated the temporal gene expression pattern of Mic1 after infection and the corresponding transcriptional responses of its host. Moreover, the identification of a less-infective Mic1 and a Mic1-resistant host strain provided the evolved genes in the phage-host coevolution during the multiple-generation cultivation in the laboratory. Our findings enrich the knowledge on the interplay and coevolution of M. aeruginosa and its cyanophages and lay the foundation for the future application of cyanophage as a potential eco-friendly and bio-safe agent in controlling the succession of harmful cyanobacterial blooms.

Aerobic anoxygenic phototrophs play important roles in nutrient cycling within cyanobacterial Microcystis bloom microbiomes.

BACKGROUND: During the bloom season, the colonial cyanobacterium Microcystis forms complex aggregates which include a diverse microbiome within an exopolymer matrix. Early research postulated a simple mutualism existing with bacteria benefitting from the rich source of fixed carbon and Microcystis receiving recycled nutrients. Researchers have since hypothesized that Microcystis aggregates represent a community of synergistic and interacting species, an interactome, each with unique metabolic capabilities that are critical to the growth, maintenance, and demise of Microcystis blooms. Research has also shown that aggregate-associated bacteria are taxonomically different from free-living bacteria in the surrounding water. Moreover, research has identified little overlap in functional potential between Microcystis and members of its microbiome, further supporting the interactome concept. However, we still lack verification of general interaction and know little about the taxa and metabolic pathways supporting nutrient and metabolite cycling within Microcystis aggregates. RESULTS: During a 7-month study of bacterial communities comparing free-living and aggregate-associated bacteria in Lake Taihu, China, we found that aerobic anoxygenic phototrophic (AAP) bacteria were significantly more abundant within Microcystis aggregates than in free-living samples, suggesting a possible functional role for AAP bacteria in overall aggregate community function. We then analyzed gene composition in 102 high-quality metagenome-assembled genomes (MAGs) of bloom-microbiome bacteria from 10 lakes spanning four continents, compared with 12 complete Microcystis genomes which revealed that microbiome bacteria and Microcystis possessed complementary biochemical pathways that could serve in C, N, S, and P cycling. Mapping published transcripts from Microcystis blooms onto a comprehensive AAP and non-AAP bacteria MAG database (226 MAGs) indicated that observed high levels of expression of genes involved in nutrient cycling pathways were in AAP bacteria. CONCLUSIONS: Our results provide strong corroboration of the hypothesized Microcystis interactome and the first evidence that AAP bacteria may play an important role in nutrient cycling within Microcystis aggregate microbiomes. Video Abstract.

Sphingomonas lacusdianchii sp. nov., an attached bacterium inhibited by metabolites from its symbiotic cyanobacterium.

An alpha-proteobacterial strain JXJ CY 53 T was isolated from the cyanosphere of Microcystis sp. FACHB-905 (MF-905) collected from Lake Dianchi, China. JXJ CY 53 T was observed to be an aerobic, Gram-stain-negative, oval shaped, and mucus-secreting bacterium. It had C18:1ω7c and C16:0 as the major cellular fatty acids, Q-10 as the predominant ubiquinone, and sphingoglycolipid, diphosphatidylglycerol, phosphatidylcholine, and phosphatidylmethylethanolamine as the polar lipids. The G + C content of DNA was 65.85%. The bacterium had 16S rRNA gene sequence identities of 98.9% and 98.7% with Sphingomonas panni DSM 15761 T and Sphingomonas hankookensis KCTC 22579 T, respectively, while less than 97.4% identities with other members of the genus. Further taxonomic analysis indicated that JXJ CY 53 T represented a new member of Sphingomonas, and the species epithet was proposed as Sphingomonas lacusdianchii sp. nov. (type strain JXJ CY 53 T = KCTC 72813 T = CGMCC 1.17657 T). JXJ CY 53 T promoted the growth of MF-905 by providing bio-available phosphorus and nitrogen, plant hormones, vitamins, and carotenoids. It could modulate the relative abundances of nonculturable bacteria associated with MF-905 and influence the interactions of MF-905 and other bacteria isolated from the cyanobacterium, in addition to microcystin production characteristics. Meanwhile, MF-905 could provide JXJ CY 53 T dissolved organic carbon for growth, and control the growth of JXJ CY 53 T by secreting specific chemicals other than microcystins. Overall, these results suggest that the interactions between Microcystis and its attached bacteria are complex and dynamic, and may influence the growth characteristics of the cyanobacterium. This study provided new ideas to understand the interactions between Microcystis and its attached bacteria. KEY POINTS: • A novel bacterium (JXJCY 53 T) was isolated from the cyanosphere of Microcystis sp. FACHB-905 (MF-905) • JXJCY 53 T modulated the growth and microcystin production of MF-905 • MF-905 could control the attached bacteria by specific chemicals other than microcystins (MCs).

Phenotypic and Genotypic Methods for the Identification and Quantification of Cyanobacteria in Lake Water.

Early monitoring of Microcystis, a cyanobacterium that produces microcystin, is paramount in order to confirm the presence of Microcystis spp. Both phenotypic and genotypic methods have been used. The phenotypic methods provide the presence of the microcystis but do not confirm its species type and toxin produced. Additionally, phenotypic methods cannot differentiate toxigenic from non-toxigenic Microcystis. Therefore, the current protocol also describes genetic methods based on PCR to detect toxigenic Microcystis spp. based on microcystin synthetase E (mcy E) gene and 16-23S RNA genes for species-specific identification, which can effectively comprehend distinct lineages and discrimination of potential complexity of microcystin populations. The presence of these microcystin toxins in blood, in most cases, indicates contamination of drinking water by cyanobacteria. The methods presented herein are used to identify microcystin toxins in drinking water and blood.

The synchronicity of bloom-forming cyanobacteria transcription patterns and hydrogen peroxide dynamics.

Hydrogen peroxide is a reactive oxygen species (ROS) naturally occurring at low levels in aquatic environments and production varies widely across different ecosystems. Oxygenic photosynthesis generates hydrogen peroxide as a byproduct, of which some portion can be released to ambient water. However, few studies have examined hydrogen peroxide dynamics in relation to cyanobacterial harmful algal blooms (cHABs). A year-long investigation of algal succession and hydrogen peroxide dynamics was conducted at the Caloosahatchee River, Florida, USA. We aimed to identify potential biological mechanisms responsible for elevated hydrogen peroxide production during cHAB events through the exploration of the freshwater microbial metatranscriptome. Hydrogen peroxide concentrations were elevated from February to September of 2021 when cyanobacteria were active and abundant. We observed one Microcystis cHAB event in spring and one in winter. Both had distinct nutrient uptake and cyanotoxin gene expression patterns. While meaningful levels of microcystin were only detected during periods of elevated hydrogen peroxide, cyanopeptolin was by far the most expressed cyanotoxin during the spring bloom when hydrogen peroxide was at its yearly maxima. Gene expressions of five microbial enzymes (Rubisco, superoxide dismutase, cytochrome b559, pyruvate oxidase, and NADH dehydrogenase) positively correlated to hydrogen peroxide concentrations. Additionally, there was higher nitrogen-fixing gene (nifDKH) expression by filamentous cyanobacteria after the spring bloom but no secondary bloom formation occurred. Overall, elevated environmental hydrogen peroxide concentrations were linked to cyanobacterial dominance and greater expression of specific enzymes in the photosynthesis of cyanobacteria. This implicates cyanobacterial photosynthesis and growth results in increased hydrogen peroxide generation as reflected in measured environmental concentrations.

CyanoStrainChip: A Novel DNA Microarray Tool for High-Throughput Detection of Environmental Cyanobacteria at the Strain Level.

Detecting cyanobacteria in environments is an important concern due to their crucial roles in ecosystems, and they can form blooms with the potential to harm humans and nonhuman entities. However, the most widely used methods for high-throughput detection of environmental cyanobacteria, such as 16S rRNA sequencing, typically provide above-species-level resolution, thereby disregarding intraspecific variation. To address this, we developed a novel DNA microarray tool, termed the CyanoStrainChip, that enables strain-level comprehensive profiling of environmental cyanobacteria. The CyanoStrainChip was designed to target 1277 strains; nearly all major groups of cyanobacteria are included by implementing 43,666 genome-wide, strain-specific probes. It demonstrated strong specificity by in vitro mock community experiments. The high correlation (Pearson's R > 0.97) between probe fluorescence intensities and the corresponding DNA amounts (ranging from 1-100 ng) indicated excellent quantitative capability. Consistent cyanobacterial profiles of field samples were observed by both the CyanoStrainChip and next-generation sequencing methods. Furthermore, CyanoStrainChip analysis of surface water samples in Lake Chaohu uncovered a high intraspecific variation of abundance change within the genus Microcystis between different severity levels of cyanobacterial blooms, highlighting two toxic Microcystis strains that are of critical concern for Lake Chaohu harmful blooms suppression. Overall, these results suggest a potential for CyanoStrainChip as a valuable tool for cyanobacterial ecological research and harmful bloom monitoring to supplement existing techniques.

Bacterial community shifts induced by high concentration hydrogen peroxide treatment of Microcystis bloom in a mesocosm study.

Hydrogen peroxide has gained popularity as an environmentally friendly treatment for cyanobacterial harmful algal blooms (cHABs) that takes advantage of oxidative stress sensitivity in cyanobacteria at controlled concentrations. Higher concentrations of hydrogen peroxide treatments may seem appealing for more severe cHABs but there is currently little understanding of the environmental impacts of this approach. Of specific concern is the associated microbial community, which may play key roles in the succession/recovery process post-treatment. To better understand impacts of a high concentration treatment on non-target microbial communities, we applied a hydrogen peroxide spray equating to a total volume concentration of 14 mM (473 mg/L, 0.04%) to 250 L mesocosms containing Microcystis bloom biomass, monitoring treatment and control mesocosms for 4 days. Cyanobacteria dominated control mesocosms throughout the experiment while treatment mesocosms experienced a 99% reduction, as determined by bacterial amplicon sequencing, and a 92% reduction in bacterial cell density within 1 day post-treatment. Only the bacterial community exhibited signs of regrowth, with a fold change of 9.2 bacterial cell density from day 1 to day 2. Recovery consisted of succession by Planctomycetota (47%) and Gammaproteobacteria (17%), which were likely resilient due to passive cell component compartmentalization and rapid upregulation of dnaK and groEL oxidative stress genes, respectively. The altered microbiome retained beneficial functionality of microcystin degradation through a currently recognized but unidentified pathway in Gammaproteobacteria, resulting in a 70% reduction coinciding with bacterial regrowth. There was also an 81% reduction of both total nitrogen and phosphorus, as compared to 91 and 93% in the control, respectively, due to high expressions of genes related to nitrogen (argH, carB, glts, glnA) and phosphorus (pntAB, phoB, pstSCB) cycling. Overall, we found a portion of the bacterial community was resilient to the high-concentration hydrogen peroxide treatment, resulting in Planctomycetota and Gammaproteobacteria dominance. This high-concentration treatment may be suitable to rapidly end cHABs which have already negatively impacted the aquatic environment rather than allow them to persist.

The Microcystis-microbiome interactions: origins of the colonial lifestyle.

Species of the Microcystis genus are the most common bloom-forming toxic cyanobacteria worldwide. They belong to a clade of unicellular cyanobacteria whose ability to reach high biomasses during blooms is linked to the formation of colonies. Colonial lifestyle provides several advantages under stressing conditions of light intensity, ultraviolet light, toxic substances and grazing. The progression from a single-celled organism to multicellularity in Microcystis has usually been interpreted as individual phenotypic responses of the cyanobacterial cells to the environment. Here, we synthesize current knowledge about Microcystis colonial lifestyle and its role in the organism ecology. We then briefly review the available information on Microcystis microbiome and propose that changes leading from single cells to colonies are the consequence of specific and tightly regulated signals between the cyanobacterium and its microbiome through a biofilm-like mechanism. The resulting colony is a multi-specific community of interdependent microorganisms.

Comparative genomic analysis of Microcystis strain diversity using conserved marker genes.

Microcystis-dominated cyanobacterial harmful algal blooms (cyanoHABs) have a global impact on freshwater environments, affecting both wildlife and human health. Microcystis diversity and function in field samples and laboratory cultures can be determined by sequencing whole genomes of cultured isolates or natural populations, but these methods remain computationally and financially expensive. Amplicon sequencing of marker genes is a lower cost and higher throughput alternative to characterize strain composition and diversity in mixed samples. However, the selection of appropriate marker gene region(s) and primers requires prior understanding of the relationship between single gene genotype, whole genome content, and phenotype. To identify phylogenetic markers of Microcystis strain diversity, we compared phylogenetic trees built from each of 2,351 individual core genes to an established phylogeny and assessed the ability of these core genes to predict whole genome content and bioactive compound genotypes. We identified single-copy core genes better able to resolve Microcystis phylogenies than previously identified marker genes. We developed primers suitable for current Illumina-based amplicon sequencing with near-complete coverage of available Microcystis genomes and demonstrate that they outperform existing options for assessing Microcystis strain composition. Results showed that genetic markers can be used to infer Microcystis gene content and phenotypes such as potential production of bioactive compounds , although marker performance varies by bioactive compound gene and sequence similarity. Finally, we demonstrate that these markers can be used to characterize the Microcystis strain composition of laboratory or field samples like those collected for surveillance and modeling of Microcystis-dominated cyanobacterial harmful algal blooms.

Hormetic effect of a short-chain PFBS on Microcystis aeruginosa and its molecular mechanism.

Short-chain Perfluorinated compounds (PFCs), used as substitutes for highly toxic long-chain PFCs, are increasingly entering the aquatic environment. However, the toxicity of short-chain PFCs in the environment is still controversial. This study investigated the effects of short-chain perfluorobutanesulfonic acid (PFBS) at different concentrations (2.5, 6, 14.4, 36, and 90 mg/L) on M. aeruginosa growth under 12-day exposure and explored the molecular mechanism of toxicity using transcriptomics. The results showed that M. aeruginosa exhibited hormetic effects after exposure to PFBS. Low PFBS concentrations stimulated algal growth, whereas high PFBS concentrations inhibited it, and this inhibitory effect became progressively more pronounced with increasing PFBS exposure concentrations. Transcriptomics showed that PFBS promoted the pathways of photosynthesis, glycolysis, energy metabolism and peptidoglycan synthesis, providing the energy required for cell growth and maintaining cellular morphology. PFBS, on the other hand, caused growth inhibition in algae mainly through oxidative stress, streptomycin synthesis, and genetic damage. Our findings provide new insights into the toxicity and underlying mechanism of short-chain PFCs on algae and inform the understanding of the hormetic effect of short-chain PFCs, which are crucial for assessing their ecological risks in aquatic environments.

Harmful algal blooms in Cayuga lake, NY: From microbiome analysis to eDNA monitoring.

The global increase in harmful algal blooms (HABs) has become a growing concern over the years, and New York State (NYS) is no exception. The Finger Lakes region in NYS has been identified as a hotspot for HABs, with Cayuga Lake having the highest number of blooms reported. The Cayuga Lake HABs Monitoring Program has been tracking cHABs (dominant bloom taxa, chlorophyll A, and microcystin levels) since 2018. However, limited research has been conducted on the microbiome of HABs in this region. In this study, the microbiome of HABs in the Cayuga Lake was surveyed and compared with non-HAB baseline samples. Using 16S rDNA community analysis, common bloom-forming cyanobacteria, were identified, with Microcystis being the dominant taxa in high toxin blooms. Further, this study evaluated the ability of Microcystis mcyA qPCR to detect elevated levels of potential toxigenic Microcystis in water samples using both benchtop and handheld qPCR devices. The results showed good performance of the qPCR assay as a screening for high toxin versus low/no toxin blooms. Additionally, the handheld qPCR device holds potential for in-field rapid (<1 h) screenings for high toxin blooms. This study provides insights into the microbiome of HABs in Cayuga Lake and offers a potential tool for rapid screening of high toxin blooms.

Response of particle-attached and free-living bacterial communities to Microcystis blooms.

The massive proliferation of Microcystis threatens freshwater ecosystems and degrades water quality globally. Understanding the mechanisms that contribute to Microcystis growth is crucial for managing Microcystis blooms. The lifestyles of bacteria can be classified generally into two groups: particle-attached (PA; > 3 µm) and free-living (FL; 0.2-3.0 µm). However, little is known about the response of PA and FL bacteria to Microcystis blooms. Using 16S rRNA gene high-throughput sequencing, we investigated the stability, assembly process, and co-occurrence patterns of PA and FL bacterial communities during distinct bloom stages. PA bacteria were phylogenetically different from their FL counterparts. Microcystis blooms substantially influenced bacterial communities. The time decay relationship model revealed that Microcystis blooms might increase the stability of both PA and FL bacterial communities. A contrasting community assembly mechanism was observed between the PA and FL bacterial communities. Throughout Microcystis blooms, homogeneous selection was the major assembly process that impacted the PA bacterial community, whereas drift explained much of the turnover of the FL bacterial community. Both PA and FL bacterial communities could be separated into modules related to different phases of Microcystis blooms. Microcystis blooms altered the assembly process of PA and FL bacterial communities. PA bacterial community appeared to be more responsive to Microcystis blooms than FL bacteria. Decomposition of Microcystis blooms may enhance cooperation among bacteria. Our findings highlight the importance of studying bacterial lifestyles to understand their functions in regulating Microcystis blooms. KEY POINTS: • Microcystis blooms alter the assembly process of PA and FL bacterial communities • Microcystis blooms increase the stability of both PA and FL bacterial communities • PA bacteria seem to be more responsive to Microcystis blooms than FL bacteria.

Devosia lacusdianchii sp. nov., an attached bacterium inhibited by metabolites from its symbiotic Microcystis.

A novel alphaproteobacterial strain JXJ CY 41T was isolated from a culture mass of Microcystis, collected from Lake Dianchi, south-west, China. Strain JXJ CY 41T was gram-strain-negative, aerobic, motile, with rod-shaped cells (0.4-1.0 × 1.7-3.5 µm). It was positive for catalase and starch hydrolysis, negative for oxidase and hydrolysis of Tweens (20, 40, and 80). Growth occurred at 10-44 °C, pH 5.0-10.0, and 0-5.0% (w/v) NaCl. Major fatty acids included C16:0 (28.1%), 11-methyl C18:1 ω7c (36.7%) and C18:1 ω7c (20.8%). Q10 was the sole ubiquinone. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, glycolipid, and an unidentified lipid. The DNA G + C content was 63.1%. Its 16S rRNA gene sequence showed high similarities with Devosia oryziradicis G19T (99.5%; not validly published), D. yakushimensis Yak96BT (98.3%) and D. ginsengisoli Gsoil 520T (98.1%), and less than 98.1% similarities with other members of the genus Devosia. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strain JXJ CY 41T and its 5 closest similar strains were 19.9-24.1% and 75.7-80.5%, respectively. Based on the data above, strain JXJ CY 41T was identified as a novel species of the genus Devosia, for which the epithet Devosia lacusdianchii sp. nov. was proposed. The type strain is JXJ CY 41T (= KCTC 72812T = CGMCC 1.17502T). Strain JXJ CY 41T exhibited different interactions with Microcystis aeruginosa FACHB-905 (Maf) under different conditions, and Maf could control the bacterial cellular density by secreting unknown specific chemical compounds according to its nutritional requirements.

Spatio-temporal variation of toxin-producing gene abundance in Microcystis aeruginosa from Poyang Lake.

Microcystis aeruginosa (M. aeruginosa) causes massive blooms in eutrophic freshwater and releases microcystin. Poyang Lake is the largest freshwater lake in China and has kept a mid-nutrient level in recent years. However, there is little research on microcystin production in Poyang Lake. In this study, water and sediment samples from ten sampling sites in Poyang Lake were collected from May to December in 2020, and from January to April in 2021 respectively. Microcystis genes (mcyA, mcyB, 16 s rDNA) were quantified by real-time fluorescence quantitative PCR analysis, and then the spatial and temporal variation of mcy genes, physicochemical factors, and bacterial population structure in the lake was analyzed. The relationship between the abundance of mcy genes and physicochemical factors in water column was also revealed. Results indicated that the microcystin-producing genes mcyA and mcyB showed significant differences in spatial and temporal levels as well, which is closely related to the physicochemical factors especially the water temperature (p < 0.05) and the nitrogen content (p < 0.05). The abundance of mcy genes in the sediment in December affected the abundance of mcy genes in the water column in the next year, while the toxic Microcystis would accumulate in the sediment. In addition to the toxic Microcystis, we also found a large number of non-toxic Microcystis in the water column and sediment, and the ratio of toxic to non-toxic species can also affect the toxicity production of M. aeruginosa. Overall, the results showed that M. aeruginosa toxin-producing genes in Poyang Lake distributed spatially and temporally which related to the physicochemical factors of Poyang Lake.