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.

Temperature rise and its influence on the toxic effects caused by cyanotoxins in a neotropical catfish.

Potentially toxic cyanobacterial blooms (cyanoHABs) have become a problem in public water supply reservoirs. Temperature rise caused by climate change can increase the frequency and intensity of blooms, which may influence the cyanotoxins concentration in the environment. This study aimed to evaluate the effect of the temperature on the responses of a Neotropical catfish exposed to a neurotoxin-rich cyanobacterial crude extract (Raphidiopsis raciborskii T3). Juveniles of Rhamdia quelen were exposed to four treatments, based on study data: control at 25 °C (C25), control at 30 °C (C30), crude extract equivalent to 105 cells.mL-l of R. raciborskii at 25 °C (CE25) and 30 °C (CE30). After 96 h of exposure, the fish were anesthetized and blood was taken. After euthanasia, the gill, posterior kidney, brain, muscle, liver and gonad were sampled for hematological, biochemical, genotoxic and histopathological biomarker analysis. Liver was sampled for proteomic analysis for identification of proteins related to energy production. Water samples were collected at the beginning and the end of the experiment for neurotoxins quantification. Different parameters in both males and females were altered at CE25, evidencing the effects of neurotoxins in freshwater fish. At CE30, a water warming scenario, more effects were observed in females than at 25 °C, such as activation of saxitoxin metabolism pathway and genotoxicity. More damage to macromolecules was observed in females at the higher temperature, demonstrating that the increase in temperature can aggravate the toxicity of neurotoxins produced by R. raciborskii T3.

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.

Potential gap in understanding cyanoHABs: Light-dependent morphological variations in colonial cyanobacterium Microcystis.

Colony formation is a crucial characteristic of Microcystis, a cyanobacterium known for causing cyanobacterial harmful algal blooms (cyanoHABs). It has been observed that as Microcystis colonies grow larger, they often become less densely packed, which correlates with a decrease in light penetration. The objective of this study was to investigate the effects of light limitation on the morphological variations in Microcystis, particularly in relation to the crowded cellular environment. The results indicated that when there was sufficient light (transmittance = 100 %) to support a growth rate of 0.11±0.01 day-1, a significant increase in colony size was found, from 466±15 µm to 1030±111 µm. However, under light limitation (transmittance = 50 % - 1 %) where the growth rate was lower than 0, there was no significant improvement in colony size. Microcystis in the light limitation groups exhibited a loose cell arrangement and even the presence of holes or pores within the colony, confirming the negative correlation between colony size and cell arrangement. This pattern is driven by regional differences in growth within the colony, as internal cells have a significantly lower frequency of division compared to peripheral cells, due to intra-colony self-shading (ICSS). The research demonstrates that Microcystis can adjust its cell arrangement to avoid excessive self-shading, which has implications for predicting and controlling cyanoHABs. These findings also contribute to the understanding of cyanobacterial variations and can potentially inform future research on the diverse phycosphere.

Assessment of in-situ monitoring and tracking the vertical migration of cyanobacterial blooms using LISST-HAB.

Cyanobacterial harmful algal blooms (cyanoHABs) are becoming increasingly common in aquatic ecosystems worldwide. However, their heterogeneous distributions make it difficult to accurately estimate the total algae biomass and forecast the occurrence of surface cyanoHABs by using traditional monitoring methods. Although various optical instruments and remote sensing methods have been employed to monitor the dynamics of cyanoHABs at the water surface (i.e., bloom area, chlorophyll a), there is no effective in-situ methodology to monitor the dynamic change of cell density and integrated biovolume of algae throughout the water column. In this study, we propose a quantitative protocol for simultaneously measurements of multiple indicators (i.e., biovolume concentration, size distribution, cell density, and column-integrated biovolume) of cyanoHABs in water bodies by using the laser in-situ scattering and transmissometry (LISST) instrument. The accuracy of measurements of the biovolume and colony size of algae was evaluated and exceeded 95% when the water bloom was dominated by cyanobacteria. Furthermore, the cell density of cyanobacteria was well estimated based on total biovolume and mean cell volume measured by the instrument. Therefore, this methodology has the potential to be used for broader applications, not only to monitor the spatial and temporal distribution of algal biovolume concentration but also monitor the vertical distribution of cell density, biomass and their relationship with size distribution patterns. This provides new technical means for the monitoring and analysis of algae migration and early warning of the formation of cyanoHABs in lakes and reservoirs.

Temporal Dynamics of Cyanobacterial Bloom Community Composition and Toxin Production from Urban Lakes.

With a long evolutionary history and a need to adapt to a changing environment, cyanobacteria in freshwater systems use specialized metabolites for communication, defense, and physiological processes. However, the role that these metabolites play in differentiating species, maintaining microbial communities, and generating niche persistence and expansion is poorly understood. Furthermore, many cyanobacterial specialized metabolites and toxins present significant human health concerns due to their liver toxicity and their potential impact to drinking water. Gaps in knowledge exist with respect to changes in species diversity and toxin production during a cyanobacterial bloom (cyanoHAB) event; addressing these gaps will improve understanding of impacts to public and ecological health. In the current project, we utilized a multiomics strategy (DNA metabarcoding and metabolomics) to determine the cyanobacterial community composition, toxin profile, and the specialized metabolite pool at three freshwater lakes in Providence, RI during summer-fall cyanoHABs. Species diversity decreased at all study sites over the course of the bloom event, and toxin production reached a maximum at the midpoint of the event. Additionally, LC-MS/MS-based molecular networking identified new toxin congeners. This work provokes intriguing questions with respect to the use of allelopathy by organisms in these systems and the presence of emerging toxic compounds that can impact public health.

Environmental drivers behind the exceptional increase in cyanobacterial blooms in Okavango Delta, Botswana.

The Okavango Delta region in Botswana experienced exceptionally intense landscape-wide cyanobacterial harmful algal blooms (CyanoHABs) in 2020. In this study, the drivers behind CyanoHABs were determined from thirteen independent environmental variables, including vegetation indices, climate and meteorological parameters, and landscape variables. Annual Land Use Land Cover (LULC) maps were created from 2017 to 2020, with ∼89% accuracy to compute landscape variables such as LULC change. Generalized Additive Models (GAM) and Structural Equation Models (SEM) were used to determine the most important drivers behind the CyanoHABs. Normalized Difference Chlorophyll Index (NDCI) and Green Line Height (GLH) algorithms served as proxies for chlorophyll-a (green algae) and phycocyanin (cyanobacteria) concentrations. GAM models showed that seven out of the thirteen variables explained 89.9% of the variance for GLH. The models showcased that climate variables, including monthly precipitation (8.8%) and Palmer Severity Drought Index- PDSI (3.2%), along with landscape variables such as changes in Wetlands area (7.5%), and Normalized Difference Vegetation Index (NDVI) (5.4%) were the determining drivers behind the increased cyanobacterial activity within the Delta. Both PDSI and NDVI showed negative correlations with GLH, indicating that increased drought conditions could have led to large increases in toxic CyanoHAB activity within the region. This study provides new information about environmental drivers which can help monitor and predict regions at risk of future severe CyanoHABs outbreaks in the Okavango Delta, Botswana, and other similar data-scarce and ecologically sensitive areas in Africa. Plain Language Summary: The waters of the Okavango Delta in Northern Botswana experienced an exceptional increase in toxic cyanobacterial activity in recent years. Cyanobacterial blooms have been shown to affect local communities and wildlife in the past. To determine the drivers behind this increased bloom activity, we analyzed the effects of thirteen independent environmental variables using two different statistical models. Within this research, we focused on vegetation indices, meteorological, and landscape variables, as previous studies have shown their effect on cyanobacterial activity in other parts of the world. While driver determination for cyanobacteria has been done before, the environmental conditions most important for cyanobacterial growth can be specific to the geographic setting of a study site. The statistical analysis indicated that the increases in cyanobacterial bloom activity within the region were mainly driven by persistent drier conditions. To our knowledge, this is the first study to determine the driving factors behind cyanobacterial activity in this region of the world. Our findings will help to predict and monitor areas at risk of future severe cyanobacterial blooms in the Okavango Delta and other similar African ecosystems.