The haplotype-resolved Prymnesium parvum (type B) microalga genome reveals the genetic basis of its fish-killing toxins.

The catastrophic loss of aquatic life in the Central European Oder River in 2022, caused by a toxic bloom of the haptophyte microalga Prymnesium parvum (in a wide sense, s.l.), underscores the need to improve our understanding of the genomic basis of the toxin. Previous morphological, phylogenetic, and genomic studies have revealed cryptic diversity within P. parvum s.l. and uncovered three clade-specific (types A, B, and C) prymnesin toxins. Here, we used state-of-the-art long-read sequencing and assembled the first haplotype-resolved diploid genome of a P. parvum type B from the strain responsible for the Oder disaster. Comparative analyses with type A genomes uncovered a genome-size expansion driven by repetitive elements in type B. We also found conserved synteny but divergent evolution in several polyketide synthase (PKS) genes, which are known to underlie toxin production in combination with environmental cues. We identified an approximately 20-kbp deletion in the largest PKS gene of type B that we link to differences in the chemical structure of types A and B prymnesins. Flow cytometry and electron microscopy analyses confirmed diploidy in the Oder River strain and revealed differences to closely related strains in both ploidy and morphology. Our results provide unprecedented resolution of strain diversity in P. parvum s.l. and a better understanding of the genomic basis of toxin variability in haptophytes. The reference-quality genome will enable us to better understand changes in microbial diversity in the face of increasing environmental pressures and provides a basis for strain-level monitoring of invasive Prymnesium in the future.

The Effect of pH and Salinity on the Toxicity and Growth of the Golden Alga, Prymnesium parvum.

Bioassays using cultures of the toxic haptophyte Prymnesium parvum and the ciliate Cyclidium sp. as prey were conducted to test the effect of pH (range = 6.5 - 8.5), salinity (range = 1.50 - 7.50‰), and a combination of pH and salinity on the toxicity of P. parvum. pH had a significant effect on P. parvum toxicity. Toxicity was rapidly (within 24 hr) induced by increasing pH of the medium, or reduced by lowering pH. Conversely, lowering salinity reduced toxicity, albeit less effectively compared to pH, and P. parvum cells remained toxic at the lowest values tested (1.50‰ at pH 7.5). An additional effect between pH and salinity was also observed: low salinity combined with low pH led to not only decreased toxicity, but also resulted in lower P. parvum growth rates. Such effects of pH and salinity on P. parvum growth and toxicity provide insight into the environmental factors supporting community dominance and toxic blooms of the alga.

Gill Transcriptomic Responses to Toxin-producing Alga Prymnesium parvum in Rainbow Trout.

The gill of teleost fish is a multifunctional organ involved in many physiological processes, including protection of the mucosal gill surface against pathogens and other environmental antigens by the gill-associated lymphoid tissue (GIALT). Climate change associated phenomena, such as increasing frequency and magnitude of harmful algal blooms (HABs) put extra strain on gill function, contributing to enhanced fish mortality and fish kills. However, the molecular basis of the HAB-induced gill injury remains largely unknown due to the lack of high-throughput transcriptomic studies performed on teleost fish in laboratory conditions. We used juvenile rainbow trout (Oncorhynchus mykiss) to investigate the transcriptomic responses of the gill tissue to two (high and low) sublethal densities of the toxin-producing alga Prymnesium parvum, in relation to non-exposed control fish. The exposure time to P. parvum (4-5 h) was sufficient to identify three different phenotypic responses among the exposed fish, enabling us to focus on the common gill transcriptomic responses to P. parvum that were independent of dose and phenotype. The inspection of common differentially expressed genes (DEGs), canonical pathways, upstream regulators and downstream effects pointed towards P. parvum-induced inflammatory response and gill inflammation driven by alterations of Acute Phase Response Signalling, IL-6 Signalling, IL-10 Signalling, Role of PKR in Interferon Induction and Antiviral Response, IL-8 Signalling and IL-17 Signalling pathways. While we could not determine if the inferred gill inflammation was progressing or resolving, our study clearly suggests that P. parvum blooms may contribute to the serious gill disorders in fish. By providing insights into the gill transcriptomic responses to toxin-producing P. parvum in teleost fish, our research opens new avenues for investigating how to monitor and mitigate toxicity of HABs before they become lethal.

Growth of the harmful alga, Prymnesium parvum (Prymnesiophyceae), after gradual and abrupt increases in salinity.

Prymnesium parvum is a euryhaline, toxin-producing microalga. Although its abundance in inland waters and growth potential in the laboratory is reduced at high salinity (>20), the ability of inland strains to adjust their growth after long-term residence in high salinity is uncertain. An inland strain of P. parvum maintained at salinity of 5 in modified artificial seawater medium (ASM-5) was subjected to the following treatments over five sequential batch culture rounds: ASM-5 (control); modified ASM at salinity of 30, raised with NaCl; modified ASM at salinity incrementally increased to 30 with NaCl; and Instant Ocean® at salinity of 30 (IO-30). Exponential growth rate (r) was reduced when salinity was increased from 5 to 30 in ASM but returned to control values during the second round. When salinity was incrementally increased, a reduction in r still occurred when salinity reached 25-30. Maximum density was reduced at salinity of 30 in ASM upon abrupt transfer or incremental increase, and compensation did not occur. Growth performance in IO-30 was comparable to control values. In conclusion, (i) long-term compensation for acute inhibitory effects of high salinity occurred for r but not maximum density, (ii) incremental increases in salinity did not prevent growth inhibition, suggesting the existence of a salinity threshold of 25-30 for onset of salinity stress, and (iii) the presence of a seawater-like salt mixture prevented growth inhibition by high salinity. These findings provide new insights on P. parvum's long-term ability to adjust its growth in environments of different salinity and ionic composition.

Low-dose stimulation of growth of the harmful alga, Prymnesium parvum, by glyphosate and glyphosate-based herbicides.

Glyphosate-based herbicides (GBH) are widely used around the globe. While generally toxic to phototrophs, organic phosphorus in glyphosate can become available to glyphosate-resistant phytoplankton and contribute to algal bloom development. Few studies have examined the effects of GBH on growth of eukaryotic microalgae and information for the toxic bloom-forming haptophyte, Prymnesium parvum, is limited. Using a batch-culture system, this study examined the effects on P. parvum growth of a single application of Roundup Weed and Grass Killer Super Concentrate Plus® (Roundup SC), Roundup Weed and Grass Killer Ready-to-Use III® (Roundup RtU), and technical-grade glyphosate at low concentrations [0-1000 µg glyphosate acid equivalent (ae) l-1]. Roundup formulations differ in the percent of glyphosate as active ingredient (Roundup SC, ∼50%; Roundup RtU, 2%), allowing indirect evaluation of the influence of inactive ingredients. Roundup SC enhanced exponential growth rate at 10-1000 µg glyphosate ae l-1, and a positive monotonic association was noted between Roundup SC concentration and early (pre-exponential growth) but not maximum cell density. Glyphosate and both Roundup formulations enhanced growth rate at 100 µg glyphosate l-1, but only Roundup SC and glyphosate significantly stimulated early and maximum density. This observation suggests the higher concentration of inactive ingredients and other compounds in Roundup RtU partially counteracts glyphosate-dependent growth stimulation. When phosphate concentration was varied while maintaining other conditions constant, addition of Roundup SC and glyphosate at 100 µg l-1 influenced growth more strongly than equivalent changes in phosphate-associated phosphorus. It appears, therefore, that low doses of glyphosate stimulate growth by mechanisms unrelated to the associated small increases in total phosphorus. In conclusion, glyphosate and GBH stimulate P. parvum growth at low, environmentally relevant concentrations. This finding raises concerns about the potential contribution to P. parvum blooms by glyphosate-contaminated runoff or by direct application of GBH to aquatic environments.