Phytool, a ShinyApp to homogenise taxonomy of freshwater microalgae from DNA barcodes and microscopic observations

Methods for biomonitoring of freshwater phytoplankton are evolving rapidly with eDNA-based methods, offering great complementarity with microscopy. Metabarcoding approaches have been more commonly used over the last years, with a continuous increase in the amount of data generated. Depending on the researchers and the way they assigned barcodes to species (bioinformatic pipelines and molecular reference databases), the taxonomic assignment obtained for HTS DNA reads might vary. This is also true for traditional taxonomic studies by microscopy with regular adjustments of the classification and taxonomy. For those reasons (leading to non-homogeneous taxonomies), gap-analyses and comparisons between studies become even more challenging and the curation processes to find potential consensus names are time-consuming. Here, we present a web-based application (Phytool), developed with ShinyApp (Rstudio), that aims to make the harmonisation of taxonomy easier and in a more efficient way, using a complete and up-to-date taxonomy reference database for freshwater microalgae. Phytool allows users to homogenise and update freshwater phytoplankton taxonomical names from sequence files and data tables directly uploaded in the application. It also gathers barcodes from curated references in a user-friendly way in which it is possible to search for specific organisms. All the data provided are downloadable with the possibility to apply filters in order to select only the required taxa and fields (e.g. specific taxonomic ranks). The main goal is to make accessible to a broad range of users the connection between microscopy and molecular biology and taxonomy through different ready-to-use functions. This study estimates that only 25% of species of freshwater phytoplankton in Phytobs are associated with a barcode. We plead for an increased effort to enrich reference databases by coupling taxonomy and molecular methods. Phytool should make this crucial work more efficient. The application is available at https://caninuzzo.shinyapps.io/phytool_v1/

Some strains from microalgae collection IBASU-A (Ukraine) as an object of biotechnology

An information on the collection of strains of biotechnological application as an integral part of Microalgal Culture Collection of the M.G. Kholodny Institute of Botany of NAS of Ukraine (IBASU-A) is given. The base of its funds contains some green algal strains belonging to the families of Dunaliellaceae, Chlorellaceae, Scenedesmaceae and Selenastraceae. They have been isolated from different regions of Ukraine in order to find cultures of phototrophic microorganisms – promising for biotechnology, in particular, obtaining biologically active additives for the needs of the food industry, medicine, agriculture, raw materials for the production of biofuels, as well as bioindication, biomonitoring, bioremediation of aquatic objects of the environment, etc. Overall, this special collection includs 90 strains of halophile and freshwater microalgae of 30 species, 15 genera, 7 families, 4 orders, 2 classes. All of them are considered as important objects for industrial cultivation, solution of environmental problems, and the basis for further biotechnological research.

Phycoremediation of fish farm wastewater by Chlorella sorokiniana and autochthonous microalgae

With the disorderly increase in global environmental problems, the cultivation of aquatic organisms is a promising path for sustainable food production. The quality of water, both at the entrance and exit of the production of aquatic animals, needs to be maintained following the parameters specified by local legislation. This study aimed to investigate the removal of contaminants from fish farming wastewater associated with the production of freshwater microalgae biomass. Six completely randomized treatments were used in triplicate: with the addition of microalgae C. sorokiniana in fish farm wastewater (W+Cs), the addition of C. sorokiniana in wastewater enriched with NPK fertilizing (W+F+Cs) or sugarcane vinasse (W+V+Cs), only wastewater (W), wastewater supplemented with fertilizer (W+F) or vinasse (W+V). The wastewater was used in natura to allow the development of autochthonous microalgae. The microalgae C. sorokiniana grew rapidly in effluents enriched with NPK and vinasse. After 28 days of bioassay, the concentrations of several contaminants in the water were reduced: zinc (20 to 88%), lead (5 to 83%), aluminum (56 to 75%), manganese (56 to 72%), cadmium (9 to 52%), calcium (16 to 24%) and magnesium (12 to 33%). Our results indicated that the production of microalgae biomass can be integrated with the treatment of fish farming effluents to reduce the environmental burden and increase the economic bonus for adopting a sustainable production method. However, our results also indicated the importance of introducing a microalgae strain with high productive performance and supplementing the wastewater to obtain rapid biomass.

Development of wound dressing film using methanolic extracts of freshwater microalgae and determining its wound healing ability

Wound healing is a complex process in which bacterial infection is a major cause for delayed wound healing. Occurrence of drug resistance among bacterial pathogens led to discovery of new antimicrobial agents from new sources. The present study aimed to identify microalgal metabolites with antibacterial activity and to develop a wound dressing film with their potential healing activity. Microalgal samples were collected from three different freshwater habitats, isolated, made them pure cultures and physico-chemical properties of water samples from the respective sampling sites were analzed. Among the three microalgal isolates, Chlorella sp. NRMC-F-0350 showed antibacterial activity against clinical isolates of E. coli, Klebsiella pneumoniae and Staphylococcus aureus. Additionally, functional groups present in Chlorella sp. NRMC-F-0350 were identified using FTIR and GC-MS analysis. Several antibacterial compounds viz. benzoxazole, 1,2 Benzene dicarboxylic acid, sistosterol, 9-Octadecanoic acid, eicosane and hexadecane were identified. Wound dressing films were developed and showed evident antibacterial as well as significant wound healing activity (84.5%). Therefore, the developed films can be used as a potential wound dressing material.

Infection with intracellular parasite Amoeboaphelidium protococcarum induces shifts in associated bacterial communities in microalgae cultures

In recent years microalgae products have developed increasing market demand, but sustainable industrial production is still challenged by biological stability of large-scale production plants. Yet the relationships between algal hosts, associated microbiomes, and contaminants in photobioreactors remains widely understudied. The aim of this study was to investigate the temporal development of microbiomes of four freshwater microalgae species Scenedesmus vacuolatus, Desmodesmus quadricauda, Chlorella sorokiniana, and Botryococcus braunii, in presence and absence of the zoosporic parasite Amoeboaphelidium protococcarum. To compare the effects of sterile and nonsterile culture conditions, infection experiments were performed in sterile laboratory (sterile) and simulated industrial conditions (open). Algal growth (dry weight, optical density, and nutrient consumption) was observed for 21 days, and samples of the associated microbiome were collected for bacterial 16S rRNA gene Illumina MiSeq sequencing. Infection patterns of A. protococcarum were algae species-specific, irrespectively of culture conditions. Bacterial community analysis demonstrated distinct and stable bacterial communities for each algae species, which were mostly dominated by α- and γ-Proteobacteria. Upon aphelid parasitosis, bacterial diversity increased, and community compositions diverged algae-specific over time. Moreover, bacterial functional traits shifted to detoxification, degradation, and cellulolysis once algae were infected. This study provides a first insight into the close connection between algae, associated bacterial microbiomes and appearing contaminants in photobioreactor systems.

Freshwater alga Raphidocelis subcapitata undergoes metabolomic changes in response to electrostatic adhesion by micrometer-sized nylon 6 particles

Nylon powders are a type of microplastic (MP) used in personal care products such as cosmetics and sunscreens. To determine the effects of nylon polymers on freshwater microalgae, we investigated the effects of two types of micrometer-sized nylon polymers, i.e., powdered nylon 6 (Ny6-P) and nylon 12 (Ny12), and four other micrometer-sized MPs, i.e., low-density polyethylene, polyethylene terephthalate, polystyrene, and ultra-high-molecular-weight polyethylene, on the microalga Raphidocelis subcapitata. The results showed that Ny6-P inhibited R. subcapitata growth more than the other MPs; R. subcapitata growth was inhibited by 54.2% with 6.25 mg/L Ny6-P compared with the control. Ny6-P in the culture media adhered to R. subcapitata cells electrostatically, which may have disrupted growth and photosynthetic activity. Metabolomic analysis revealed that many metabolites related to the amino acid catabolic pathway and γ-glutamyl cycle were induced, which might trigger responses to avoid starvation and oxidative stress. Our study provides important information on the effects of Ny6-P on algae in freshwater environments.