Freshwater plastisphere: a review on biodiversity, risks, and biodegradation potential with implications for the aquatic ecosystem health.

The plastisphere, a unique microbial biofilm community colonizing plastic debris and microplastics (MPs) in aquatic environments, has attracted increasing attention owing to its ecological and public health implications. This review consolidates current state of knowledge on freshwater plastisphere, focussing on its biodiversity, community assembly, and interactions with environmental factors. Current biomolecular approaches revealed a variety of prokaryotic and eukaryotic taxa associated with plastic surfaces. Despite their ecological importance, the presence of potentially pathogenic bacteria and mobile genetic elements (i.e., antibiotic resistance genes) raises concerns for ecosystem and human health. However, the extent of these risks and their implications remain unclear. Advanced sequencing technologies are promising for elucidating the functions of plastisphere, particularly in plastic biodegradation processes. Overall, this review emphasizes the need for comprehensive studies to understand plastisphere dynamics in freshwater and to support effective management strategies to mitigate the impact of plastic pollution on freshwater resources.

Microparticles from dental calculus disclose paleoenvironmental and palaeoecological records.

Plants have always represented a key element in landscape delineation. Indeed, plant diversity, whose distribution is influenced by geographic/climatic variability, has affected both environmental and human ecology. The present contribution represents a multi-proxy study focused on the detection of starch, pollen and non-pollen palynomorphs in ancient dental calculus collected from pre-historical individuals buried at La Sassa and Pila archaeological sites (Central Italy). The collected record suggested the potential use of plant taxa by the people living in Central Italy during the Copper-Middle Bronze Age and expanded the body of evidence reported by previous palynological and palaeoecological studies. The application of a microscopic approach provided information about domesticated crops and/or gathered wild plants and inferred considerations on ancient environments, water sources, and past health and diseases. Moreover, the research supplied data to define the natural resources (e.g., C4-plant intake) and the social use of the space during that period. Another important aspect was the finding of plant clues referable to woody habitats, characterised by broad-leaved deciduous taxa and generally indicative of a warm-temperate climate and grassy vegetation. Other unusual records (e.g., diatoms, brachysclereids) participated in defining the prehistoric ecological framework. Thus, this work provides an overview on the potential of the human dental calculus analysis to delineate some features of the ancient plant ecology and biodiversity.

Long-term oscillations in the normalized biomass-size spectrum reveal the impact of oligotrophication on zooplankton trophic structure in the North Atlantic Subtropical Gyre.

Ocean warming of the North Atlantic Subtropical Gyre (NASG) induced oligotrophication and a decrease in integrated net primary production during the 2010s, potentially affecting higher trophic levels. We analyzed long-term records (1994-2019) of daytime and nighttime zooplankton biomass in five size classes from the NASG. Daytime biomass decreased in the three largest size classes during the 2010s, while decrease in nighttime biomass was less evident due to the relative stability in diel vertical migrator biomass. We used the normalized biomass size spectrum (NBSS) to estimate the relative transfer efficiency between trophic levels. The steepness of the NBSS slope at the end of the time series increased by 14% (daytime) and 24% (nighttime) from the maximum observed annual average values (2011 and 2009, respectively). This suggests oligotrophication during the 2010s led to a significant reduction in the transfer of biomass across trophic levels, with negative impacts on the NASG planktonic food web.

Production of lipids and proteins from marine diatoms under changing pH and silica.

Diatom algae are increasingly explored as an alternative sustainable source for functional biomolecules likes fucoxanthin, and eicosapentaenoic acid. But biomolecule quantity and quantity are influenced by growth conditions. So, effect of differential silica concentration (0-120 mg L-1) and medium pH (5.5-9.5) on growth and cellular biochemical composition of commercially important marine diatom species were studied. Growth rate of Thalassiosira sp., Skeletonema sp., and Chaetoceros sp., was higher with 30 mg L-1 Si at a pH of 7.5-8.5. Highest carbohydrate (153.71 mg g-1) and protein (17.34 mg g-1) content was found in Skeletonema sp. Silica concentration positively influenced chlorophyll and carotenoid content in a dose dependent manner. A medium pH of 8.5 and Si concentration between 60 and 120 mg L-1 was ideal for lipid production. The optimum concentration of Si and pH for maximum biomolecule production have been reported with further scope of utilizing these conditions in commercial scale systems.

Keeping Track of Phaeodactylum tricornutum (Bacillariophyta) Culture Contamination by Potentiometric E-Tongue.

The large-scale cultivation of microalgae provides a wide spectrum of marketable bioproducts, profitably used in many fields, from the preparation of functional health products and feed supplement in aquaculture and animal husbandry to biofuels and green chemistry agents. The commercially successful algal biomass production requires effective strategies to maintain the process at desired productivity and stability levels. Hence, the development of effective early warning methods to timely indicate remedial actions and to undertake countermeasures is extremely important to avoid culture collapse and consequent economic losses. With the aim to develop an early warning method of algal contamination, the potentiometric E-tongue was applied to record the variations in the culture environments, over the whole growth process, of two unialgal cultures, Phaeodactylum tricornutum and a microalgal contaminant, along with those of their mixed culture. The E-tongue system ability to distinguish the cultures and to predict their growth stage, through the application of multivariate data analysis, was shown. A PLS regression method applied to the E-tongue output data allowed a good prediction of culture growth time, expressed as growth days, with R2 values in a range from 0.913 to 0.960 and RMSEP of 1.97-2.38 days. Moreover, the SIMCA and PLS-DA techniques were useful for cultures contamination monitoring. The constructed PLS-DA model properly discriminated 67% of cultures through the analysis of their growth media, i.e., environments, thus proving the potential of the E-tongue system for a real time monitoring of contamination in microalgal intensive cultivation.

Wastewater Biofilm Photosynthesis in Photobioreactors.

Photosynthetic performance of algal-bacterial biofilms from an Italian wastewater treatment plant was studied in a flow-lane photobioreactor at different irradiances, temperatures, and flow regime to evaluate the effects of these environmental parameters on biofilms' functioning, in view of application of these communities in wastewater biological treatment. Pulse amplitude modulated fluorescence was used to estimate the effective quantum yield of PSII (ΔF/Fm') of the light-acclimated biofilms and to perform rapid light curves (RLCs) for the determination of the photosynthetic parameters (rel.ETRmax, α, Ik). Chl a, ash free dry weight (AFDW), and dry weight (DW) were measured to assess phototrophic and whole biofilm biomass development over time. From the analysis of photosynthetic parameter variation with light intensity, temperature and flow rate, it was possible to identify the set of experimental values favoring biofilm photosynthetic activity. Biomass increased over time, especially at the highest irradiances, where substrata were fastly colonized and mature biofilms developed at all temperatures and flow conditions tested.

Scaling-up of wastewater bioremediation by Tetradesmus obliquus, sequential bio-treatments of nutrients and metals.

Microalgae may be exploited in water or wastewater treatment facilities to reduce excess concentrations of nutrients and metals to comply with regulatory limits. In this study, we characterized the growth and phosphorus (P) removal capacity of an isolated strain of Tetradesmus obliquus VRUC280. Investigations were carried out from laboratory scale (50 mL) up to a 100 L outdoor photobioreactor (PBR). After 10 days, batch cultures removed up to 74% of the media P, while in the PBR, 95% removal was achieved within five days. The harvested biomass was then inactivated (freeze-dried) and used for metal adsorption tests, employing solutions containing 6.0 mg Cu L-1 or 4.8 mg Ni L-1. Metal removal rates were evaluated after 15, 30, 60 and 120 min by the analysis of liquid and biomass metal contents. For the latter, a specific biomass digestion method was developed. Cu removal ranged between 50% and 65%, while for Ni, removal varied between 30% and 50%. 300-400 mg Cu Kg DW-1 and 130-250 mg Ni Kg DW-1 were rapidly adsorbed on the cell surface of T. obliquus (ca. 15-30 min incubations). This study demonstrates the potential of microalgae, in this case T. obliquus, to remove sequentially P and metals from aqueous media.

UV-shielding and wavelength conversion by centric diatom nanopatterned frustules.

Diatoms can represent the major component of phytoplankton and contribute massively to global primary production in the oceans. Over tens of millions of years they developed an intricate porous silica shell, the frustule, which ensures mechanical protection, sorting of nutrients from harmful agents, and optimization of light harvesting. Several groups of microalgae evolved different strategies of protection towards ultraviolet radiation (UVR), which is harmful for all living organisms mainly through the formation of dimeric photoproducts between adjacent pyrimidines in DNA. Even in presence of low concentrations of UV-absorbing compounds, several diatoms exhibit significant UVR tolerance. We here investigated the mechanisms involved in UVR screening by diatom silica investments focusing on single frustules of a planktonic centric diatom, Coscinodiscus wailesii, analyzing absorption by the silica matrix, diffraction by frustule ultrastructure and also UV conversion into photosynthetically active radiation exerted by nanostructured silica photoluminescence. We identified the defects and organic residuals incorporated in frustule silica matrix which mainly contribute to absorption; simulated and measured the spatial distribution of UVR transmitted by a single valve, finding that it is confined far away from the diatom valve itself; furthermore, we showed how UV-to-blue radiation conversion (which is particularly significant for photosynthetic productivity) is more efficient than other emission transitions in the visible spectral range.

Trichormus variabilis (Cyanobacteria) Biomass: From the Nutraceutical Products to Novel EPS-Cell/Protein Carrier Systems.

A native strain of the heterocytous cyanobacterium Trichormus variabilis VRUC 168 was mass cultivated in a low-cost photobioreactor for a combined production of Polyunsaturated Fatty Acids (PUFA) and Exopolymeric Substances (EPS) from the same cyanobacterial biomass. A sequential extraction protocol was optimized leading to high yields of Released EPS (REPS) and PUFA, useful for nutraceutical products and biomaterials. REPS were extracted and characterized by chemical staining, Reversed Phase-High-Performance Liquid Chromatography (RP-HPLC), Fourier Transform Infrared Spectroscopy (FT-IR) and other spectroscopic techniques. Due to their gelation property, REPS were used to produce a photo-polymerizable hybrid hydrogel (REPS-Hy) with addition of polyethylene glycol diacrylated (PEGDa). REPS-Hy was stable over time and resistant to dehydration and spontaneous hydrolysis. The rheological and functional properties of REPS-Hy were studied. The enzyme carrier ability of REPS-Hy was assessed using the detoxification enzyme thiosulfate:cyanide sulfur transferase (TST), suggesting the possibility to use REPS-Hy as an enzymatic hydrogel system. Finally, REPS-Hy was used as a scaffold for culturing human mesenchymal stem cells (hMSCs). The cell seeding onto the REPS-Hy and the cell embedding into 3D-REPS-Hy demonstrated a scaffolding property of REPS-Hy with non-cytotoxic effect, suggesting potential applications of cyanobacteria REPS for producing enzyme- and cell-carrier systems.

Non-enzymatic portable optical sensors for microcystin-LR.

We present here the development of an all-solid-state optical sensor based on phenyl-substituted diaza-18-crown-6 hydroxyquinoline (DCHQ-Ph) for the indirect selective detection of microcystin-LR (MC-LR), reaching a very low detection limit of 0.05 µg L-1, well below the World Health Organisation (WHO) guideline value (1 µg L-1) in potable water. We demonstrate the potential applicability of the developed method in fast and low-cost water toxicity estimation.

Culturing Toxic Benthic Blooms: The Fate of Natural Biofilms in a Microcosm System.

A microcosm designed for culturing aquatic phototrophic biofilms on artificial substrata was used to perform experiments with microphytobenthos sampled during summer toxic outbreaks of Ostreopsis cf. ovata along the Middle Tyrrhenian coast. This dynamic approach aimed at exploring the unique and complex nature of O. cf. ovata bloom development in the benthic system. Epibenthic assemblages were used as inocula for co-cultures of bloom organisms on polycarbonate slides at controlled environmental conditions. Biofilm surface adhesion, growth, and spatial structure were evaluated along with shifts in composition and matrix production in a low disturbance regime, simulating source habitat. Initial adhesion and substratum colonisation appeared as stochastic processes, then community structure and physiognomy markedly changed with time. Dominance of filamentous cyanobacteria and diatoms, and dense clusters of Amphidinium cf. carterae at the mature biofilm phases, were recorded by light and confocal microscopy, whilst O. cf. ovata growth was visibly limited in the late culture phases. Life-form strategies, competitiveness for resources, and possibly allelopathic interactions shaped biofilm structure during culture growth. HPLC (High Performance Liquid Chromatography) analysis of exopolysaccharidic matrix revealed variations in sugar total amounts and composition. No toxic compounds were detected in the final communities tested by LC-MS (Liquid Chromatography- Mass Spectrometry) and MALDI-TOF MS (Matrix Assisted Laser Desorption Ionization Time OF Flight Mass Spectroscopy) techniques.

Phosphatase activities of a microepiphytic community during a bloom of Ostreopsis cf. ovata in the northern Adriatic Sea.

It is becoming more apparent that increased organic nutrient loads deriving from anthropogenic activities and natural processes frequently cause the eutrophication of coastal waters. Concurrently, an increasing number of phototrophs have been shown to make use of organic nutrients, mainly through indirect studies of surface enzyme activities or through direct studies of growth in media containing organic-only nutrients. The potential utilization of dissolved organic phosphorus (DOP) by microepiphytic-mats associated with frequently problematic, toxic dinoflagellate Ostreopsis cf. ovata, was investigated throughout a full cycle of a bloom that occurs annually (over the last few decades) during summer along several stretches of the Mediterranean coast. Measurements of phosphomonoesterase (PMEase) and phosphodiesterase (PDEase) activities of the epiphytic mats (including cells and exopolymeric substances) and a range of chemico-physical parameters were made from late summer to early autumn at a eutrophic site. Analyses of ambient nutrient fractions revealed very high aquatic N:P values (TN:TP = 178 ± 50), very low filterable reactive P (FRP) concentrations (13 out 19 under the limits of detection) and DOP concentrations that were on average 85% of the total dissolved P. We recorded a rapid increase in PMEase and PDEase activities in the epiphytic microalgal community that coincided with the onset of a proliferation of the Ostreopsis population. Chromogenic staining of samples showed that activity was closely associated with the Ostreopsis cells, located both extracellularly (cell surface and within the EPS) and intracellularly (ventral cytoplasm). The increase in both phosphatases indicates that Ostreopsis can utilise a wide range of DOP types. The intense activity in the EPS was suggested to aid in the efficient entrapment and processing of high concentration nutrient pulses, for extracellular processing of larger organic materials and to prevent loss of substrates and products to the surrounding water. Based on the present findings, Ostreopsis seems to have adaptations that allow it to thrive in P-limited environments where organic P is the main source of P.

The Diatom Staurosirella pinnata for Photoactive Material Production.

A native isolate of the colonial benthic diatom Staurosirella pinnata was cultivated for biosilica production. The silicified cell walls (frustules) were used as a source of homogeneous and structurally predictable porous biosilica for dye trapping and random laser applications. This was coupled with the extraction of lipids from biomass showing potential to fabricate photoactive composite materials sustainably. The strain was selected for its ease of growth in culture and harvesting. Biosilica and lipids were obtained at the end of growth in indoor photobioreactors. Frustules were structurally characterized microscopically and their chemistry analyzed with Fourier Transform Infrared Spectroscopy. Frustule capacity of binding laser dyes was evaluated on a set of frustules/Rhodamine B (Rho B) solutions and with respect to silicon dioxide and diatomite by Fluorescence Spectroscopy demonstrating a high affinity for the organic dye. The effect of dye trapping property in conveying Rho B emission to frustules, with enhancement of scattering events, was analyzed on Rho B doped polyacrylamide gels filled or not with frustules. Amplified spontaneous emission was recorded at increasing pump power indicating the onset of a random laser effect in frustule filled gels at lower power threshold compared to unfilled matrices.

Dataset exploited for the development and validation of automated cyanobacteria quantification algorithm, ACQUA.

The estimation and quantification of potentially toxic cyanobacteria in lakes and reservoirs are often used as a proxy of risk for water intended for human consumption and recreational activities. Here, we present data sets collected from three volcanic Italian lakes (Albano, Vico, Nemi) that present filamentous cyanobacteria strains at different environments. Presented data sets were used to estimate abundance and morphometric characteristics of potentially toxic cyanobacteria comparing manual Vs. automated estimation performed by ACQUA ("ACQUA: Automated Cyanobacterial Quantification Algorithm for toxic filamentous genera using spline curves, pattern recognition and machine learning" (Gandola et al., 2016) [1]). This strategy was used to assess the algorithm performance and to set up the denoising algorithm. Abundance and total length estimations were used for software development, to this aim we evaluated the efficiency of statistical tools and mathematical algorithms, here described. The image convolution with the Sobel filter has been chosen to denoise input images from background signals, then spline curves and least square method were used to parameterize detected filaments and to recombine crossing and interrupted sections aimed at performing precise abundances estimations and morphometric measurements.

ACQUA: Automated Cyanobacterial Quantification Algorithm for toxic filamentous genera using spline curves, pattern recognition and machine learning.

Toxigenic cyanobacteria are one of the main health risks associated with water resources worldwide, as their toxins can affect humans and fauna exposed via drinking water, aquaculture and recreation. Microscopy monitoring of cyanobacteria in water bodies and massive growth systems is a routine operation for cell abundance and growth estimation. Here we present ACQUA (Automated Cyanobacterial Quantification Algorithm), a new fully automated image analysis method designed for filamentous genera in Bright field microscopy. A pre-processing algorithm has been developed to highlight filaments of interest from background signals due to other phytoplankton and dust. A spline-fitting algorithm has been designed to recombine interrupted and crossing filaments in order to perform accurate morphometric analysis and to extract the surface pattern information of highlighted objects. In addition, 17 specific pattern indicators have been developed and used as input data for a machine-learning algorithm dedicated to the recognition between five widespread toxic or potentially toxic filamentous genera in freshwater: Aphanizomenon, Cylindrospermopsis, Dolichospermum, Limnothrix and Planktothrix. The method was validated using freshwater samples from three Italian volcanic lakes comparing automated vs. manual results. ACQUA proved to be a fast and accurate tool to rapidly assess freshwater quality and to characterize cyanobacterial assemblages in aquatic environments.

Phosphorus removal coupled to bioenergy production by three cyanobacterial isolates in a biofilm dynamic growth system.

In the present study a closed incubator, designed for biofilm growth on artificial substrata, was used to grow three isolates of biofilm-forming heterocytous cyanobacteria using an artificial wastewater secondary effluent as the culture medium. We evaluated biofilm efficiency in removing phosphorus, by simulating biofilm-based tertiary wastewater treatment and coupled this process with biodiesel production from the developed biomass. The three strains were able to grow in the synthetic medium and remove phosphorus in percentages, between 6 and 43%, which varied between strains and also among each strain according to the biofilm growth phase. Calothrix sp. biofilm turned out to be a good candidate for tertiary treatment, showing phosphorus reducing capacity (during the exponential biofilm growth) at the regulatory level for the treated effluent water being discharged into natural water systems. Besides phosphorus removal, the three cyanobacterial biofilms produced high quality lipids, whose profile showed promising chemical stability and combustion behavior. Further integration of the proposed processes could include the integration of oil extracted from these cyanobacterial biofilms with microalgal oil known for high monounsaturated fatty acids content, in order to enhance biodiesel cold flow characteristics.

Optical properties of diatom nanostructured biosilica in Arachnoidiscus sp: micro-optics from mother nature.

Some natural structures show three-dimensional morphologies on the micro- and nano-scale, characterized by levels of symmetry and complexity well far beyond those fabricated by best technologies available. This is the case of diatoms, unicellular microalgae, whose protoplasm is enclosed in a nanoporous microshell, made of hydrogenated amorphous silica, called frustule. We have studied the optical properties of Arachnoidiscus sp. single valves both in visible and ultraviolet range. We found photonic effects due to diffraction by ordered pattern of pores and slits, accordingly to an elaborated theoretical model. For the first time, we experimentally revealed spatial separation of focused light in different spots, which could be the basis of a micro-bio-spectrometer. Characterization of such intricate structures can be of great inspiration for photonic devices of next generation.

Shedding light on diatom photonics by means of digital holography.

Diatoms are among the dominant phytoplankters in the world's oceans, and their external silica investments, resembling artificial photonic crystals, are expected to play an active role in light manipulation. Digital holography allowed studying the interaction with light of Coscinodiscus wailesii cell wall reconstructing the light confinement inside the cell cytoplasm, condition that is hardly accessible via standard microscopy. The full characterization of the propagated beam, in terms of quantitative phase and intensity, removed a long-standing ambiguity about the origin of the light confinement. The data were discussed in the light of living cell behavior in response to their environment.