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- From piggery wastewater to wheat using microalgae towards zero wastePublication . Ferreira, Alice; Figueiredo, Daniel; Ferreira, Francisca; Marujo, Ana; Bastos, Carolina; Martin-Atanes, Guillermo; Ribeiro, Belina; Štěrbová, Karolína; Marques-dos-Santos, Cláudia; Acién, F. Gabriel; Gouveia, LuisaMicroalgae production is still expensive, driving the need to lower costs while strengthening the industry's environmental sustainability. Microalgae are recognized tools for efficient wastewater treatment, offering the recycling of nutrients and water for agriculture, and producing biomass rich in growth-promoting compounds to improve plant productivity and resistance to adverse conditions. The use of wastewater can reduce cultivation costs as it is a source of nutrients and water. Alternative low-cost methods can significantly decrease harvesting costs, which represents one of the most expensive steps of the whole process.The goal of this work was to evaluate the potential of wastewater-grown microalga biomass for agriculture purposes. To reduce production costs, the microalga Tetradesmus obliquus was produced in pre-treated photo-Fenton (PF) piggery wastewater in combination with the use of different harvesting techniques - electro-coagulation, flocculation, and centrifugation, and different combinations. From the wastewater treatment pro-cess, two fractions (biomass and supernatant) were evaluated for germination and growth of wheat (Triticum aestivum L.) plants and compared to non-harvested microalga culture (MC), distilled water, and Hoagland (synthetic) solution. The concentrated resulting from PF was also tested as a biofertilizer.The results confirm that both biomass and supernatants are useful for agricultural applications. The obtained biomass elicited a 20-105 % increase in germination index compared to the control, while supernatants were inhibiting. The opposite trend was observed at later stages of wheat growth, where the nutrient-enriched su-pernatants and the PF concentrate (PF-CC) increased the number of tillers (3-5) and leaves (30-42) after 83 days. Wheat plants treated with MC and PF-CC produced similar number of ears (3.4 & PLUSMN; 0.5 and 6.0 & PLUSMN; 4.1 ears per plant, respectively) than the synthetic control (5.7 & PLUSMN; 1.4) after 182 days. All fractions obtained from the process can be used in a zero-waste process.
- Impact of high-pressure homogenization on the cell integrity of tetradesmus obliquus and seed germinationPublication . Ferreira, Alice; Figueiredo, Daniel; Ferreira, Francisca; Ribeiro, Belina; Reis, Alberto; da Silva, Teresa Lopes; Gouveia, LuisaMicroalgae have almost unlimited applications due to their versatility and robustness to grow in different environmental conditions, their biodiversity and variety of valuable bioactive compounds. Wastewater can be used as a low-cost and readily available medium for microalgae, while the latter removes the pollutants to produce clean water. Nevertheless, since the most valuable metabolites are mainly located inside the microalga cell, their release implies rupturing the cell wall. In this study, Tetradesmus obliquus grown in 5% piggery effluent was disrupted using high-pressure homogenization (HPH). Effects of HPH pressure (100, 300, and 600 bar) and cycles (1, 2 and 3) were tested on the membrane integrity and evaluated using flow cytometry and microscopy. In addition, wheat seed germination trials were carried out using the biomass at different conditions. Increased HPH pressure or number of cycles led to more cell disruption (75% at 600 bar and 3 cycles). However, the highest increase in wheat germination and growth (40-45%) was observed at the lowest pressure (100 bar), where only 46% of the microalga cells were permeabilised, but not disrupted. Non-treated T. obliquus cultures also revealed an enhancing effect on root and shoot length (up to 40%). The filtrate of the initial culture also promoted shoot development compared to water (21%), reinforcing the full use of all the process fractions. Thus, piggery wastewater can be used to produce microalgae biomass, and mild HPH conditions can promote cell permeabilization to release sufficient amounts of bioactive compounds with the ability to enhance plant germination and growth, converting an economic and environmental concern into environmentally sustainable applications.
- Microalgae-associated stenotrophomonas maltophilia enhances lutein production and biostimulant activity in Monoraphidium sp.Publication . Manoel, João Artur Câmara; Šterbová, Karolína; Saini, Mohit Kumar; Bárcenas-Pérez, Daniela; Cheel, José; Grivalský, Tomáš; Lakatos, Gergely Ernő; Lukeš, Martin; Urajová, Petra; Ferreira, Alice; Rodrigues Figueiredo, Daniel; Gouveia, Luisa; Masojídek, Jiří; Saurav, KumarPhotosynthetic organisms like microalgae can collect solar energy and transform it into biochemical compounds as other forms of energy that can be utilized in metabolic processes. In nature, microalgae coexist with bacterial communities and may maintain a symbiotic relationship. In the current study, a heterotrophic bacterium, Stenotrophomonas maltophilia was isolated from the phycosphere of a cold-adapted green microalga Monoraphidium sp. (further abbreviated as Monoraphidium). By using advanced liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS), we were able to detect homoserine lactones (HSLs): 3OHC12-HSL, 3OHC10-HSL, 3OHC14-HSL, C10-HSL, C8-HSL, and OC14-HSL, produced by S. maltophilia. Further, the role of this bacterium in establishing intricate relationships and its implication on biotechnological potential was evaluated. Significant improvements were found in the lutein production of the Monoraphidium culture with bacterial supplements, achieving about 19.3 ± 0.88 mg g− 1 DW of this carotenoid compared to 13.7 ± 1.87 mg g− 1 DW in the control, which represents an increase of about 40 %. Furthermore, the biostimulant potential of Monoraphidium was evaluated using the germination tests with tomato and barley seeds. A higher germination index was observed with improvements of 55 % in tomato and 110 % in barley, respectively, as compared to the control culture, which was related to the microalgae’s growth stage. The role of the bacterium was evaluated in how the intricate relationships with the microalgal culture can affect its biotechnological potential (e.g., biostimulant activity and lutein production). The current work expands our knowledge towards designing an efficient polyculture based on complementary traits and metabolic potential to maximize the yield and bioactivity in algal biotechnology.
- Optimizing chlorella vulgaris production and exploring its impact on germination through microalga-N2-fixing bacteria consortiaPublication . Sanchez-Zurano, Ana; Vilaró-Cos, Silvia; Rodrigues Figueiredo, Daniel; Melkonyan, Lusine; Ferreira, Alice; Acién, Francisco Gabriel; Lafarga, Tomas; Gouveia, LuisaMicroalgal biomass is increasingly valued in industrial and agricultural sectors due to its bioactive compounds. However, large-scale production remains costly, mainly due to nitrogen fertilizer expenses. A promising sustainable alternative is co-cultivation with N2-fixing bacteria, capable of supplying biologically available nitrogen. In this study, Chlorella vulgaris was grown in synthetic medium with and without nitrogen, as well as in co-culture with three different N2-fixing bacteria in nitrogen-free medium. Microalgal growth was assessed by dry weight, Fv/Fm ratio, and flow cytometry, which also allowed evaluation of population dynamics and cell viability. Biomass composition (proteins, carbohydrates, lipids, chlorophyll, and carotenoids) was analyzed under all conditions. Co-cultures in nitrogen-free medium showed comparable biomass productivity to nitrogensupplemented controls, although Fv/Fm values indicated physiological stress in some cases. Moreover, the agricultural potential of the resulting biomass and supernatants was evaluated through germination bioassays using lettuce seeds. All cultures tested at 0.2 g⋅L− 1 significantly improved the germination index. Also, applying the culture supernatant (biomass removed) also yielded positive effects, with GI increases exceeding 40 %. These results suggest that co-cultivation with N2-fixing bacteria can support efficient microalgal production while generating biomass and supernatants with biostimulant potential, contributing to sustainable agriculture and circular bioeconomy strategies.