Green tides refer to harmful algal bloom (HAB) phenomena caused by the explosive proliferation or high accumulation of some green algae species under specific environmental conditions, resulting in substantial global marine environmental problems (Rybak, 2018). Since 2007, the green tide in the Yellow Sea of China caused by Ulva prolifera has occurred annually (Xiao et al., 2020; Yuan et al., 2022) and is considered the largest green tide outbreak event in the world (Zhao et al., 2019). The green tide typically forms in the South Yellow Sea around mid-April every year and subsequently drifts northward, propelled by ocean currents and monsoons, finally reaching its peak distribution and coverage area in the sea of Shandong Province by June (Xiao et al., 2013). As the seawater temperature increases and the nutrient salt content decreases by late July, the buoyancy, activity, and photosynthetic rates of the alga plummet, indicating that the floating green algae are demising. Only a fraction of U. prolifera are collected before they settle to the bottom of the ocean and demise in Shandong Province, off the shore of the cities of Qingdao, Rizhao, Weihai, etc. The demise of U. prolifera results in the release of large amounts of nitrogen and phosphorus nutrients that are absorbed during its migration across the Yellow Sea (Wang Z. et al., 2023). This process not only has a substantial effect on the socioeconomic development of coastal regions but also has irreversible ecological repercussions on marine ecosystems (Bastos et al., 2019; Li et al., 2019; Wang et al., 2021).

In algal ecosystems, the concept of the “phycosphere” is similar to that of the plant “rhizosphere” and refers to the microenvironment immediately surrounding an algae cell that is enriched in organic molecules exuded by the cell into the surrounding water (Seymour et al., 2017). Studies have demonstrated that phycospheric microorganisms interact with algae in multiple ways during the different stages of algae growth, forming symbiotic, competitive, and harmful relationships with them (Daly et al., 2022; Zhao et al., 2023). As hosts, algae provide habitats, vitamins, inorganic salts and carbohydrates for phycospheric microorganisms (Krug et al., 2020). In contrast, phycospheric microorganisms provide amino acids, polypeptides, nucleotides, fatty acids and dimethylsulfoniopropionate for algae to promote their growth (Caruana and Malin, 2014; Variem and Kizhakkedath, 2021). Therefore, the phycospheric microenvironment provides a key medium for the interaction between algae and microorganisms (Noreen et al., 2023), and can facilitate the colonization of specific microorganisms within it (Han et al., 2021). This microenvironment also includes many algae-lysing bacteria, which can cleave algal bloom-causing species through physical contact or secrete algae-lysing substances (including proteases, alkaloids, benzoic acid, amino acids, etc.) and compete for nitrogen and phosphorus, promoting the reduction of HABs (Nair et al., 2022; Thoré et al., 2023). For example, Saprospira can directly destroy the diatom cell wall, leading to algal cell dissolution and death (Wang M. et al., 2020).

Although interactions between algae and bacteria occur in the microscale environment, they influence the entire process of HABs and can also drive and affect the cycle of marine elements in the outbreak area. The large amount of dissolved organic carbon (DOC) secreted by algae provides support for the rapid growth and reproduction of phycospheric microorganisms during the early stage of HABs (Piontek et al., 2011). The DOC released by algae is composed mainly of polysaccharides, and phycospheric microorganisms also use polysaccharides in a variety of ways. For example, Flavobacterium mainly ingests and degrades complex polysaccharides and other high-molecular-weight organic carbon, while Alteromonadaceae can extract organic carbon by degrading polysaccharides or monosaccharide types with low complexity (Qu et al., 2021). Furthermore, dissolved organic nitrogen (DON) and dissolved organic phosphorus (DOP) are remineralized by phycospheric microorganisms, providing substantial nutrients for algae (Krug et al., 2020; Zhao et al., 2023). When the HABs enter the late stage, the number of algae begin to decrease, and the change in algae source material not only affects the composition of the phycospheric microbial community but also changes the content of biogenic elements in the ocean. For example, heterotrophic bacteria in the phycosphere have a great effect on the degradation and remineralization of organic matter during the late stage of diatom blooms, thus significantly affect the marine carbon cycle (Buchan et al., 2014; Wu et al., 2023). Thus, the relationship between algae and microorganisms plays a crucial role in regulating biogeochemical element cycles in the phycospheric microenvironment.

The microbial community can exhibit obvious specificity during HABs, and the different development stages of HABs affect the structural variation and element cycles of the microbial community (Ma et al., 2024). For example, Bacteroidetes and Proteobacteria dominate the microbial communities in red tide areas, whereas Proteobacteria dominate in non-red tide areas (Delmont et al., 2014). After a dinoflagellate bloom (Gymnodinium catenatum) enters the outbreak stage, the microbial abundance involved in the reduction of the citrate cycle, carbon degradation and nitrification transport increases significantly (Zhou et al., 2020). During the extinction period of algal blooms, the majority of bacteria are Pseudomonas and Bacillus, which may have denitrification functions (Zheng et al., 2008). Because of the complex relationship between microorganisms and algae, it is widely believed that the beginning, persistence and disappearance of HABs involve interactions with microbial communities (Qu et al., 2021). Therefore, we focused on a primary site associated with the demise of green tides caused by U. prolifera in the Yellow Sea, and the composition structure, functional changes and related gene functional groups of microbial communities in seawater were analyzed via metagenomic technology coupled with environmental physicochemical factors. Our primary objectives were to explore (1) the changes in microbial community structure composition during the demise stage of U. prolifera green tides and (2) the element cycles that microorganisms participated in the phycospheric microenvironment. This research is crucial for a comprehensive understanding of the ecological effects exerted by phycospheric microorganisms during the green tide demise period.

This study analyzed the changes in the structure of the microbial community within the phycospheric microenvironment and biogeochemical effects induced in the late stage of U. prolifera green tides. Our results revealed that during the demise stage of green tides, significant changes have taken place in the structure and diversity of phycospheric microorganisms. Meanwhile, founded that this changes will further affect the element cycling process in the phycospheric microenvironment.

We found that the demise of U. prolifera can change the environmental factors and microbial community diversity in the phycospheric microenvironment. Generally, U. prolifera decomposes could release many organic compounds and consume oxygen in the late stage of green tide (Zhou et al., 2019; DeRose et al., 2021). Our study also obtained similar findings that the concentration of DO decreased, and the concentrations of DOC and nitrogenous nutrients increased significantly in the phycospheric microenvironment (Figure 1). Consistent with previous findings showing that some bacteria can use the organic matter released by algae to reproduce rapidly and gain an advantage in the competition, such as Algoriphagus and Marinobacter (Liang et al., 2021; Kim et al., 2023). Our results showed that the microbial diversity in the phycosphere decreased significantly (Figure 3). This could be the large amount of organic matter released by U. prolifera will promote the growth of some specific microbe rapidly, thus occupy a dominant position in the phycospheric microenvironment, ultimately lead to a decrease in microbial diversity.

There was a significant difference in the composition of the microbial community between the phycospheric microenvironment and surrounding seawater environment during the demise of U. prolifera. In our research, Proteobacteria and Bacteroidota at the phylum level together accounted for more than 80% of the total bacteria in the phycosphere. Flavobacteriaceae at the family level and Alteromonas, Maribacter and Vibrio at the genus level increased significantly during the demise stage of the green tide (Figures 2, 3). Proteobacteria and Bacteroidota are the two most common phyla in the phycosphere and exhibit a wide range of functional diversity as well as ecological adaptability (Coyne et al., 2022). Many isolated algae-lysing bacteria belong to Proteobacteria and Bacteroidota (Gao et al., 2020; Wang M. et al., 2020). In addition, relevant studies have shown that Flavobacteriaceae inhibits the growth of many HAB species, such as Microcystis aeruginosa and Prococentrum marinum (Rishiram et al., 2016). This is because the genome of Flavobacteriaceae species contains many genes encoding polysaccharide-degrading enzymes, so Flavobacteriaceae can use and decompose a variety of complex carbon sources (Gavriilidou et al., 2020). In recent years, a variety of algae-lysing bacteria in genera such as Streptomyces, Maribacter, Pseudoalteromonas, Alteromonas, Shewanella, and Vibrio, which are important organic degradation agents for complex biological macromolecules and algae fragments, have been discovered (Kim et al., 2023; Zheng et al., 2018). Attention worthy, we detected high abundance of Maribacter, Alteromonas and Vibrio in the phycosphere, which can promote carbon and nitrogen cycling by degrading aromatic compounds into simpler compounds (Ahmad et al., 2020). On the basis of these findings, we inferred that the significant increase in algae-lysing bacteria in the phycospheric microenvironment may participate in the degradation of algal cells, further decomposing the organic carbon and nitrogen substances released by them. Therefore, biodegradability may be an important reason for the enrichment of algae-lysing bacteria in the phycospheric microenvironment during the late stage of U. prolifera green tide.

Previous studies have shown that seagrass and some macroalgae surfaces can selectively regulate the adhesion and sedimentation of microorganisms in the surrounding environment to establish specific microbial communities, often referred to as the “host effect” (Roth-Schulze et al., 2016; Roth-Schulze et al., 2018). In our research, the topological network diagram revealed that U. prolifera in the demise stage had a similar mechanism (Figure 4). The number of nodes and edges in the phycosphere network is lower than that in the nonalgae-covered network, and the complexity of the network structure is also lower, which is consistent with the abovementioned findings that the microbial community diversity of the phycosphere decreases significantly in the late period. The lower average degree and graph density in the phycosphere network indicated that there were fewer connections in the symbiotic network of microorganisms in the phycosphere; therefore, U. prolifera entering the demise stage may reduce the interaction of microbial communities in the phycosphere. However, the specific regulatory mode still needs further experimental investigation.

In addition, we found that microorganisms play important roles in regulating the process of carbon, nitrogen and sulfur element transformation in the phycospheric microenvironment during the demise stage of green tide. Previous studies have found that phycospheric microorganisms can rapidly consume polysaccharides because of the presence of many polysaccharide utilization loci and specific substrate-binding proteins (Francis et al., 2021). In our research, CAZy enzymes were highly enriched in the phycosphere (Figure 5). Particularly, the abundance of GHs and PLs with decompose and utilize polysaccharides function in the phycospheric microenvironment is significantly higher than that in the surrounding seawater environment. This conclusion is consistent with the PLs enzyme play a major role in cleaving complex polysaccharides into simple oligomers or monomers (Chen et al., 2022; Mühlenbruch et al., 2018). Recent studies have demonstrated that various bacteria in the phycosphere display substrate preferences for the decomposition and utilization of polysaccharides (Lu et al., 2023; Zhao et al., 2022). Therefore, we infer that when U. prolifera demise, the release of a large amount of organic matter attracts microorganisms that hydrolyse or cleave this organic matter to gather and grow rapidly in seawater, which also allocates organic carbon resources. This can increase the metabolic efficiency of carbon in the phycospheric microenvironment, ultimately influencing the carbon transformation process.

Nitrogen transformation is an important part of the marine biogeochemical cycle (Zehr and Capone, 2020). In our study, genes associated with the processes of DNRA, denitrification and ANRA were found to be abundant within the phycosphere (Figure 6), highlighting the diversity of nitrogen transformation processes in this microenvironment. The nrfA is essential for DNRA process, facilitating the conversion of nitrate to ammonium, thus influencing nitrogen retention in ecosystems (Wang S. et al., 2020). The presence of nirS/K and norC are linked to denitrification, which convert nitrite to nitric oxide. This process effectively eliminate fixed nitrogen from the environment (Liao et al., 2024). U. prolifera consumes a lot of oxygen in the process of demise, which will lead to the formation of hypoxia or anaerobic environment, therefore it may be beneficial to the anaerobic metabolic process such as DNRA and denitrification (Liu et al., 2021). Additionally, the abundance of nrfA and nirS have been correlated with the rates of DNRA and denitrification, suggesting that these genes are crucial for understanding nitrogen dynamics in environment (Pandey et al., 2020). Furthermore, nasA involved in the ANRA process have the capacity to convert nitrite into ammonia, further contributing to nitrogen transformation (Wang et al., 2024). Previous findings showing that, a large amount of organic matter released during the decomposition of algae provide a rich source of nutrients for microorganisms and can promote the growth and reproduction of them (Broman et al., 2021). Thus for microbes involved in DNRA, denitrification and ANRA processes, the increase of nutrients may promote their activity and further increase the abundance of related genes. Consequently, the form and distribution of nitrogen will be altered, the process of nitrogen transformation will be accelerated, and the nitrogen cycle within the phycospheric microenvironment will be significantly impacted.

The abundances of the genes cysN/D/H/J, sir, sat and dsrA/B in the phycosphere were relatively high (Figure 5), indicating that the ASR and DSR pathways were significantly activated in the phycosphere. These two pathways are the core pathways of the marine sulfur cycle (Wang S. et al., 2023), and genes involved in these pathways can assimilate sulfide and convert it into cysteine, which is closely related to carbohydrate metabolism, carbon and nitrogen balance, protein synthesis and secondary metabolism, etc. (Sakurai et al., 2010). Thus, the sulfur reduction process is active in the phycospheric microenvironment, and there is a close relationship between the sulfur cycle and the carbon and nitrogen cycles (Figure 7).

Our research indicates that after U. prolifera green tides enter the demise stage, they can affect the phycospheric microenvironment, resulting in a distinct microbial community composition and structure. A large proportion of algae-lysing bacteria in the phycosphere also have unique functions and are crucial for niche adaptation and interactions. In addition, phycospheric microorganisms can increase the efficiency of carbon and nitrogen metabolism and thus significantly influence marine biogeochemical processes. Therefore, U. prolifera green tides during the demise stage could affect potential biogeochemical processes in the Yellow Sea by driving changes in phycospheric microbes. However, the role of microorganisms in this process within such a large-scale green tide, laden with organic biogenic elements, warrants further investigation through additional experiments.