我们进行了一项实地研究,以调查在淡水湖(犹他州湖)盛开的各个阶段营养缺乏期间,严格响应在蓝细菌和共存的浮游细菌中的作用。使用宏基因组学和超转录组学分析,我们检查了蓝藻生态学和与严格响应相关的重要功能基因的表达,N和P代谢,和监管。我们的发现标志着在理解有毒蓝细菌在氮(N)和磷(P)限制下生存和增殖的机制方面取得了重大进展。我们成功地鉴定并分析了显性水华形成蓝细菌的宏基因组组装基因组(MAG),即,圆形Dolichospermum,Aphanizomenonflos-aquaeUKL13-PB,Planktothrixagardhii,和铜绿微囊藻。通过将RNA-seq数据映射到MAG的编码序列,我们观察到,这四种流行的蓝藻物种激活多种功能,以适应无机营养素的消耗。在开花期间和之后,四种优势蓝藻物种表达了与毒素产生相关的高水平转录本,如微囊藻毒素(mcy),Anatoxins(ana),和cylindrospermopsin(cyr)。此外,与多磷酸盐(poly-P)储存和严格响应alarmone(p)ppGpp合成/水解相关的基因,包括ppk,rela,和spot,在蓝细菌和浮游细菌中均高度激活。在N缺乏下,在某些蓝细菌中,主要的N途径从浮游细菌中的反硝化和异化硝酸盐还原转变为N2固定和同化硝酸盐还原,群落组成也相应变化。P剥夺触发了蓝细菌和浮游细菌中spoT依赖性(p)ppGpp积累和Pho调节子激活介导的严格反应,促进无机和有机磷的吸收。显性蓝细菌MAG表现出多种碱性磷酸酶(APase)转录本的存在(例如,在Dolichospermum中的phoA,phoX在浮游生物中,和微囊藻),表明它们具有合成和释放APase酶以将环境有机P转化为生物可利用形式的能力。相反,与浮游细菌为主的途径如反硝化相关的转录本很低,并且与强烈的氰化HAB的发生不一致。在N之间观察到的强相关性,P,严格的响应代谢和由优势蓝细菌物种引起的水华演替提供了证据,由营养限制引起,可能激活产生毒素的蓝细菌中独特的N和P功能,从而维持cyanoHABs。
We conducted a field study to investigate the role of stringent response in cyanobacteria and coexisting bacterioplankton during nutrient-deprived periods at various stages of bloom in a freshwater lake (Utah Lake) for the first time. Using metagenomics and metatranscriptomics analyses, we examined the cyanobacterial ecology and expression of important functional genes related to stringent response, N and P metabolism, and regulation. Our findings mark a significant advancement in understanding the mechanisms by which toxic cyanobacteria survive and proliferate during nitrogen (N) and phosphorus (P) limitations. We successfully identified and analyzed the metagenome-assembled genomes (MAGs) of the dominant bloom-forming cyanobacteria, namely, Dolichospermum circinale, Aphanizomenon flos-aquae UKL13-PB, Planktothrix agardhii, and Microcystis aeruginosa. By mapping RNA-seq data to the coding sequences of the MAGs, we observed that these four prevalent cyanobacteria species activated multiple functions to adapt to the depletion of inorganic nutrients. During and after the blooms, the four dominant cyanobacteria species expressed high levels of transcripts related to toxin production, such as microcystins (mcy), anatoxins (ana), and cylindrospermopsins (cyr). Additionally, genes associated with polyphosphate (poly-P) storage and the stringent response alarmone (p)ppGpp synthesis/hydrolysis, including ppk, relA, and spoT, were highly activated in both cyanobacteria and bacterioplankton. Under N deficiency, the main N pathways shifted from denitrification and dissimilatory nitrate reduction in bacterioplankton toward N2-fixing and assimilatory nitrate reduction in certain cyanobacteria with a corresponding shift in the community composition. P deprivation triggered a stringent response mediated by spoT-dependent (p)ppGpp accumulation and activation of the Pho regulon in both cyanobacteria and bacterioplankton, facilitating inorganic and organic P uptake. The dominant cyanobacterial MAGs exhibited the presence of multiple alkaline phosphatase (APase) transcripts (e.g., phoA in Dolichospermum, phoX in Planktothrix, and Microcystis), suggesting their ability to synthesize and release APase enzymes to convert ambient organic P into bioavailable forms. Conversely, transcripts associated with bacterioplankton-dominated pathways like denitrification were low and did not align with the occurrence of intense cyanoHABs. The strong correlations observed among N, P, stringent response metabolisms and the succession of blooms caused by dominant cyanobacterial species provide evidence that the stringent response, induced by nutrient limitation, may activate unique N and P functions in toxin-producing cyanobacteria, thereby sustaining cyanoHABs.