| 10 Oct 2025
Fluvial biofilm phosphorus entrapment shifts from intracellular to extracellular dominance along a multifactorial longitudinal river gradient
Abstract. Phosphorus (P) entrapment by biofilms can be distinguished into the intracellular P entrapment (the P uptake by microbial cells) and the extracellular P entrapment via the extracellular polymeric substances. It is unknown how these two P entrapment pathways behave in natural ecosystems in which gradients of environmental drivers such as P, labile dissolved organic matter (DOM) and light occur. Another key aspect in P dynamics at the sediment-water interface is microbial activity. Microbially mediated mineralization of DOM and microbial activity in general is suspected to be a driver of internal P mobilization from the sediments. Here we brought such expectations into a real-world context by analysing the P entrapment patterns of benthic biofilms and the P release potential from the sediments along a longitudinal gradient in a third order river in front of the background of key microbial metabolic variables. The gradient consists of increasing P availability, DOM lability and light availability.
We found a gradual shift in the dominance of P entrapment from higher intracellular P entrapment in the upstream biofilms to higher extracellular P entrapment in the downstream biofilms. This shift towards dominance of extracellular P entrapment was accompanied by an increase in the P release potential from the sediment. The increasing P release potential was also connected to high extracellular enzyme activity of alkaline phosphatase, an enzyme involved in the mineralization of P from organic compounds. We further found that a balanced ratio between intracellular and extracellular P was connected to a higher C metabolic diversity.
All this evidence suggests an influence of the benthic biofilms on P dynamics at the sediment-water interface. This research advocates for a more integrated perspective that accounts for both intracellular and extracellular biofilm-mediated processes.
The authors present data on phosphorus and carbon turnover characterization in benthic biofilms of a river stretch. The data include results from a large number of laboratory classification approaches (including biofilm composition, enzyme activity and P release experiments) for 7 positions in a 30km stream stretch, sampled once in autumn of 2023 under baseflow. The results are imbedded in a research question that surrounds the role of the biofilms as P retainers of various form (cells and their biofilm) and how their relation is driven.
The manuscript is well structured, characterizing the field site and its boundary conditions and then successively introducing the dynamic biofilm characteristics. The discussion section describes the systems effectively without repetition from the results section. Methods and statistics seem rigorous.
Despite the technical benefits, I struggle with the overall novelty of the manuscript. I study aquatic biogeochemistry across various environments, not stream biofilms in particular, and it seems to me as if this manuscript suffers from an overly deep insight paired with too limited spatial and temporal coverage. With their dataset, the authors try to establish a novel connection between carbon metabolism and P retention mechanisms. It escaped me, unfortunately, what the larger benefit of the knowledge of such a tight-knit dependence would be. This may be due to my limited insight to biofilm mechanics, but also to the limited structure of the introduction section. Especially the “open questions” are mixed with background knowledge and presented independent from the (too many and unclear) hypotheses. I’d like to motivate the authors to review this section with great care. This assessment it seems to me is mirrored in the discussion section. Next to the (solid, but to a large extent already published) presentation of the riverine gradient, there is a lot of text that revolves around the argumentation why the analyzed system is three-fold partitioned and why some of the other patterns observed in the previous studies trigger some of the observations made here (e.g. DOM quality, 490f). Section 4.5, which would be the heartpiece of the manuscript as currently presented, is not sufficiently clear. Overall my conclusion is that the authors go beyond what the limited observational density can provide.
For a revision, I think this manuscript needs a better focused introduction and a better imbedding of the results. I understand that the many analyses were time-consuming and could not be rolled out to the full network as in Weitere et al. network description. Despite that, I think the data has the potential to couple the observations from the 7 sites with the findings across the larger network and maybe even pair it with numerical data from other sites and studies. I don’t want this review to be read as a straight-out recommendation for rejection, as I see merit in the rigorous work. However, I feel for a strong broad-topic biogeo journal as BG, the study motivation, depths and conclusions should be more accessible to a broad community of researchers, which I think is not achieved in the current form, but authors and data have the potential for.
Some more specific comments
46-51 is it possible to add quantities to these mechanistic background. It becomes more tangible if the pool sizes of intra and extra-P are better known
57 “increasing gradients” is maybe not a good terminology here. It is gradients of increasing XYZ
64 not sure if Hupfer and Lewandowski, 2008 really had dissolved OM in mind, it is more about solid-phase OM
60-80 maybe it is possible to state the 2 open research question in a more systematic fashion (e.g. in two paragraphs, each focused on one of the topics: labile DOM->APA and the P entrapment gradient)
100 write out upstream and downstream
149 do you put the sediments in the P buffer solutions?
250 I do not understand why the zero-inflation was necessary in the data structure
401 Increasing gradient.. again this seems like a wrong terminology. What is “light DOM availability”
404 As an example here but also elsewhere, the discussion section could be clearer if it more directly links to the results figure in question.
447 the biofilm as natural P buffer… this is a relatable concept that may be useful also elsewhere
459 -465 Unclear what this statement is leading to. Discussion is not developing more than just the presentation of results.