the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
Phosphorus regulates fungal biomass production in a Norway spruce forest
- 1Lund University, Microbial Ecology, Dept of Biology, SE-223 62 Lund, Sweden
- 2Sveriges Lantbruksuniversitet (SLU), Department of Ecology, Ulls Väg 17, Uppsala, Sweden
- 3School of Science and Technology, Örebro University, SE- 701 82, Örebro, Sweden
- 4Center for Environmental and Climate Research, Lund University, SE-22362 Lund, Sweden
- 1Lund University, Microbial Ecology, Dept of Biology, SE-223 62 Lund, Sweden
- 2Sveriges Lantbruksuniversitet (SLU), Department of Ecology, Ulls Väg 17, Uppsala, Sweden
- 3School of Science and Technology, Örebro University, SE- 701 82, Örebro, Sweden
- 4Center for Environmental and Climate Research, Lund University, SE-22362 Lund, Sweden
Abstract. Ectomycorrhizal fungi (EMF) are important components of the soil microbial communities and EMF biomass can potentially increase carbon (C) stocks by accumulating in the soils as necromass and producing recalcitrant structures. EMF growth depends on the C allocated belowground by the host trees and the nutrient limitation on tree growth is expected to influence this allocation. Therefore, studying EMF production and understanding the factors that regulates it in natural soils is important to understand C cycling in forests.
Ingrowth meshbags are commonly used to estimate EMF production, but these measurements might not reflect the total EMF production since turnover rates of the hyphae are not considered. Here we estimated fungal production and turnover in response to P fertilization in a Norway spruce forest where nitrogen (N) deposition has resulted in phosphorus (P) limitation of plant production by using a combination of meshbags with different incubation periods and with Bayesian inferences. To test how localized patches of N and P influence EMF production and turnover we amended some bags with a nitrogen source (methylene urea) or P source (apatite). Additionally, the Bayesian model tested the effect of seasonality (time of meshbag harvesting) on fungal production and turnover.
We found that turnover of EMF and was not affected by P fertilization or meshbag amendment. P fertilization had a negative effect on EMF production in all the meshbag amendments suggesting a reduced belowground C allocation to the extramatrical mycelium under high P status. Apatite amendment significantly increased EMF biomass production in comparison with the pure quartz bags in the control plots but not in the P-fertilized plots. This indicates that P-rich patches enhance EMF production in P limited forests, but not when P is not limiting. Urea amendment had a general positive effect on EMF production, but this was significantly reduced by P fertilization, suggesting that a decrease in EMF production under high P status also will affect N foraging. Seasonality had a significant effect on fungal production and the differences registered between the treatments were higher during the warmer months and disappeared at the end of the growing season.
Many studies highlight the importance of N for regulating belowground C allocation to EMF in northern coniferous forests, but here we show that the P status of the forest can be equally important for belowground carbon allocation to EMF production in areas with high N deposition.
Juan Pablo Almeida et al.
Status: final response (author comments only)
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RC1: 'Comment on bg-2022-165', Anonymous Referee #1, 08 Oct 2022
The study uses experiments and models to assess ectomycorrhizal fungal (EMF) biomass production and turnover during five months in a Norway spruce forest. It is combining an experimental series on the EMF production with localized N or P amendments with modelling and testing both for dependency on the growing season. It reads smoothly and the content is easy to follow. Significance of the work is already given by the importance of the host tree species and its role for boreal geochemical cycles.
Novel aspects are in acknowledging that turnover rates of fungal hyphae might differ according to season and combining phosphorus fertilization of a P-limited forest with testing localized N and P amendments at the same time. Its results confirm and extend earlier findings on EMF behaviour under different P and N regimes and are well summarized in the abstract (L21-L32).
However, there are discrepancies between the expectations the title stirs by its wording (“fungal biomass”), the aim formulated in the introduction (L128-L130), the hypotheses (L146-152) and the methods. Wording and presentation of goals of the manuscript should be unified and unique aims/results be highlighted: 1) please choose a term that can be derived from the methods and use it uniformly, e.g., “EMF biomass production”, because the distinction from similar terms (“fungal biomass”) throughout the manuscript is unclear. 2) the hypotheses are unconnected bullet-points, which is making them very generic; please integrate them into the last part of the introduction together with the reasoning behind them, especially as a body of prior studies followed similar questions. 3) turnover/seasonality was a central aspect in the study, it should be involved in a hypothesis, too.
Specific Comments:
- Title: “fungal biomass production” includes all types of fungal lifestyles, including the quantitatively very important group of saprotrophs. However, most of the manuscript uses the term “EMF production”. Therefore, I suggest to change the wording or explain in detail in the text how EMF production could be a proxy for other fungal lifestyles.
- Abstract:
L12: To me, it is not clear, how you arrive at “fungal production” here, when you specify in L10 that you estimated EMF production. This should be clarified throughout the manuscript.
- Introduction:
L86: EMM abbreviation is not explained. Do you mean extrametrical mycelium? (same in L502, L569)
L97: Ekblad et al .2016 do not use the term fungal standing biomass
- Methods:
Experiments: was there any kind of control to examine the share of non-EMF fungi in the ingrowth bags, like ingrowth bags in a root-free area of soil, or amplicon sequencing of the EMF that were found in the ingrowth bags? – It would be very helpful to have clear knowledge to which degree this experiment was able to capture the term “EMF biomass production” used throughout the manuscript.
L188-190: drilling a new hole and placing an ingrowth bag in it and re-placing an ingrowth bag by putting it into an existing hole seem to be two different kinds of disturbance. Is there any knowledge on this?
Models: I am not versed in modelling and Bayesian inference, wherefore I could not review parts based on this in detail. However, the models seem well thought trough. Unfortunately, there is no explanation of data sources. The statements “the methodology allows us to draw information from publications” (L313) and “Priors for ðk and ðk were derived from the literature” (L322) are too vague. How was this done and which publications where used? Please explicitly state if this is based on data from Hagenbo et al. 2017 (L334) or new estimations. -- All data sources should be clarified and the data made available.
- Results:
L380: do you mean “… apatite, urea, and not amended meshbags”?
L389-294 / Fig. 2: Please mention the number of data-points for each boxplot, either by plotting or mentioning n in the figure caption. As I understood the methods section, each box in figure 2 is resembling three samples (one pooled sample per plot), accordingly n = 3. In this case, a boxplot is not very useful in summarizing the data and another type of graph could better be chosen.
L398-403 / Fig. 3: having connecting lines between the points, on first glance indicates a time series with one starting point, but in fact there are for example several 30 and 60 day starting points and the samples are independent of each other. Therefore, deleting the lines (or making them dotted) would be useful. Additionally, labelling the x-axis with “incubation time of ingrowth bags [days]” would also help for understanding.
- Discussion:
L481-482: “The fact that more incubation periods and a larger number of bags were used makes the present study more reliable.” Please clarify: more reliable than what study? And what is the difference implied by “more … periods” and “larger number”?
L486: the term “extrametrical mycelium” has not been introduced in the text, so far.
L486-487 “P as a nutrient regulating fungal growth in boreal forest was not reported before”: Please exactly define what fungal growth stands for in this case or reword for avoiding conflicts with earlier studies. For example, Aleida et al. 2019 (ref. in this manuscript) already wrote for the same forest: “Soil EMF communities responded more strongly to P than to N” which can be read as P is regulating fungal fungal growth. Not to mention the body of literature therein: “Ekblad et al. (1995) found that the production of extramatrical mycelium peaked under low P conditions. In a field study comparing Norway spruce (Picea albies) forests of varying P status, Rosenstock et al. (2016) observed greatly enhanced EMF biomass from ingrowth meshbags in the P limited forest […] in P-limited forests, fungal biomass is enhanced by the presence of mineral P sources like apatite (Hagerberg et al., 2003; Berner et al., 2012; Rosenstock et al., 2016). Bahr et al. (2015) reported that apatite addition stimulated ingrowth of EMF in meshbags, especially in N-fertilized plots.” (Almeida et al. 2019, https://doi.org/10.1016/j.funeco.2018.05.008). Why aren’t those references seen as reports of P as nutrient which is regulating fungal growth?
L504: It would be worth mentioning that an independent second method measuring the decrease in belowground C allocation due to P is needed for verification in further studies.
Technical corrections:
L21: “EMF and was” – missing word
L65: missing period
L118: EFM?
L389-294 / Fig. 2: please label left and right panels (a, b). Remove the cluttering design of R-ggplot’s standard output (grid lines, grey facet-boxes. Y-axis label: what is “per g” referring to? Quartz? Please choose a more exact way to label the y-axis like “µg [Ergosterol] / µg […]”
L398-403 / Fig. 3: Remove the legend and explain in the caption. Remove the cluttering design of R-ggplot’s standard output (grid lines, grey facet-boxes. Y-axis label: what is “per g” referring to? Quartz? Please choose a more exact way to label the y-axis like “µg [Ergosterol] / µg […]”
L604 typo: not --> no
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AC1: 'Reply on RC1', Juan Almeida, 03 Dec 2022
The comment was uploaded in the form of a supplement: https://bg.copernicus.org/preprints/bg-2022-165/bg-2022-165-AC1-supplement.pdf
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AC1: 'Reply on RC1', Juan Almeida, 03 Dec 2022
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RC2: 'Comment on bg-2022-165', Anonymous Referee #2, 14 Oct 2022
This is an interesting and timely manuscript, but the presentation needs to better structured and more succinct. The ms revisits Almeida et al. 2018 with many design and analytical improvements (and more untested assumptions) and some different findings.
The Discussion in particular is too long, it should be broken into sections e.g. hypotheses, limitations. The paper overall needs to be more readable, easier to navigate for the reader. Redundancy/repetition should be reduced. There are also too many figures and tables, need to select the most important ones and move the rest to supplement.
Unfortunately, no links are made to recent papers on nutritional thresholds or tipping points in EM for European (incl. spruce Swedish) forests. There are no links made to recent studies of P limitation (i.e. beech in Germany). There is no mention of mycorrhizal fungi supressing decomposers (Gadgil effect). There is frequent mention of tree C allocation, as if this was being measured directly, but this is not the case. It is indirect and incomplete (via fungal growth and without respiration); so this needs to be qualified consistently throughout, e.g. "potential tree C allocation". It is often unclear if apatite amendment is the same as P fertilization, which it is not; both are used interchangeably, which is confusing.
The abstract needs to mention how P fertilization was done, e.g superphosphate twice.
24 - high P status is vague - of soil, fungus, tree? What is the standard/threshold used? What is the evidence for P limitation?
72. Quantifying biomass and growth?
78. Is there evidence for nearly zero non-ECM growth into bags?
91-3. Is there any evidence supporting these assumptions?
157. Total experimental site area?
163. Is this above published thresholds for EM?
166. Why 200kg?
171. Why 50um?
172. Acid-washed quartz?
173. Why 2%, 50-250nm, 0.5%?
181. Why 30, 60, 90, 120 or 150d?
215-6. Would there be significant disturbance effects?
219. Up to how long?
221. Why not chitin or nucleic acids?
302. Which relative (related?) publications?
367. Were there normal average temp and ppt at the site in 2015 summer/autumn?
Figure 2. Why show this as 1 vs 2 months? Redundant?
Figure 3. Could saprobes use carbon in urea?
Discussion: Any practical recommendations for future studies? Tradeoffs in replication/design/power?479. shown indirectly in the current results?
480. P means superphosphate and/or apatite?
505. and/or shifted EM community? Could these cross the EM tipping point?
511. P-limitation - were trees tested, e.g. foliar chemistry?
523-524 - observed or assumed?
528. smooth mantle, not mycelium
514-538. This paragraph could be shorter (less speculative) and more sophisticated/precise. Why talk about Tylospora when no fungal community data is presented? I am hoping there won't be a follow up paper where fungal community data is presented separately, if it is available, it should be presented here.
601. But not measured, is this this a hypothesis?
616-617 - explain in methods section.
700 - P - meaning superphosphate and/or apatite?-
AC2: 'Reply on RC2', Juan Almeida, 03 Dec 2022
The comment was uploaded in the form of a supplement: https://bg.copernicus.org/preprints/bg-2022-165/bg-2022-165-AC2-supplement.pdf
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AC2: 'Reply on RC2', Juan Almeida, 03 Dec 2022
Juan Pablo Almeida et al.
Juan Pablo Almeida et al.
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