Exchangeable manganese regulates carbon storage in the humus layer of the boreal forest

Abstract

The huge carbon stock in humus layers of the boreal forest plays a critical role in the global carbon cycle. However, there remains uncertainty about the factors that regulate below-ground carbon sequestration in this region. Notably, based on evidence from two independent but complementary methods, we identified that exchangeable manganese is a critical factor regulating carbon accumulation in boreal forests across both regional scales and the entire boreal latitudinal range. Moreover, in a novel fertilization experiment, manganese addition reduced soil carbon stocks, but only after 4 y of additions. Our results highlight an underappreciated mechanism influencing the humus carbon pool of boreal forests.

Full text

PNAS 2024 Vol. 121 No. 13 e2318382121 https://doi.org/10.1073/pnas.2318382121 1 of 3
BRIEF REPORT
|
Author aliations: aKey Laboratory of Forest Ecology
and Management, Institute of Applied Ecology, Chinese
Academy of Sciences, Shenyang 110016, China; bCollege
of Resources and Environment, University of Chinese
Academy of Sciences, Beijing 100049, China; cDepartment
of Ecology, Evolution and Behavior, University of Minne
sota, St. Paul, MN 55108; dEarth and Climate Sciences
Division, The Nicholas School of the Environment, Duke
University, Durham, NC 27710; eDepartment of Forest
Sciences, University of Helsinki, Helsinki FIN- 00014,
Finland; fQingyuan Forest Chinese Ecosystem Research
Network, National Observation and Research Station,
Liaoning Province, Shenyang 110016, China; gLiaoning
Key Laboratory for Management of Non- commercial
Forests, Shenyang 110016, China; hKey Laboratory of
Terrestrial Ecosystem Carbon Neutrality, Liaoning Province,
Shenyang 110016, China; iChinese Academy of Sciences-
Campbell Scientic Inc. Joint Laboratory of Research and
Development for Monitoring Forest Fluxes of Trace Gases
and Isotope Elements, Shenyang 110016, China; and jSino-
USA Joint Laboratory of Forest Ecology and Silviculture,
Shenyang 110016, China
Author contributions: T.S. designed research; Y.Z. and
T.S. performed research; Y.Z. and T.S. analyzed data; and
Y.Z., S.E.H., W.H.S., B.B., and J.Z. wrote the paper.
The authors declare no competing interest.
Copyright © 2024 the Author(s). Published by PNAS.
This open access article is distributed under Creative
Commons Attribution- NonCommercial- NoDerivatives
License 4.0 (CC BY- NC- ND).
1To whom correspondence may be addressed. Email:
sunt@iae.ac.cn.
This article contains supporting information online at
https://www.pnas.org/lookup/suppl/doi:10.1073/pnas.
2318382121/ /DCSupplemental.
Published March 19, 2024.
ECOLOGY
Exchangeable manganese regulates carbon storage in the
humus layer of the boreal forest
YunyuZhanga,b , SarahE.Hobbiec, WilliamH.Schlesingerd, BjörnBerge, TaoSuna,1, and JiaojunZhua,f,g,h,i,j
Edited by Simon Levin, Princeton University, Princeton, NJ; received December 20, 2023; accepted February 21, 2024
e huge carbon stock in humus layers of the boreal forest plays a critical role in the
global carbon cycle. However, there remains uncertainty about the factors that regulate
below- ground carbon sequestration in this region. Notably, based on evidence from two
independent but complementary methods, we identified that exchangeable manganese
is a critical factor regulating carbon accumulation in boreal forests across both regional
scales and the entire boreal latitudinal range. Moreover, in a novel fertilization experi-
ment, manganese addition reduced soil carbon stocks, but only after 4 y of additions.
Our results highlight an underappreciated mechanism influencing the humus carbon
pool of boreal forests.
manganese | carbon sequestration | decomposition | biogeochemical cycle | humus layer
e boreal forest is estimated to hold 30% of global terrestrial carbon (C) stocks, and
60% of this C is located underground (1). Approximately one- third of that soil C is found
in organic soil horizons (2). us, the stability of boreal soils is critical to understanding
C cycle feedbacks to climate change and soil management (3). Yet, regulation of humus
C sequestration in the boreal forests globally is not entirely understood.
Previous studies proposed that the potential controls of humus C storage in boreal
regions include climate (2, 4), exchangeable manganese (Mn) and potassium (K) (5), and
mycorrhizal fungi (6). Recent studies have emphasized that soil C cycling in boreal forests
depends on mycorrhizal associations (6) and enzymatic depolymerization by microorgan-
isms (7). Emerging research on fungal, chemical, and climatic inuences on soil C has
concluded that Mn is an especially important control over organic matter decomposition
and boreal C stocks at local to regional scales (5, 8, 9), yet is often overlooked in global
analyses (7). ough climate has always been recognized to play a critical role in regulating
the persistence and decomposition of soil organic matter (SOM) in boreal regions (4),
the availability of Mn could overshadow its inuence (5, 8). Earlier investigations suggest
that exchangeable Mn and C storage in organic horizons are negatively correlated (8, 10).
Keiluweit et al. (11) identied that Mn- redox cycling mediated by litter- decomposing
fungi enhanced the oxidative breakdown of lignin and other aromatic litter components,
thus implying that Mn bioavailability to the fungal community signicantly reduces
humus C storage by stimulating the rate and extent of organic matter decomposition.
Notably, Mn appears to be the dominant factor facilitating decomposition of the most
stable fraction of litter (5, 11), suggesting that it could play a major role in regulating C
stocks in organic horizons of boreal soils.
Yet, most of the past work on Mn eects on decomposition has focused on litter
decomposition; there has been limited work examining Mn regulation of soil organic C
stocks in boreal forests (6) on broader spatial scales. Nor have there been experimental
studies evaluating the role of Mn as a primary driver regulating C sequestration in the
boreal forest humus. In this paper, we combine meta- analysis and a novel long- term
experiment to investigate the eect of Mn on boreal forest humus C stocks. Our results
reveal a long- overlooked problem—the stimulation of the degradation of soil C by Mn
within a broader ecosystem spatiotemporal context. is issue is particularly signicant
when considering the eect of human activities on cycling of Mn through vegetation and
soils.
Results
We rst ascertained how climate and element concentrations relate to boreal humus C
pools using a meta- analysis of boreal forest data encompassing the whole boreal zone
(Fig. 1A). Among exchangeable cations considered (Mn, K, Ca, Na, and Mg), we found
that C stocks in the humus layer were primarily and negatively correlated with exchange-
able Mn (Mn held on soil cation exchange sites) (R2 = 0.372, P < 0.0001, Fig. 1B).
Exchangeable Ca had a weaker correlation (R2 = 0.010, P < 0.05), whereas other
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2 of 3 https://doi.org/10.1073/pnas.2318382121 pnas.org
exchangeable elements (K, Na, and Mg) showed no signicant
relationship with humus C stocks (P > 0.05). Among total cations
(Mn, K, Ca, Mg, S, Al, and Fe), Mn, Mg, Al, and Fe were also
related to C stocks (R2 = 0.053, 0.022, 0.031, and 0.013, respec-
tively; P < 0.05). e C/N ratio was also related to soil C stocks,
but only explained 1.5% of the variation in stored soil C (P <
0.05, R
2
= 0.015). ere were no signicant relationships between
C storage and pH, mean annual precipitation, or mean annual
temperature (P > 0.05). Among all of the parameters tested above,
exchangeable Mn concentration best predicted boreal humus C
pools.
To experimentally examine the eect of Mn on humus soil C
stocks, we conducted a long- term Mn fertilization study in a 50- y- old
Larix gmelinii plantation at Huzhong Forest Bureau of Daxing'an
Mountains (52°02′N, 123°36′E). Over the 14- y experimental
period, humus C storage in both control and plots fertilized with
MnSO
4
showed a signicant and overall increase (P < 0.001, Fig. 2).
ere was no dierence between control and Mn- fertilized plots in
humus C stocks during the rst 4 y (2009 to 2012, P > 0.05).
However, over the next decade, Mn fertilization reduced the size of
humus C stocks signicantly compared to control plots, by 13.3%
loss (−3.27 t/ha, P < 0.05, Fig. 2).
Discussion
ese complementary investigations oer consistent evidence that
Mn was a signicant factor controlling long- term humus C
sequestration in boreal forest soils and provide the rst generalized
Mn eect on organic soil C stocks for boreal forests on a global
scale (P < 0.0001, Fig. 1B). While climatic controls have long
been recognized as important for C storage in high- latitude
regions (4), our ndings add to growing evidence of the impor-
tance of Mn. After subdividing the data of Sweden into 9 climate
regions, Stendahl et al. (5) similarly found that in soils with high
Mn concentrations, C storage in sites of all climate regions stabi-
lized at equally low levels, indicating limited climatic inuence in
C storage when available Mn was high. e presence of specic
ectomycorrhizal fungi is closely related to organic topsoil C stocks,
as the decomposition of this C pool requires specic peroxidases
produced by the fungi (6). A reasonable explanation for the
dependence of organic matter decomposition in humus layers on
Mn is that Mn stimulates both production and activity of Mn
peroxidase (MnP) (9, 11, 12), a crucial extracellular enzyme only
produced by specic members of the Agaricomycetes (13). Dierent
from lignin peroxidase, which attacks lignin bonds directly, MnP
has a regulating eect on the long- term decomposition of litter
and soil organic C by taking an electron from Mn2+, then driving
the Mn redox cycle (5, 11, 14), and penetrating the phenolic
structure of lignin and lignin- like compounds (12). Litter-
decomposing fungi oxidize accumulated Mn
2+
into highly reactive
Mn3+ that is coupled tightly to the loss of saccharides and the
transformation of aromatic litter and soil C components (11). In
summary, Mn- dependent oxidation constitutes the major mech-
anism for degradation of stable residues (12). Consistent with this
interpretation, exchangeable Mn was shown to increase the lability
of soil C in a boreal forest recovering from wildre (8). Notably,
other potential regulators of the C pool, such as soil N concen-
tration (6) and N deposition (9), were secondary in their eects
on soil C pools and had indirect eects through altering fungal
activity. In addition to boreal forests (5, 6, 8, 10), negative corre-
lations between Mn and humus C pool have been documented
in temperate forests (8–10).
e response of the humus C pool to Mn fertilization provided
direct evidence for the Mn eect on C storage (P < 0.001, Fig. 2),
consistent with the positive response of MnP activities to Mn
fertilization found in other studies (9). Notably, the eects of Mn
addition on soil C stocks were not apparent until after 4 y (Fig. 2),
possibly because it took time for Agaricomycetes to increase in
Fig.1. Relationship between exchangeable Mn concentrations and soil C stocks in humus layers of boreal forests. (A) Area of the boreal forest and location of
study sites in the database (n = 2,538). (B) Log–log regression between C stocks and exchangeable Mn in the humus layer (n = 2,437).
Fig.2. Eect of annual experimental Mn fertilization (MnSO4) on humus C
storage of the boreal forest over 14 y. Points show mean soil C stocks in
organic horizons (humus) in control and Mn- fertilized plots in a Larix gmelinii
stand in boreal China (Huzhong Forest Bureau of Daxing’an Mountains). Bars
are means ± SEM.

PNAS 2024 Vol. 121 No. 13 e2318382121 https://doi.org/10.1073/pnas.2318382121 3 of 3
abundance and activity in response to the increased Mn availability
and changes in the soil environment (6, 8). ese ndings under-
score the need for additional long- term Mn fertilization studies
to better understand the generality and magnitude of the eects
demonstrated here. Emerging evidence (15) shows that long- term
industrial airborne pollution has diminished the Mn concentra-
tion of coniferous litterfall in some northern pine and spruce
forests, resulting from the long- term impact of acidic precipitation
leaching Mn from plant tissues (15–17). As Mn is not reabsorbed
during leaf senescence, litterfall is the essential pathway for Mn
to return to soil (7). is direct anthropogenic disturbance on
Mn- plant cycling in boreal forest ecosystems could be important
to forest management, the state and ecological function of boreal
forest ecosystems, and climate feedbacks over ecologically realistic
time frames (>10 y).
Using two dierent approaches, our work demonstrates that in
both the global boreal latitudinal zone and in a 10- y experiment at
a single site, Mn plays a fundamental role in boreal forest humus
preservation, leading to substantially enhanced understanding of
variation in soil C stocks worldwide. Our work provides a critical
unifying framework regarding the eect of exchangeable Mn on the
long- term persistence and decomposition of SOM, which has impli-
cations for the formation and stabilization of litter- derived C within
the humus. Our eorts thus suggest that a revised representation of
Mn- mediated processes inuencing C storage in biogeochemical
models would boost condence in the structure, parameter esti-
mates, and projections of those models related to the processes of
litter decomposition and SOM formation.
Materials and Methods
Data Collection. We compiled a database of humus C (t/ha). See SIAppendix
for details of data collection.
Mn Fertilization. We established a fertilization experiment at Daxing'an
Mountains (52°02′N, 123°36′E) in 2009. See SIAppendix for research site char-
acteristics, experimental design, and laboratory analyses.
Data Analysis. Forest cover data were accessed via EarthEnv (18). All analyses
were performed in SPSS 22 (α = 0.05). Data displayed in figures were extracted
with WebPlotDigitizer4.6 and Get Data Graph Digitizer. Plots were drawn using
ArcMap10.8 and Origin2023b. See SIAppendix for statistical analyses.
Data, Materials, and Software Availability.
All study data are included in the
article and/or SIAppendix.
ACKNOWLEDGMENTS. This research was supported by the National Natural
Science Foundation of China (32192432 and 32022054), the National Key R&D
Program of China (2022YFD2201300 and 2020YFA0608100), the International
Partnership Program of CAS (151221KYSB20210005), and the Excellent Member
of Youth Innovation Promotion Association of CAS to T.S.
1. Y. Pan et al., A large and persistent carbon sink in the world’s forests. Science 333, 988–993 (2011).
2. M. Fröberg et al., Mean residence time of O horizon carbon along a climatic gradient in Scandinavia
estimated by C- 14 measurements of archived soils. Biogeochemistry 104, 227–236 (2011).
3. W. H. Schlesinger, E. S. Bernhardt, "Chapter5–The carbon cycle of terrestrial ecosystems" in
Biogeochemistry, W. H. Schlesinger, E. S. Bernhardt, Eds. (Academic Press, ed. 4, 2020), p. 180.
4. S. E. Hobbie, J. P. Schimel, S. E. Trumbore, J. R. Randerson, Controls over carbon storage and
turnover in high- latitude soils. Global Change Biol. 6, 196–210 (2000).
5. J. Stendahl, B. Berg, B. D. Lindahl, Manganese availability is negatively associated with carbon
storage in northern coniferous forest humus layers. Sci. Rep. 7, 15487 (2017).
6. B. D. Lindahl et al., A group of ectomycorrhizal fungi restricts organic matter accumulation in boreal
forest. Ecol. Lett. 24, 1341–1351 (2021).
7. H. Li, F. Santos, K. Butler, E. Herndon, A critical review on the multiple roles of manganese
in stabilizing and destabilizing soil organic matter. Environ. Sci. Technol. 55, 12136–12152
(2021).
8. B. Andrieux, D. Pare, J. Beguin, P. Grondin, Y. Bergeron, Boreal- forest soil chemistry drives soil
organic carbon bioreactivity along a 314- year fire chronosequence. Soil 6, 195–213 (2020).
9. E. D. Whalen, R. G. Smith, A. S. Grandy, S. D. Frey, Manganese limitation as a mechanism for reduced
decomposition in soils under atmospheric nitrogen deposition. Soil. Biol. Biochem. 127, 252–263
(2018).
10. F. Santos, E. Herndon, Plant- soil relationships influence observed trends between manganese and
carbon across biomes. Global Biogeochem. Cycles 37, e2022GB007412 (2023).
11. M. Keiluweit et al., Long- term litter decomposition controlled by manganese redox cycling. Proc.
Natl. Acad. Sci. U.S.A. 112, E5253–E5260 (2015).
12. M. Hofrichter, Review: Lignin conversion by manganese peroxidase (MnP). Enzyme Microb. Technol.
30, 454–466 (2002).
13. D. Floudas et al., The Paleozoic origin of enzymatic lignin decomposition reconstructed from 31
fungal genomes. Science 336, 1715–1719 (2012).
14. F. Passardi et al., Prokaryotic origins of the non- animal peroxidase superfamily and organelle- mediated
transmission to eukaryotes. Genomics 89, 567–579 (2007).
15. E. A. Ivanova, N. V. Lukina, V. E. Smirnov, L. G. Isaeva, Effect of industrial airborne pollution on the
chemical composition of pine needle litterfall at the northern distribution limit of pine forests.
Contemp. Probl. Ecol. 15, 851–862 (2022).
16. N. V. Lukina et al., Mass- loss rates from decomposition of plant residues in spruce forests near the
northern tree line subject to strong air pollution. Environ. Sci. Pollut. Res. Int. 24, 19874–19887 (2017).
17. J. Kopáček et al., Composition of Norway spruce litter and foliage in atmospherically acidified and
nitrogen- saturated Bohemian Forest stands, Czech Republic. Boreal Environ. Res. 15, 413–426 (2010).
18. M.- N. Tuanmu, W. Jetz, A global 1- km consensus land- cover product for biodiversity and ecosystem
modelling. Global Ecol. Biogeogr. 23, 1031–1045 (2014).