Hydrological transitions drive dissolved organic matter quantity and composition in a temporary Mediterranean stream

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Hydrological transitions drive dissolved organic matterquantity and composition in a temporary Mediterraneanstream

Daniel von Schiller • Daniel Graeber •

Miquel Ribot • Xisca Timoner • Vicenc Acuna •

Eugenia Martı • Sergi Sabater • Klement Tockner

Received: 1 September 2014 / Accepted: 29 January 2015 / Published online: 7 February 2015

� Springer International Publishing Switzerland 2015

Abstract The implications of stream flow intermitten-

cy for dissolved organic matter (DOM) are not well

understood despite its potential significance for water

quality and ecosystem integrity. We combined intensive

sampling with liquid chromatography and spectroscopic

techniques to follow changes in DOC and DON concen-

trations as well as in DOM size fractions and spectro-

scopic properties in a temporary stream during an entire

contraction–fragmentation–expansion hydrological cy-

cle. DOC and DON concentrations remained low

(range = 1.4–5.2 mg C L-1 and 0.05–0.15 mg N L-1)

during hydrological contraction and fragmentation, with

concomitant increases in the proportion of high mole-

cularweight substances (HMWS) during contraction and

of DOM aromaticity during fragmentation. DOC and

DON concentrations abruptly increased (up to

8.8 mg C L-1 and 0.37 mg N L-1) at the end of the

fragmentation phase, with a concomitant increase in the

non-humic, microbial and aquatic character of DOM.

Upon rewetting, the DOC and DON concentrations

reached their highest values (up to 12.7 mg C L-1 and

0.39 mg N L-1), with concomitant increases in the

proportion of HMWS and in the humic, aromatic and

terrestrial character of DOM. Subsequently, DOC and

DON concentrations recovered to values similar to those

at the contraction phase, while DOM composition

variables indicated the prevalence of a DOM of humic

and terrestrial character during the whole expansion

phase. Overall, our results emphasize the importance of

hydrological transitions forDOMdynamics in temporary

streams, and point to the potential response of perennial

streams under future water scarcity scenarios.

Responsible Editor: Sujay Kaushal.

D. von Schiller (&) � X. Timoner � V. Acuna �S. Sabater

Catalan Institute for Water Research, Emili Grahit 101,

17003 Girona, Spain

e-mail: [email protected]

D. Graeber

Department of Bioscience, Aarhus University, Vejlsøvej

25, 8600 Silkeborg, Denmark

M. Ribot � E. Martı

Integrative Freshwater Ecology Group, Centre for

Advanced Studies of Blanes (CEAB-CSIC), Acces a la

Cala St. Francesc 14, 17300 Blanes, Spain

X. Timoner � S. SabaterInstitute of Aquatic Ecology, University of Girona,

17071 Girona, Spain

K. Tockner

Leibniz-Institute of Freshwater Ecology and Inland

Fisheries, Muggelseedamm 301, 12587 Berlin, Germany

K. Tockner

Institute of Biology, Freie Universitat Berlin, Takustraße

3, 14195 Berlin, Germany

123

Biogeochemistry (2015) 123:429–446

DOI 10.1007/s10533-015-0077-4

Author's personal copy

Keywords Drying � Rewetting � Dissolved organic

matter � Dissolved organic carbon � Temporary

stream � Drought

Introduction

Temporary streams are waterways that cease to flow at

some points in space and time along their course

(Acuna et al. 2014). A major part of the world’s river

networks consists of temporary streams (Larned et al.

2010; Datry et al. 2014). For instance, more than half

of the total river length in South Africa (Uys and

O’Keefe 1997), USA (Nadeau and Rains 2007), and

Greece (Tzoraki et al. 2007) is subject to flow

intermittency, and most Alpine, Arctic, and Antarctic

rivers are temporary (Datry et al. 2014). A recent

global estimate shows that about two-thirds of the

first-order streams and one-third of the larger, fifth-

order rivers below 60� latitude experience flow

intermittency (Raymond et al. 2013). However, tem-

porary streams are widely neglected in policy imple-

mentation (Nikolaidis et al. 2013; Acuna et al. 2014).

At the same time, the biogeochemical implications of

stream flow intermittency are still not well understood,

despite of their potential relevance for stream ecosys-

tem functioning (Lake 2011; Steward et al. 2012).

Temporary streams are the dominant surface water

type in the Mediterranean Basin (Bonada and Resh

2013; Prat et al. 2014), and the duration and extent of

stream flow intermittency is expected to increase in

this region, as well as in other regions, in response to

climate and land use change (IPCC 2013). Mediter-

ranean temporary streams exhibit a highly dynamic

hydrological regime with a characteristic seasonal

drought period (Bernal et al. 2013). Typically, stream

flow gradually decreases in late spring to early

summer (contraction phase), followed by flow cessa-

tion and the formation of isolated pools (fragmentation

phase) before the stream completely dries up at the

surface (dry phase). At the end of summer, when

evapotranspiration in the catchment decreases and

intense precipitation events are frequent, stream flow

is gradually or abruptly re-established (expansion

phase). The drying and rewetting cycle controls both

the longitudinal hydrological connectivity along the

stream and the lateral hydrological connectivity

between the stream and its valley, with potentially

important but widely unexplored effects on in-stream

biogeochemical cycling (Dahm et al. 2003; von

Schiller et al. 2011; Vazquez et al. 2011).

Dissolved organic matter (DOM) is a complex

mixture of organic compounds, which represents the

largest pool of transported organic matter in streams

and plays an essential role in aquatic ecosystem

biogeochemistry (Findlay and Sinsabaugh 2003;

Prairie 2008; Tank et al. 2010). This role depends

not only on the quantity of DOM but also on its

composition, which result from a combination of

inputs from the catchment (i.e. allochthonous sources)

and from in-stream production (i.e. autochthonous

sources) (Webster and Meyer 1997). Among several

key functions, DOM supplies carbon and nitrogen for

heterotrophic production (Keil and Kirchman 1991;

Wetzel 1992), mediates the availability of metals

(Yamashita and Jaffe 2008), and modifies the optical

properties of waters (Rodrıguez-Zuniga et al. 2008).

Therefore, it is of fundamental interest to understand

the effect of flow intermittency on DOM quantity and

composition. Little is known, however, on the dy-

namics of DOM before and after flow cessation.

Hydrological contraction and fragmentation causes

the disruption of the longitudinal and lateral hydro-

logical pathways (Sabater and Tockner 2010), which

results in low-oxygen and acidic in-stream conditions

(Boulton and Lake 1990; von Schiller et al. 2011). In

addition, contraction and fragmentation influences the

availability of dissolved inorganic nutrients (Dent and

Grimm 1999; von Schiller et al. 2011) and promotes the

accumulation of particulate organicmatter (Boulton and

Lake 1992; Acuna et al. 2005). During these hydro-

logical phases, allochthonous DOM input from the

catchment is interrupted, which may lead to low

dissolved organic carbon (DOC) and nitrogen (DON)

concentrations when there is no relevant autochthonous

DOM source (Freeman et a. 1994; Dahm et al. 2003).

Hydrological contraction and fragmentation may sti-

mulate biotic processes such as algal blooms and

microbial uptake (Lake 2003; Fellman et al. 2011). In

addition, abiotic processes that affect in-stream DOM

such as evaporation and reduction of the dilution

capacity (Sabater and Tockner 2010), leaching of

particulate organic matter (McMaster and Bond 2008),

photodegradation (Rodrıguez-Zuniga et al. 2008) and

adsorption–desorption (Dahm 1981) may be enhanced.

In general, allocthonous DOM dominates the bulk pool

of DOM, but the proportion of DOM deriving from in-

430 Biogeochemistry (2015) 123:429–446

123


streammicrobial production andmineralization tends to

increase, while the proportions of organic nitrogen,

polysaccharides and amino acids decrease (Ylla et al.

2010; Fellman et al. 2011; Vazquez et al. 2011).

Moreover, stream fragmentation into isolated pools

tends to increase DOC concentrations and DOM

biodegradability, while enhancing the spatial hetero-

geneity of DOM quantity and composition (Fellman

et al. 2011; Vazquez et al. 2011).

Hydrological expansion reestablishes hydrological

connectivity, thereby increasing the concentration of

allochthonous DOC and DON derived from the

leaching of sediments/soils and particulate organic

matter accumulated on the streambed and the hill

slopes during the dry phase (Jacobson et al. 2000;

Inamdar et al. 2011; Catalan et al. 2013). The DOM

pool upon rewetting is thus typically characterized by

high concentrations of labile high molecular weight

substances (Romanı et al. 2006; Vazquez et al. 2007;

Inamdar et al. 2011), with a high proportion of proteins

(Inamdar et al. 2011; Singh et al. 2014) and polysac-

charides (Ylla et al. 2010), and a low proportion of

aromatic and basic amino acids (Ylla et al. 2011). In

general, changes in stream DOM quantity and com-

position following dry periods are more pronounced

than those observed for other events (e.g. storms)

during the rest of the year (Bernal et al. 2005; Vazquez

et al. 2007; Inamdar et al. 2011). The quantity and

composition of DOM during rewetting events in

temporary streams may thus differ significantly from

that measured more routinely during storm-events in

perennial streams (Fellman et al. 2009; Nguyen et al.

2010; Pellerin et al. 2012).

To date, there has been little work on the dynamics

of DOM in temporary streams that consider both DOM

quantity and composition. Some studies have focused

on DOM dynamics upon rewetting at the start of the

expansion phase (e.g. Bernal et al. 2005; Vazquez

et al. 2007; Inamdar et al. 2011; Catalan et al. 2013).

Much less is known on the dynamics of DOM during

hydrological contraction and, especially, during hy-

drological fragmentation (but see e.g. Ylla et al. 2010;

Vazquez et al. 2011: Fellman et al. 2011). Moreover,

because of lack of temporal sampling and/or limited

sampling frequency, most of these studies have missed

the short-term dynamics and the transitions between

hydrological phases (i.e. contraction–fragmentation

and fragmentation–dry), which may represent biogeo-

chemical ‘‘hot moments’’ (sensu McClain et al. 2003).

The objective of this study was to examine the

effect of flow intermittency on the short-term dynam-

ics of stream DOM quantity and composition, with

special emphasis on the transitions between hydro-

logical phases. We combined intensive sampling with

a variety of analytical techniques to follow changes in

DOC and DON concentrations as well as in DOM size

fractions and spectroscopic properties in a temporary

Mediterranean stream during an entire contraction–

fragmentation–expansion hydrological cycle. Our fi-

nal goal was to gain information that may help us

develop better conceptual and mechanistic models of

DOM biogeochemistry in temporary streams, and thus

facilitate the sustainable management of these ecosys-

tems. Furthermore, our results may serve as a template

for understanding and managing the potential biogeo-

chemical response of stream ecosystems under future

water scarcity scenarios.

Methods

Study site

The Fuirosos stream drains a 15.2-km2 granitic

catchment in the Montnegre-Corredor Natural Pro-

tected Area (NE Iberian Peninsula). The climate is

semiarid Mediterranean, with monthly mean air

temperature ranging from 5 �C (January) to 24 �C(August) and a seasonally and inter-annually highly

variable precipitation (annual mean = 750 mm)

(Ninyerola et al. 2000). The catchment is mostly

forested and human land use is restricted to disperse

agricultural fields (\2 % of the catchment area).

Evergreen forests dominate at the lower parts of the

catchment, whereas deciduous forests are common at

higher elevations. Riparian vegetation is well devel-

oped, with a leaf input peak in autumn, although

intense summer hydric stress causes additional leaf

fall during the dry phase (Acuna et al. 2007).

Stream flow is seasonally intermittent, with a dry

phase during summer of variable duration and spatial

extent among years (Vazquez et al. 2013). The

concentration of DOC (annual mean *6 mg C L-1)

peaks during the transition from dry to wet conditions

in autumn, whereas the concentration of DON (annual

mean *0.2 mg N L-1) shows no clear seasonal

pattern (Bernal et al. 2005). At storm flow conditions,

DOC and DON concentrations tend to increase and the

Biogeochemistry (2015) 123:429–446 431

123


highest concentrations occur in autumn (mean

*8 mg C/L and *0.6 mg N/L; Bernal et al. 2005).

DOM composition is also highly responsive to

seasonal hydrological changes in this stream (Bernal

et al. 2005; Romanı et al. 2006; Vazquez et al. 2007;

Ylla et al. 2010; Vazquez et al. 2011).

For this study, we selected a 300-m long reach,

representative of the third-order section of the

Fuirosos stream (UTM coordinates at the center of

the reach = 31T 464934E 4616159 N; eleva-

tion = 123 m a.s.l.). The selected reach had a mod-

erate slope (0.063 m m-1) with alternance of pools

and riffles. Substrate was composed of sand (60 %)

and boulders (30 %), intermixed with patches of

cobbles, pebbles, gravel, and bedrock. The streambed

was mostly covered by biofilms, with an algal

community dominated by diatoms and cyanobacteria

(Tornes and Sabater 2010).

Field sampling

We sampled surfacewater at 3 to 4-day intervals (totally

24 dates) during the contraction, fragmentation and

expansion phases (see Fig. 1 for representative photos

of the different hydrological phases). To account for

spatial heterogeneity, we took the samples from 4

locations along the study reach (80, 160, 240 and 280 m

from the top of the study reach). Surface water was

present at all locations during the contraction phase (8

June–4 July; 8 sampling dates) and the expansion phase

(23 October–19 November; 9 sampling dates), while it

was absent from the location at 80 m on the last two

sampling dates of the fragmentation phase (5 July–23

July; 7 sampling dates). Sampling was carried out

between0900 h and1200 h tominimize the influence of

potential diel changes inDOM(Wilson andXenopoulos

2013).

We measured dissolved oxygen concentration

(DO), pH and water temperature at the mid-channel

area of each sampling location on each date using

WTW (Weilheim, Germany) hand-held probes. At the

same spots, we collected surface water using 100-mL

syringes (flushed three times prior to use), and filtered

it in the field through ashed Albet (Barcelona, Spain)

GF55 glass fiber filters (0.7 lm pore size) into pre-

washed plastic containers (1 sample per location). We

chose glass fiber filters because they are inert, allow

rapid filtration in the field and can be ashed. We stored

Fig. 1 Representative photos of the hydrological phases in the

Fuirosos stream (NE Iberian Peninsula) during the study period

a contraction phase (29 June 2014), b fragmentation phase (6

July 2014), c dry phase (22 August 2014), and expansion phase

(27 October 2014). Photos were taken at the sampling location

180 m


123

personal copy

the samples on ice in the dark, transported them

immediately to the laboratory and froze them at

-20 �C. At the end of the samplings, we transported

all samples frozen to the Leibniz-Institute of Fresh-

water Ecology and Inland Fisheries (Berlin, Germany)

and analyzed them within 1 month. To minimize the

effects of freezing and thawing, all samples were

subject to only one freezing-thawing cycle, and

thawing was conducted in a refrigerator at 4 �C(Fellman et al. 2008; Hudson et al. 2009; Spencer et al.

2010).

We estimated discharge from measurements of

water column depth at the bottom of the study reach at

10-min intervals using a YSI (Ohio, USA) 600-OMS-

V2 multiparameter sonde. We constructed an em-

pirical depth-discharge relationship based on several

additions of a conservative tracer (i.e. chloride) during

the sampling period (Gordon et al. 2004).

Laboratory analyses

Size exclusion chromatography

We measured total DOC and DON concentrations and

characterized the DOM size fractions on a DOC-Labor

Huber (Karlsruhe, Germany) liquid chromatography–

organic carbon–organic nitrogen detection system

(LC-OCD-OND) (Huber et al. 2011; Graeber et al.

2012a). The LC-OCD-OND system was driven by a

Knauer (Berlin, Germany) S-100 HPLC pump and

consisted of a MLE (Dresden, Germany) autosampler

and a YSK HW (Toso, Japan) chromatographic

column. Each sample was measured after passing

and bypassing the chromatographic column with a

Knauer (Berlin, Germany) S-200 UV detector. Sub-

sequently, flow was further divided into two streams.

One stream went to a UV reactor to measure nitrogen

at 220 nm after oxidation to nitrate. The second stream

went to a thin-film reactor where DOCwas oxidized to

CO2 and then to an infrared CO2 detector. The mobile

phase used was a phosphate buffer of pH 6.85 (2.5 g

KH2PO4 ? 1.5 g Na2HPO4 9 2H2O to 1 L).

We calibrated the LC-OCD-OND system according

to the manufacturer’s instructions (DOC-Labor Huber,

Karlsruhe, Germany), and analyzed the DOM size

fractions using the software FIFFIKUS (DOC-Labor

Huber, Karlsruhe, Germany). Following Attermeyer

et al. (2014), we classified the DOM into three

fractions: (i) high molecular weight substances

(HMWS), including polysaccharides, (ii) humic or

humic-like substances (HS), including buildings

blocks, and (iii) low molecular weight substances

(LMWS), which summarize low molecular weight

acids and low molecular weight neutral substances.

We expressed each fraction as the percent contribution

to total DOC and total DON.While DOCwas detected

in all DOM size fractions, DON was only detected in

the HMWS and HS fractions (Graeber et al. 2012a).

Thus, the percent contribution to total DON and the

DOC:DON molar ratios were only reported for these

two fractions. Noteworthy, the LC-OCD-OND system

allowed the direct determination of DON, which is less

prone to errors than indirect DON calculations at high

dissolved inorganic nitrogen concentrations (Graeber

et al. 2012a), such as those typically found during the

end of the fragmentation phase and the start of the

expansion phase (von Schiller et al. 2011).

Spectroscopic measurements

We determined the specific UV absorbance (SUVA; L

mg-1m-1), a surrogate forDOMaromaticity (Weishaar

et al. 2003), directly on the LC-OCD-OND system

(Huber et al. 2011). To determine other spectroscopic

indices, we conducted absorbance measurements on a

Shimadzu (Kyoto, Japan) UV-2401 UV/VIS spec-

trophotometer and fluorescence measurements on a

Perkin-Elmer (Waltham, MA, USA) LS-50b fluores-

cence spectrometer. We measured excitation from 240

to 450 nm (5 nm steps) and emission from 300 to

600 nm (2 nm steps) with a slit width of 5 nm to

produce excitation-emission-matrices (EEMs; Stedmon

andMarkager 2005).Wemeasured the samples at room

temperature and corrected the absorbance spectra

(800–190 nm) for instrument baseline offset (Green

and Blough 1994). Daily measurements of the area

under the Raman peak for MilliQ water (Millipore,

Schwalbach, Germany) indicated instrument stability

(Lawaetz and Stedmon 2009). Fluorescence and ab-

sorbance readings were within the linear range of the

spectrometers and we accounted for primary and

secondary inner-filter effects by inner-filter correction

(Lakowicz 2006). Moreover, we corrected the spectra

for excitation by using the correction provided by the

manufacturer and for emission by using the BAM

fluorescence calibration kit (Pfeifer et al. 2006), and

normalized them by the area under the Raman peak at

350 nm excitation wavelength (Lawaetz and Stedmon


123


2009). These corrections provide the best possible

comparability to other DOM fluorescence studies

(Lawaetz and Stedmon 2009).

From absorbance spectra we calculated 4 indices: a)

the ratio of absorptions at 250 and 365 nm (E2:E3),

with values ranging *4–8 (De Haan and De Boer

1987), (b) the spectral slope for 275–295 nm

(S275–295), with values ranging *0.01–0.05 (Helms

et al. 2008) (c) the spectral slope for 350–400 nm

(S350–400), with a similar range of values as S275–295(Helms et al. 2008), and d) the ratio of these slopes

(SR), with values ranging*0.5–3 (Helms et al. 2008).

The indices E2:E3, S275–295, S350–400 and SR have been

found to be inversely related to DOM molecular size;

thus, the higher the index value, the lower the

molecular size (De Haan and De Boer 1987; Peu-

ravuori and Pihlaja 1997; Helms et al. 2008). In

addition, S275–295 can be used as a tracer of photo-

bleaching (Helms et al. 2008) and of terrestrial DOC

(Fichot and Benner 2012).

FromEEMswecalculated 3 indices: (a) humification

index (HIX), forwhich values*1–2 are associatedwith

non-humified material and values[10 are typical for

fulvic acid extracts (Zsolnay et al. 1999), (b) fluores-

cence index (FI), for which values *1.3 suggest the

dominance of terrestrial higher-plant DOM sources and

values*1.8 suggest the dominance of microbial DOM

sources (McKnight et al. 2001), and (c) freshness index

(b:a), for which values[1 correspond to DOM freshly

released into water, whereas values *0.6–0.7 corre-

spond to lower DOM production in natural waters and

higher input of terrestrial origin (Parlanti et al. 2000;

Wilson and Xenopoulos 2009).

To further characterize changes in DOM fluores-

cence, we created a parallel factor analysis (PAR-

AFAC) model with the EEMs using the DOM Fluor

Toolbox (version 1.7; Stedmon and Bro 2008) for

Matlab (version R2009b, MathWorks, Ismaning,

Germany). We identified and split-half validated four

PARAFAC components (hereafter referred to as C1–

C4; Fig. 2, Table 1) and established the best model fit

by random initialization (Stedmon and Bro 2008). We

expressed DOM fluorescence of the PARAFAC

components as percent contribution of each compo-

nent to the sum of fluorescence by all components.

Data analysis

We examined differences among hydrological phases

for the measured physical, chemical and DOM quantity

Fig. 2 Excitation-emission

fluorescence spectra of the

four identified PARAFAC

components


123


and composition variables using non-parametric Krus-

kal–Wallis tests.Wegrouped themedianvalues for each

date by hydrological phase (i.e. contraction phase = 8

sampling dates, fragmentation phase = 7 sampling

dates, and expansion phase = 9 sampling dates). If the

testwas significant,we performed post hoc comparisons

of mean ranks of all pairs of groups using two-sided

significance levels with a Bonferroni adjustment. To

examine the temporal patterns of themeasured variables

under the different hydrological conditions,weexplored

the relationship between the median of these variables

along the reach and the sampling date using non-

parametric Spearman-rank correlations for each hydro-

logical phase separately. We ran all statistical analyses

with Statistica 6.1 (Statsoft, Tulsa, OK, USA). We

considered the statistical results significant if p\ 0.05.

Results

Physical and chemical variables

Stream discharge decreased during hydrological contrac-

tion (rs = -0.95, p\0.001; Fig. 3a). The subsequent

fragmentation phase was only interrupted by a short flow

pulse (10 July) that reconnected the isolated pools for

3 days. The study reach remained dry for 91 days until

stream flow reconnection occurred with a moderate

increase in discharge at the beginning of the expansion

phase (23 October). Subsequently, discharge decreased

until the endof the study (rs = -0.85, p\0.001; Fig. 3a).

Temperature, conductivity,DOandpHdifferedamong

hydrological phases (H[11.40, p\0.05; Fig. 3b, c).

Temperature and conductivity were lower and higher,

respectively, during hydrological expansion (post hoc,

p\0.01) while DO and pH were lower during the

fragmentation phase than during the other hydrological

phases (post hoc, p\0.01). Conductivity showed a clear

increase during hydrological contraction (rs = ? 0.93,

p\0.001), which continued during hydrological frag-

mentation (rs = ? 0.89, p = 0.007; Fig. 3b). Conversely,

temperature, DO and pH showed no clear temporal

patterns during these hydrological phases (p[0.05,

Fig. 3b, c). After flow reconnection, conductivity de-

creased (rs = -0.82, p = 0.007; Fig. 3b), while the rest

of physicochemical variables did not show a clear

temporal pattern (p[0.05; Fig. 3b, c).

Total DOM

Total DOC and DON concentrations did not differ

among hydrological phases (p[ 0.05; Fig. 4a). Total

DOC and DON concentrations remained low

(* 3 mg C L-1 and *0.1 mg N L-1, respectively)

Table 1 Emission (Em. max) and excitation (Ex. max) maxima, previous identifications and literature-based tentative interpretation

of the parallel factor analysis (PARAFAC) components

Component Em.

max.

(nm)

Ex. max. (nm) Similar components

identified in previous

studies

Tentative interpretation of components

C1 416 \240 (315) C1c, C3d, C2e Humic-acid fluorophore; aromatic, terrestrial originc; found in

marine DOM; positively related to forage crope, arable

farming and wetlands in small catchmentsd

C2 490 \240 (400) C2d Fulvic acid fluorophored; highly conjugated, terrestrial origind

C3 450 370 (255,\240) SQ2a, C4c, C4d Semi-quinone/fulvic acid fluorophorea,d; terrestrial origind,

microbially transformeda, high aromaticity, reduced statea,

positively related to arable farming in small catchmentsc

C4 340 \240 (275) C8a, C8b, C5c, C7d,

C5eTryptophan-like fluorophorea, b, d; probably derived from

aquatic microbial productiond, high bioavailability, not

humifiedb, positively related to forest in small catchmentsc

The values in the Ex. max column indicate primary and secondary (with brackets) peaksa Cory and McKnight (2005)b Fellman et al. (2009)c Graeber et al. (2012b)d Stedmon and Markager (2005)e Williams et al. (2010)


123


during hydrological contraction and fragmentation

(p[ 0.05; Fig. 4a). Noteworthy, some sampling

locations exhibited a sharp increase in total DOC

and DON concentrations just before surface drying. At

the beginning of the expansion phase, the total DOC

and DON concentrations reached the highest values

(*10 mg C L-1 and *0.3 mg N L-1, respectively)

and then decreased to background values (rs = -0.97,

p\ 0.001 and rs = -0.90, p\ 0.001, respectively;

Fig. 4a). The median total DOC:DON molar ratio

(range = 23–39), did not differ among hydrological

phases (p[ 0.05) and lacked a distinct pattern during

the entire study period (p[ 0.05; Fig. 4b).

DOM size fractions

The largest part of DOC was in the form of HS

(median = 80.6 %, quartiles = 78.2–82.1 %), followed

by LMWS (16.7, 15.5–19.2 %) and HMWS (2.2,

1.5–3 %). These proportions did not differ among

hydrological phases (p[0.05; Fig. 5a). Yet, the propor-

tion of DOC in HMWS slightly decreased (from 3.6 to

1.9 %) during hydrological contraction (rs = -0.98,

p\0.001), while the proportion of DOC in HS and

LMWS showed no significant temporal pattern

(p[0.05; Fig. 5a). There were no significant temporal

patterns in the proportion of the different fractions of

DOCduring the fragmentationphase (p[0.05; Fig. 5a).

However, some sampling locations exhibited a sharp

increase in the proportion of DOC in HS and a sharp

increase in the proportion ofDOC inLMWSandHMWS

prior to complete surface drying (Fig. 5a). After flow

reconnection, the proportion of DOC in HMWS sharply

increased and then decreased (from 6.1 to 1.7 %; rs =

-0.83, p = 0.005), while the proportion of DOC in HS

and LMWS remained stable until the end of the study

(p[0.05; Fig. 5a).

The largest part of DON was also in the form of HS

(median = 87.9 %, quartiles = 83.0–91.4 %) and to

(a)

(b)

(c)

Fig. 3 Temporal variation

of physical and chemical

variables: a daily discharge,

b temperature and

conductivity, and

c dissolved oxygen and pH.

Different shadings separate

the hydrological phases:

contraction (8 June–4 July),

fragmentation (5 July–23

July), and expansion (23

October–19 November).

The break in the x-axis

indicates the dry (no surface

water) phase. Values are the

median and range for each

sampling date


123


a lesser extent in the form of HMWS (12.1,

8.6–17.0 %). The proportion of DON in HS and

HMWS did not differ among hydrological phases

(p[ 0.05) and did not show any clear temporal pattern

during any of the investigated hydrological phases

(p[ 0.05). Noteworthy, in all sampling locations

there was a decrease in the proportion of DON in HS

and an increase in the proportion of DON in HMWS

before complete surface drying (Fig. 5b).

The DOC:DON ratio in HS was similar to the

DOC:DON ratio of total DOM and higher than the

DOC:DON ratio in HMWS (Fig. 5c). The DOC:DON

ratios in HS and HMWS differed among hydrological

phases (H = 9.11, p = 0.011 and H = 9.07,

p = 0.011, respectively). For HS, the DOC:DON

ratio was lower during the expansion phase than

during the other hydrological phases (post hoc,

p\ 0.037). For HMWS, the DOC:DON ratio was

higher during the expansion phase than during the

fragmentation phase (post hoc, p = 0.009), but there

were no differences between the contraction phase and

the other hydrological phases (post hoc, p[ 0.05).The

DOC:DON ratio in HS did not show any clear

temporal pattern throughout the study period

(p[ 0.05; Fig. 5c). Similarly, the DOC:DON ratio

in HMWS did not show any clear temporal pattern

during hydrological contraction and fragmentation

(p[ 0.05); however, it sharply increased and then

gradually decreased during hydrological expansion

(rs = -0.80, p = 0.010; Fig. 5c).

DOM spectroscopic properties

Most spectroscopic indices differed among hydro-

logical phases (H[ 7.90, p\ 0.020; Fig. 6). SUVA

was higher and SR, FI and b:a were lower during the

expansion phase than during the other hydrological

phases (post hoc, p\ 0.022, p\ 0.007, p\ 0.033,

p\ 0.042, respectively; Fig. 6). S275–295 was lower

during the expansion phase than during the fragmen-

tation phase (post hoc, p = 0.019), but there were no

differences between the contraction phase and the

other hydrological phases (post hoc, p[ 0.05;

Fig. 6b). HIX was higher during the expansion phase

than during the contraction phase (post hoc,

p = 0.019), but there were no differences between

the fragmentation phase and the other hydrological

phases (post hoc, p[ 0.05; Fig. 6d). During hydro-

logical contraction, E2:E3, S275–295 and S350–400increased (rs = ?0.76, p = 0.028; rs = ?0.88,

p = 0.004; and rs = ?0.76, p = 0.028, respectively;

Fig. 6 a, b). All other spectroscopic indices showed no

clear temporal pattern during this hydrological phase

(p[ 0.05; Fig. 6). Among all spectroscopic indices,

only SUVA decreased during hydrological fragmen-

tation (rs = -0.93, p = 0.003; Fig. 6a). Yet, abrupt

(a)

(b)


of total dissolved organic

matter (DOM)

concentrations: a Dissolved

organic carbon (DOC) and

dissolved organic nitrogen

(DON), b DOC:DON molar

ratio. Different shadings

separate the hydrological

phases (see Fig. 3 for

details). Values are the


sampling date


123


changes for most spectroscopic indices occurred on

the last sampling date before complete surface drying.

After flow reconnection, E2:E3, S275–295 and S350–400were lowest and then increased until the end of the

study (rs = ?0.93, p\ 0.001; rs = ?0.90,

p\ 0.001; and rs = ?0.83, p = 0.005, respectively;

Fig. 6a, b, c). In contrast, SUVA and FI were highest

just after flow reconnection, and then decreased

(rs = -0.80, p = 0.010 and rs = -0.87, p = 0.002,

respectively; Fig. 6a). The rest of spectroscopic vari-

ables showed no consistent temporal patterns during

hydrological expansion (p[ 0.05; Fig. 6).

All PARAFAC components except C1 showed

differences among hydrological phases (H[ 9.42,

p\ 0.010; Fig. 7). The components C2 and C4 showed

consistently higher and lower values, respectively,

during the expansion phase than during the contraction

and fragmentation phases (post hoc, p\ 0.014 and

(a)

(b)

(c)

Fig. 5 Temporal variation of dissolved organic matter (DOM)

size fractions: a the percent contribution to total dissolved

organic carbon (DOC), b the percent contribution to total

dissolved organic nitrogen (DON), and c the DOC:DON molar

ratio of each fraction. HMWS = high molecular weight

substances, including polysaccharides. HS = humic or humic-

like substances, including buildings blocks. LMWS = low

molecular weight substances, which summarize low molecular

weight acids and low molecular weight neutral substances.

Different shadings separate the hydrological phases (see Fig. 3

for details). Values are the median and range for each sampling

date


123


p\ 0.009, respectively; Fig. 7). The component C3

showed higher values during the expansion phase than

during the fragmentation phase (post hoc, p = 0.006),

but values did not differ between the contraction phase

and the other hydrological phases (post hoc, p[ 0.05;

Fig. 7). The component C3 decreased during hydro-

logical contraction (rs = -0.83, p = 0.010) while the

rest of PARAFAC components remained relatively

stable (p[ 0.05; Fig. 7). None of the PARAFAC

components showed a clear temporal pattern during

hydrological fragmentation and expansion (p[ 0.05;

Fig. 7). Noteworthy, abrupt changes in all PARAFAC

components occurred just before complete surface

drying.

Discussion

Flow intermittency was a key determinant of DOM

biogeochemistry in the study stream. There were

notable differences in DOM quantity and composition

and their dynamics among the different hydrological

phases, with the most relevant changes occurring at

the transitions between the individual phases.

(a)

(b)

(c)

(d)


of spectroscopic indices:

a Specific UV absorbance

(SUVA) and the ratio of

absorption at 250–365 nm

(E2:E3), b spectral slope

from 275 to 295 nm

(S275–295) and from 350 to

400 nm (S350–400), c spectralslope ratio (SR) and

freshness index (b:a), andd fluorescence index (FI)

and humification index

(HIX). Different shadings

separate the hydrological

phases (see Fig. 3 for

details). Values are the


sampling date


123


Hydrological contraction

During hydrological contraction the stream gradually

disconnected from its valley as well as from upstream

and downstream reaches, following a characteristic

pattern described for Mediterranean streams (Bernal

et al. 2013). Nonetheless, most physicochemical vari-

ables (i.e. temperature, DO and pH) remained fairly

stable, with only conductivity showing a clear increase

with the reduction of discharge during hydrological

contraction (Fig. 3). As a result, both total DOC and

DON concentrations remained stable, resulting in a

rather steady total DOC:DON ratio (Fig. 4), and

suggesting that the source of DOM was constant and/

or that DOM was not greatly affected by in-stream

processes during hydrological contraction (Dahm et al.

2003, Bernal et al. 2005). Our results agree with those

from previous studies in the same stream that have also

reported stable DOC and DON concentrations during

hydrological contraction (Ylla et al. 2010; von Schiller

et al. 2011). However, they contradict results from other

streams, where decreases in DOC and DON concentra-

tion due to hydrological disconnection under drought

conditions have been reported (e.g. Freeman et al. 1994;

Dahm et al. 2003). Differences between our stream and

the other streams may be attributed to the higher

heterotrophic nature of our study stream (forested)

compared to the other streams (wetland and desert).

Data from more streams during hydrological contrac-

tion or drought are needed to verify this hypothesis.

Despite there were no significant changes in DOM

quantity, some relevant changes in DOM composition

occurred during hydrological contraction.Overall, there

was a decrease in the average molecular weight of

DOM, as indicated by the decrease in the proportion of

HMWS (Fig. 5) and the concomitant increase in E2:E3,

S275–295 and S350–400 (Fig. 6) (De Haan and De Boer

1987; Peuravuori and Pihlaja 1997; Helms et al. 2008).

Thus, despite of its low contribution to the total DOC

concentration, our results suggest that the HMWS

fraction was the most biogeochemically reactive. The

high reactivity of the HMWS fraction was most likely

due to the fact that it exhibited a low DOC:DON ratio,

and was dominated by polysaccharides with some

contribution from nitrogen-containing material such as

proteins or amino sugars (Huber et al. 2011). Although

at coarser temporal scales, previous studies have also

shown a decrease in the proportion of polysaccharides in

total DOC during hydrological contraction (Ylla et al.

2010). TheHMWSfractionmay thus play a key role as a

bioavailable source of DOM in stream ecosystems

subject to flow intermittency, while the other DOM size

fractions have a more recalcitrant behavior. Nonethe-

less, we must acknowledge that we did not specifically

measure bioavailability of DOM in our samples.

Moreover, the characterization of DOMmolecular size

and its relationship with DOM reactivity is an unre-

solved topic in aquatic biogeochemistry, with often

contradictory results (e.g. Lindell et al. 1995; Amon and

Benner 1996; Sachse et al. 2001; Agren et al. 2008).


of the percent contribution

of the 4 identified parallel

factor analysis (PARAFAC)

components (C1-C4).

Different shadings separate

the hydrological phases (see

Fig. 3 for details). Values

are the median and range for

each sampling date


123


Hydrological fragmentation

In the hydrological transition between the contraction

and fragmentation phases, stream flowwas interrupted

and surface water got restricted to isolated pools

(Bernal et al. 2013). As a result, there was an abrupt

but predictable increase in acidic and low oxygen in-

stream conditions which remained for the rest of the

fragmentation phase (Boulton and Lake 1990; von

Schiller et al. 2011). Conductivity continued to

increase during the fragmentation phase, likely driven

by an increase in solute concentration through

evaporation (Sabater and Tockner 2010). Tem-

perature, however, remained rather constant probably

regulated by the shading effect of the well-developed

riparian canopy (Johnson 2004).

The significant shift in physicochemical conditions

between the contraction and the fragmentation phase

was not followed by relevant changes in DOM

quantity or composition. Total DOC and DON con-

centrations (Fig. 4), DOM size fractions (Fig. 5) and

spectroscopic properties (Fig. 6 and 7) remained

relatively constant with respect to the contraction

phase. Furthermore, among all DOM composition

variables, only the SUVA index showed a clear

temporal pattern during hydrological fragmentation

(Fig. 6). The negative relationship between SUVA

and sampling time indicated that stream DOM became

less aromatic with time since pool isolation (Weishaar

et al. 2003). These short-term temporal data support

the finding by Vazquez et al. (2011) in the same

stream, who found decreasing SUVA across a spatial

gradient of pools of increasing age. Vazquez et al.

(2011) attributed the decrease in SUVA to an increase

in the contribution of in-stream algal and microbial

processes to the total DOM pool with increasing pool

isolation time. We did not find correlations between

pool isolation time and any spectroscopic property

other than SUVA. Nonetheless, similar to Fellman

et al. (2011), who found higher protein-like fluores-

cence in highly evaporated pools in a subtropical

dryland river, we also found an abrupt increase in

protein-like fluorescence at the end of the fragmenta-

tion phase (see below).

Abrupt changes in DOM quantity and composition

occurred in most sampling locations at the end of the

fragmentation phase, just before the stream complete-

ly dried up at the surface. Both total DOC and DON

concentrations sharply increased (Fig. 4), while the

proportion of HS decreased and the proportion of

HMWS and LMWS increased (Fig. 5). These changes

indicate a major shift in the bulk composition of DOM

towards a higher proportion of non-humic substances

such as polysaccharides and low molecular weight

alcohols, aldehydes, ketones, sugars and amino acids

(Huber et al. 2011). In addition, the spectroscopic

indices indicated that the released DOM had a fresh

(high b:a; Wilson and Xenopoulos 2009), microbial

(high FI, McKnight et al. 2001), and non-humic (low

HIX; Zsolnay et al. 1999) character (Fig. 6). Further-

more, changes in the proportions of the PARAFAC

components (i.e. low C1 and high C4; Fig. 7) indicat-

ed that the DOM shifted from a more humic-acid like,

aromatic and terrestrial character to a more trypto-

phan-like, non-humified, bioavailable and aquatic

character (Stedmon and Markager 2005; Williams

et al. 2010; Graeber et al. 2012b). Together, these

results support the idea that the increase in DOC and

DON concentrations before complete surface drying

was most likely due to an abrupt microbial biofilm cell

lysis and/or DOM exudation under stress conditions

(e.g. high temperature, low DO, low pH), which could

not be mineralized (Humphries and Baldwin 2003;

Schimel et al. 2007; Timoner et al. 2012). This

biogeochemical hot moment (sensu McClain et al.

2003) at the end of the fragmentation phase has not

been captured by previous studies of DOM dynamics

during stream drying (e.g. Ylla et al. 2010; Fellman

et al. 2011; Vazquez et al. 2011), probably because

they covered coarser temporal scales than the present

study. Our results support the idea that hydrological

fragmentation, especially in its later stages, can

strongly enhance changes in stream DOM quantity

and composition, thereby increasing the spatial

heterogeneity of DOM and nutrient availability along

temporary stream networks (Dent and Grimm 1999;

Gomez et al. 2009; von Schiller et al. 2011; Vazquez

et al. 2011; Fellman et al. 2011).

Hydrological expansion

Upon rewetting, the lateral and longitudinal hydro-

logical connections along the stream channel were re-

established (Bernal et al. 2013). Flow reconnection

favored the release of high amounts of total DOC and

DON (Fig. 4), in line with previous studies in the same

stream (Bernal et al. 2005; Vazquez et al. 2007; Ylla

et al. 2010) and temporary streams elsewhere


123


(Inamdar et al. 2011; Catalan et al. 2013). These

results are also consistent with other studies performed

in a wide range of perennial streams during storm

events (Fellman et al. 2009; Nguyen et al. 2010;

Pellerin et al. 2012). Yet, we must acknowledge that

even if we sampled the stream on the first day upon

rewetting, we did not capture the rising limb of the

hydrograph; thus, our samples may not be fully

representative of the DOM that gets released just after

flow reconnection (Nguyen et al. 2010; Inamdar et al.

2011). Nonetheless, the rewetting showed a relatively

low peak discharge with DOC values similar to those

reported in other investigated post-drought rewetting

events in the same stream (Bernal et al. 2005; Romanı

et al. 2006; Vazquez et al. 2007).

In parallel to the increase in DOC and DON

concentrations upon rewetting, there was an increase

in the proportion of the HMWS fraction of DOC and

DON (Fig. 5), supported by the observation of low

E2:E3 and S275–295 values (Fig. 6), indicative of high

average molecular weight (De Haan and De Boer

1987; Helms et al. 2008). This observation is in line

with previous studies in the same stream that have

reported an increase in the proportion of HMWS after

flow reconnection (Romanı et al. 2006; Vazquez et al.

2007), most likely associated with an increase in the

relative amount of polysaccharides at rewetting (Ylla

et al. 2010). Other studies using DOM spectroscopy

have reported similar increases in the proportion of

HMWS at rewetting for a temporary stream in North

America (Inamdar et al. 2011) and for perennial

streams from several bioclimatic regions during storm

events (Nguyen et al. 2010; Spencer et al. 2010).

As indicated by other spectroscopic indices

(Fig. 6), DOM just upon rewetting was also charac-

terized by high aromaticity (high SUVA; Weishaar

et al. 2003), terrestrial origin (low S275–295; Fichot and

Benner 2012) and old source (low b:a; Wilson and

Xenopoulos 2009). Furthermore, the higher and lower

values of the PARAFAC components C2 and C4,

respectively, indicated the preponderance of a more

humic-like DOM of terrestrial origin (Fig. 7; Table 1;

Cory and McKnight 2005; Stedmon and Markager

2005). Previous studies of storm-events during post-

drought periods in temporary streams (Catalan et al.

2013) and in perennial streams (Fellman et al. 2009;

Nguyen et al. 2010) have also reported increases of

aromatic and humic DOM of terrestrial origin,

supported primarily by spectroscopic data. In contrast,

an increase in protein-like fluorescence during the

first-flush event after drought was found in a tempo-

rary stream in North America (Inamdar et al. 2011).

The authors attributed this increase to the breakdown

and production of labile organic matter in the dry

stream sediments and riparian soils during drought

(Inamdar et al. 2011). In absence of a definitive

explanation, we hypothesize that the high accumula-

tion of leaves on the streambed and riparian soils of the

Fuirosos stream in summer due to hydric stress (Acuna

et al. 2007), which was most likely not as pronounced

in the North American stream (Inamdar et al. 2011),

may explain the observed difference between the two

streams. Therefore, the timing of flow reconnection

with respect to leaf fall may be an important factor in

determining the composition of the first-flush DOM in

temporary streams.

Overall, DOM composition results confirm previ-

ous observations in the same stream that attribute the

typical increase in DOC and DON concentrations

upon rewetting to leaching of accumulated detritus on

the streambed and hill slopes (Romanı et al. 2006;

Vazquez et al. 2007; Artigas et al. 2009). Nonetheless,

the leaching of dry sediments and near-stream soils

could represent an additional DOM source upon

rewetting (Inamdar et al. 2011). Interestingly, the

plant- and sediment/soil-derived DOM observed at the

beginning of the expansion phase is probably highly

labile (McDowell 1985; McArthur and Richardson

2002) and can be rapidly used by stream microorgan-

isms that show a rapid recovery of their activity after

rewetting (Romanı et al. 2006; Artigas et al. 2009;

Timoner et al. 2012).

After the initial increase at the beginning of the

expansion phase, the total DOC and DON concentra-

tions concurrently decreased and the total DOC:DON

molar ratio remained constant (Fig. 4), indicating no

major changes in the bulk DOM source (Bernal et al.

2005). Interestingly, this temporal pattern for DOC

and DON concentrations was similar to that observed

by Inamdar et al. (2011) but unlike previous observa-

tions in the same stream (Bernal et al. 2005; Ylla et al.

2010), where it was reported that DOC concentrations

followed a different pattern than DON concentrations

during hydrological expansion. We do not have a

definitive explanation for these differences in DOC

and DON patterns during hydrological expansion;

however, we suspect that it may be due to the

technique used to measure DON concentration. We


123


determined the DON concentration directly with the

LC-OCD-OND system, which is more accurate than

the indirect DON determination technique used in

previous studies for measuring DON concentration

(Graeber et al. 2012a), especially at high dissolved

inorganic nitrogen concentrations such as those found

upon rewetting (Von Schiller et al. 2011).

In parallel to the decrease in total DOC and DON

concentrations, relevant changes in DOM composition

occurred during the expansion phase. Overall, there was

a decrease in the average molecular weight of DOM, as

indicated by the decrease in the proportion of HMWS

(Fig. 5) and the concomitant increase in E2:E3, S275–295and S350–400 (Fig. 6) (De Haan and De Boer 1987;

Peuravuori and Pihlaja 1997; Helms et al. 2008).

Simultaneously, there was a slight decrease of the

DOC:DONratio inHMWS(Fig. 5). These observations

support again the biogeochemical relevance of the

HMWS fraction, despite of its low contribution to the

total DOC concentration. The rest of DOMcomposition

variables did not show clear temporal patterns during

hydrological expansion, except for the decrease of

SUVA values (Fig. 6). This gradual reduction in stream

DOM aromaticity during hydrological expansion

(Weishaar et al. 2003)maybe at least partially explained

by a decrease in DOM leaching from the streambed and

hill slopes, but also by higher microbial processing of

DOM by recovering stream biofilms favored by

continuous flow conditions (Ylla et al. 2010; Timoner

et al. 2012). Interestingly, while some DOM composi-

tion variables recovered to values similar to those at the

contraction phase, other variables remained consistently

higher (e.g. SUVA,HIX,C2) or lower (e.g. SR, FI,HIX,

C4), indicating the prevalence of a humic-like DOM of

terrestrial origin during the whole expansion phase.

Conclusions

By applying an intensive sampling design, in combi-

nation with chromatographic and spectroscopic tech-

niques, this study shows how changes in hydrological

connectivity driven by seasonal stream flow intermit-

tency affect the dynamics of DOM quantity and

composition in a temporary Mediterranean stream.

The most relevant changes occurred at the transitions

between hydrological phases, thus highlighting the

importance of hydrological transitions for DOM

specifically, and biogeochemical processes generally,

in temporary streams. Despite the timely- and spatial-

ly-restricted nature of this study, our results suggest

that flow intermittency is a key determinant of DOM

dynamics in temporary streams, with relevant impli-

cations for the sustainable management and regulation

of these ecosystems. Changes in the quantity and

composition of DOM in surface waters have important

consequences for water quality, andmay thus affect the

ecological integrity of stream ecosystems and thewater

supply for human use. Results from this study will help

us develop better conceptual and mechanistic models

of DOM biogeochemistry in temporary streams, and

thus facilitate the sustainable management of these

ecosystems. Because DOM plays an essential role in

aquatic ecosystem biogeochemistry, while many per-

manent streams are increasingly becoming temporary;

our results may serve as a template for understanding

and managing the potential biogeochemical response

of stream ecosystems under future water scarcity

scenarios. Further research on the effects of flow

intermittency on DOM dynamics across different

geographical, hydrological and ecological settings is

needed to confirm the generality of our findings.

Acknowledgments We thank J. Rodrıguez, L. Proia, M.

Peipoch and A. Blesa for field assistance, and E. Zwirnmann, H.

J. Exner, A. Luder, S. Schell and H. Magnussen for laboratory

analyses. We are also grateful to the direction of the Montnegre-

Corredor Natural Park (Diputacio de Barcelona) for allowing

access to the sampling site. This studywas fundedby theEuropean

Union through the MIRAGE project (FP7 ENV 2007 1).

Additional funds were provided by the Spanish Ministry of

Economy and Competitiveness through the Consolider-Ingenio

projects SCARCE (CSD2009-00065) and GRACCIE (CSD2007-

00067). D. von Schiller was supported by a DAAD-‘‘laCaixa’’

fellowship and a ‘‘Juan de la Cierva’’ postdoctoral grant (JCI-

2010-06397).

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Hydrological transitions drive dissolved organic matter quantity and composition in a temporary Mediterranean stream

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