Episodic rewetting enhances carbon and nitrogen

release from chaparral soils

Amy E. Millera,*, Joshua P. Schimelb, Thomas Meixnerc, James O. Sickmand, John M. Melacka,b

aInstitute for Computational Earth System Science, University of California, Santa Barbara, CA 93106, USAbDepartment of Ecology, Evolution, and Marine Biology, University of California, Santa Barbara, CA 93106, USA

cDepartment of Hydrology and Water Resources, University of Arizona, Tucson, AZ 85721, USAdDepartment of Soil and Water Science, University of Florida, Gainesville, FL 32611-0510, USA

Received 7 April 2004; received in revised form 23 November 2004; accepted 31 March 2005

Abstract

The short-term pulse of carbon (C) and nitrogen (N) mineralization that accompanies the wetting of dry soils may dominate annual C and

N production in many arid and semi-arid environments characterized by seasonal transitions. We used a laboratory incubation to evaluate the

impact of short-term fluctuations in soil moisture on long-term carbon and nitrogen dynamics, and the degree to which rewetting enhances C

and N release. Following repeated drying and rewetting of chaparral soils, cumulative CO2 release in rewet soils was 2.2–3.7 times greater

than from soils maintained at equivalent mean soil moisture and represented 12–18% of the total soil C pool. Rewetting frequency did not

affect cumulative CO2 release but did enhance N turnover, and net N mineralization and nitrification increased with rewetting in spite of

significant reductions in nitrification potential. Litter addition decreased inorganic N release but enhanced dissolved organic nitrogen (DON)

and dissolved organic carbon (DOC) from dry soils, indicating the potential importance of a litter-derived pulse to short-term nutrient

dynamics.

q 2005 Elsevier Ltd. All rights reserved.

Keywords: Soil respiration; Litter addition; Nitrification; Nitrate; DON; DOC

1. Introduction

The rapid rewetting of dry soils is known to enhance soil

carbon (C) and nitrogen (N) mineralization, accelerating

organic matter decomposition (Sorensen, 1974; Kieft et al.,

1987; Van Gestel et al., 1993; Clein and Schimel, 1994) and

resulting in a short-term pulse of available C and N (Birch,

1958). The CO2 pulse that follows rewetting is thought to

derive from the mineralization of soil organic matter (SOM)

released via physical disruption of soil aggregates (Utomo

and Dexter, 1982; Appel, 1998; Denef et al., 2001), and/or

from the release of microbial C (Csonka and Hanson, 1991;

Halverson et al., 2000), while dissolved organic carbon

(DOC) release is thought to derive primarily from SOM C

0038-0717/$ - see front matter q 2005 Elsevier Ltd. All rights reserved.

doi:10.1016/j.soilbio.2005.03.021

* Corresponding author. Present address: National Park Service, 240

West 5th Avenue, Anchorage, AK 99501, USA. Tel.: C1 907 644 3683;

fax: C1 907 644 3809.

E-mail address: amy_e_miller@nps.gov (A.E. Miller).

(Fierer and Schimel, 2003). The complex relationship

between C availability and DOC production (Park et al.,

2002), the extent to which DOC released from labile pools

becomes available for uptake and re-immobilization

following soil rewetting (Degens and Sparling, 1995;

Lundquist et al., 1999; Magid et al., 1999), and the capacity

of the microbial community to utilize this released carbon

(McLean and Huhta, 2000; Fierer et al., 2003a,b;

LaMontgne et al., 2003), remain poorly understood.

Since surface soils often experience large seasonal

fluctuations in moisture content, especially in arid, semi-

arid, or Mediterranean-type environments, the mineraliz-

ation pulse associated with soil drying–rewetting has the

potential to significantly impact total annual C and N release

(Smith and Parsons, 1985; Groffman and Tiedje, 1988;

Mummey et al., 1997; Borken et al., 2003; Fierer and

Schimel, 2003). The rewetting events that characterize the

seasonal transition from summer drought to winter rains can

range in frequency from a few days to 1–2 months in the

foothills of the southern Sierra Nevada, CA, where early

winter flushing events result in disproportionately high

Soil Biology & Biochemistry 37 (2005) 2195–2204

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A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–22042196

stream NOK3 concentrations (Fenn et al., 2003). While such

variation in storm frequency is typical, we know little about

the cumulative effects of multiple rewetting events on soil C

and N dynamics, and how variation in rewetting frequency

may influence overall ecosystem nutrient budgets.

We conducted a series of controlled experiments on soils

from a Mediterranean-type, chaparral environment in south-

central California to examine the effect of rewetting

frequency and litter addition on C and N cycling. Using a

laboratory incubation, we addressed the following ques-

tions: How do short-term pulse dynamics impact long-term

carbon and nitrogen release from soils? How does the

frequency of rewetting events impact C and N mineraliz-

ation over time? Does C availability, as determined by litter

addition, interact with rewetting frequency to control C and

N release?

2. Methods

2.1. Soils

The soils used in this experiment were Ultic Haploxeralfs

(Huntington and Akeson, 1987) collected within the Elk

Creek Watershed of Sequoia National Park (36830 042 00N,

118842 023 00E; elevation 650 m), in late September, 2002.

The site is characterized by a Mediterranean climate, with

cool, wet winters and hot, dry summers. Mean maximum

and minimum temperatures at Ash Mountain, approxi-

mately 3 km from the study site, are 36.4 and 2.2 8C. Mean

annual precipitation is 670 mm, with the majority of rainfall

occurring between November and April. A mature stand of

Chamise (Adenostoma fasciculatum L.) chaparral comprises

the overstory vegetation. The soil used in this experiment

was a sandy clay loam with a pH of 6.2, SOM concentration

of 6.6%, and soil C and N concentrations of 2.3 and 0.1%,

respectively.

2.2. Rewetting experiment

2.2.1. Experimental design

We used a factorial design of soil moisture!litter

addition to determine whether episodic rewetting alters the

timing and magnitude of C and N release, relative to soils

maintained at constant moisture. Surface soils (0–10 cm)

were sieved to 4 mm and stored at 20 8C and field moisture

(4% of water-holding capacity (WHC)) until the initiation of

the rewetting treatments, approximately 90 d later. We

determined 100% WHC from the gravimetric water content

of soil that was saturated and then allowed to drain over 6 h.

Using 10 g dry weight soil incubated in 50 mL centrifuge

tubes, we established the following experiments: (1) a

rewetting experiment for determination of soil C and N

pools over a 16-week incubation period (T0, T1 harvests),

and (2) a leaching experiment for determination of net loss

of dissolved inorganic nitrogen (DIN), dissolved organic

nitrogen (DON), and dissolved organic carbon following the

rewetting incubation and a 9-week dry period (T2 harvest).

The litter addition was used to determine the effect of C

availability (litter C) on soil C and N release. Treatments

consisted of soils that received no litter (Klitter), or

2 mg C/g soil as native Adenostoma litter (Clitter),

equivalent to a 10% increase in soil C, or a 10% increase

in mean annual litterfall C (approximately 100 g mK2 yrK1;

A. Miller, unpublished data). Litter was collected in

August–September, 2002, air dried and ground to pass

through a 40 mesh (420 mm) screen, and was added to a

subset of the 4, 16, 60% WHC and 2-week rewet soils prior

to the start of the experiment, when all soils were still at 4%

WHC. All soils, whether they received litter or not, were

homogenized on a tumbler for 6 h prior to the start of the

rewetting treatments. Lignin, cellulose, and hemicellulose

concentrations in litter were determined by analysis of

sequential digests on an Ankom 200 Fiber Analyzer

(Ankom Technology, Fairport NY) at the University of

Minnesota. Litter carbon and nitrogen concentrations were

determined on an elemental analyzer at the University of

California, Santa Barbara.

Rewet treatments consisted of dry soils (4% WHC) that

were wetted to 60% WHC at either 2-week or 4-week

intervals, and then dried to 4% WHC (as determined by

mass) over a period of 5–6 days. Fans were used to increase

air circulation over the soils during the dry down period. We

used rewet frequencies of 2 and 4 weeks to simulate the

natural range of storm frequency, as determined from local

meteorological records (Sequoia and Kings Canyon

National Park, Division of Natural Resources,

1948–2002). Control treatments consisted of soils main-

tained at 4, 8, 16, or 60% WHC (K2.0, K1.2, K1.1, and

K0.9 MPa, respectively, for soils without litter addition).

The 8 and 16% WHC controls corresponded to the predicted

average moisture contents of soils rewet at 4- and 2-week

intervals, respectively; actual mean water holding capacity

in these soils over the course of the incubation was

estimated at 13 and 26%. The 4 and 60% WHC controls

corresponded to the extremes of soil moisture, dry and

moist, experienced by the rewet treatment soils. All soils

were maintained at 20 8C in the dark over the course of the

experiment. Rewet treatment soils subjected to leaching

were incubated at 4% WHC between the T1 harvest (16

weeks) and T2 harvest (25 weeks) without further rewetting,

while control soils were maintained at their respective

water-holding capacities (4, 8, 16, 60%) up until the time of

leaching (25 weeks).

2.2.2. Determination of C and N release

Rewetting and litter treatments were initiated in early

January, 2003. Ten days following the start of the

experiment and the first wetting–drying event, a subset of

soils from each treatment was harvested for determination

of initial (T0) microbial biomass and inorganic N pools

(nZ4). Soils were extracted with 0.5 M K2SO4

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–2204 2197

(5:1 extraction) and filtered through pre-rinsed Whatman #1

filters. Extractable NHC4 was analyzed using the diffusion

method on a Lachat autoanalyzer (Lachat Instruments,

Milwaukee, WI; Lachat Method 31-107-06-5-A), and NOK3

was analyzed using the Griess–Ilovsay reaction after Cd

reduction (Lachat Method 12-107-04-1-B). Microbial

biomass C and N were determined using the chloroform

fumigation–extraction method (Vance et al., 1987) over a

5-day fumigation period, and extracts were analyzed for

total C and N using a persulfate digestion technique (Doyle

et al., 2004). We did not apply a correction for extraction

efficiency to our estimates of biomass C and N, and thus

these values represent chloroform-labile pools of C and N

rather than the total microbial biomass.

Soil respiration was measured on rewet soils (nZ8) at

daily intervals during each rewetting event, beginning

approximately 4 h following wet-up and continuing through

the dry down period until soils again reached 4% WHC.

Respiration rates in control soils were measured every two

weeks. For each set of measurements, centrifuge tubes were

capped with gas-tight lids fitted with rubber septa and

respiration rates were calculated as the increase in head-

space CO2 over 2–3 h (rewet soils) or 24 h (control soils),

using an infrared gas analyzer with an in-line injection

system (Li-Cor LI-6252). Rewet soils were uncapped

following each set of respiration measurements to permit

continued drying. Control soils were loosely capped and

returned to humidified (8, 16, 60% WHC controls) or to dry

(4% WHC control) mason jars. All rewet soils and a subset

of control soils were weighed prior to each set of respiration

measurements in order to monitor changes in soil moisture,

if any.

Following 4–8 rewetting intervals (T1 harvestZ16

weeks), a subset of soils from each treatment was harvested

for determination of microbial biomass, inorganic N pools,

and net N mineralization and nitrification rates (nZ4). Soils

were harvested following the final wetting–drying event,

when rewet soils had returned to 4% WHC. Nitrification

potentials were determined using the chlorate–slurry

method (Belser and Mays, 1980) over a 24 h incubation

period (nZ3).

2.3. Leaching experiment

Following the final rewetting event and T1 harvest at 16

weeks, the remaining subset of soils from each soil moisture

and litter treatment was held at 20 8C for an additional 9

weeks without further rewetting disturbance (nZ4). Control

soils were held at their respective water-holding capacities

(4, 8, 16, 60% WHC), and rewet soils were maintained at

4% WHC. At the time of the leaching treatment (T2

harvestZ25 weeks), all soils were brought up to 60% WHC,

incubated for 24 h, and extracted in HPLC-grade water (5:1

extraction) with slow (50 rpm) shaking over 24 h. Soil

respiration was measured over a 2 h period, approximately

2 h following the final wet-up to 60% WHC. Water extracts

of soil were filtered through ashed and pre-rinsed GF/F

filters (0.7 mm). One aliquot of each sample was acidified

with concentrated HCl and analyzed on a Shimadzu TOC-V

(Shimadzu, Columbia, MD) at the University of California,

Riverside. The remaining aliquot was used for determi-

nation of NHC4 and NOK

3 , as above, and for determination of

total dissolved nitrogen (TDN) using the Valderrama (1981)

digestion method. Dissolved organic nitrogen was calcu-

lated by difference.

2.4. Statistical analyses

Rewetting effects on soil parameters were analyzed using

a general linear model (PROC GLM) in SAS (SAS Institute,

Carey, NC). Variables were log-transformed where necess-

ary to meet assumptions of normality and homoscedasticity.

Post hoc multiple comparison tests were performed among

rewet (2- and 4-week rewetting interval) treatments and

controls maintained at equivalent mean soil moisture (e.g.

16, 8% WHC) using adjusted least squares means. In

addition, rewet treatments were compared to 60% WHC

controls as a conservative estimate of rewetting effects. All

figures show untransformed means. Effects of litter addition

were analyzed separately within control and rewet soils

using a general linear model.

3. Results

3.1. Litter C and N concentrations

Lignin, cellulose, and hemicellulose concentrations in

litter were 22, 13, and 11%, respectively. The carbon

concentration in litter was 49.6%, and the nitrogen

concentration was 0.98%. Water extracts of litter produced

90.1 mg C g-litterK1, or approximately 18% of total litter C,

and 3.9 mg N g-litterK1, or approximately 40% of total

litter N, primarily as DON and NHC4 (data not shown).

3.2. Rewetting experiment: 16 week harvest

3.2.1. Soil respiration

Soil respiration rates peaked within 24 h of each

rewetting event and did not diminish markedly after the

first event (Fig. 1). Following each rewetting event,

respiration rates in 4-week rewet soils were approximately

twice as great as in 2-week rewets. Respiration rates

measured during the dry intervals (w4% WHC) did not

differ between the two rewet treatments (Fig. 1).

Cumulative CO2 release over the 16-week period

increased with soil moisture in the control soils but was

greatest in soils that experienced repeated drying and

rewetting (Fig. 2). When we accounted for the number of

days during which each rewet soil was moist, based on

measurements of soil mass during dry down periods,

cumulative CO2 release in rewet treatment soils was

Incubation period (days)

0 20 40 60 80 100

0 20 40 60 80 100

µ g C

g-s

oil-1

d-1

0

20

40

60

80

100

120

140

4 weeks2 weeks

Rewet interval

%W

HC

020406080

100

Rewet - Respiration rate

Rewet - Soil moisture

(a)

(b)

Fig. 1. Variation in mean soil respiration rate (a) with soil moisture (b) in 2-

and 4-week rewet (Klitter) soils. Soil respiration was measured over 2 h at

daily intervals, beginning 4 h after each rewetting event.

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–22042198

2.2–3.7 times greater than predicted by respiration from

controls maintained at equivalent mean soil moisture, and

50–60% greater than in 60% WHC controls (FZ27.33, dfZ1, P!0.001 and FZ21.55, P!0.001 for 2- and 4-week

rewets, respectively) (Fig. 2, Inset). Cumulative CO2 release

from 2- and 4-week rewet, Klitter soils did not differ.

We measured soil respiration rates in rewet treatment

soils during the final rewetting interval (16 weeks) and

again, immediately prior to leaching when all soils had been

brought up to 60% WHC (25 weeks). Respiration rates

Control - Cumulative CO2 release

4% 8% 16%

µg C

g-s

oil-1

0

1000

2000

3000

4000

R

WHC (%)

0 10 20 30 40 50 60

µ g C

g-s

oil-1

0

500

1000

1500

2000

2500

3000

REWET

CONTROL

ND

WHC

Fig. 2. Cumulative CO2 release in control and rewet treatment soils following the 16-w

different from control soils maintained at equivalent mean soil moisture (*P!0.0

(Klitter) treatments are shown by P-values. Lower case letters denote differences be

indicates that no data are available for the Clitter treatment in the 8% WHC contro

cumulative respiration in control (Klitter) soils, showing measured CO2 release in r

following the wet-up at 25 weeks were approximately five

times greater in 4% WHC controls than in all other soils,

suggesting a large release of litter-derived C, and were

lowest in 2-week rewet soils (data not shown). Litter

addition increased cumulative CO2 release at 16 weeks only

in the 60% WHC control and the 2-week rewet soil (Fig. 2).

3.2.2. Microbial C and N pools

Rewetting and litter addition had little effect on

microbial C pools measured at 16 weeks, while rewetting

slightly enhanced, and litter reduced, microbial N (Fig. 3).

While frequent (2-week) rewetting did reduce microbial C

pools relative to 16 and 60% WHC controls (Fig. 3a),

4-week rewet intervals had no effect. Litter addition

enhanced microbial C only in the 4% WHC soils, where it

increased the microbial C pool by approximately one-third,

but it reduced both microbial C and N in the 2-week rewet

soils (Fig. 3).

3.2.3. Nitrification potential, net N mineralization,

and net nitrification

Nitrification potential, an index of the size of the

NH3-oxidizer population measured at 16 weeks, increased

with soil moisture in the control treatments, but decreased

with rewetting and litter addition, (Fig. 4a). Rewetting

frequency did not affect nitrifier activity (Fig. 4a), although

2-week rewet soils showed lower nitrification potentials

than 16% (FZ33.23, dfZ1, P!0.0001) and 60% WHC

controls (FZ15.36, dfZ1, P!0.01).

Net N mineralization and nitrification measured at

16 weeks increased with soil moisture and decreased with

60%

-litter+litter

Rewet

4 week 2 week

0

***

***

***

ewet treatment

N.S.b

a b

a

ND

Rewet frequency

eek incubation (meansG1 SE). Asterisks denote rewet treatments significantly

5; **P!0.01; ***P!0.001). Differences between the 4- and 2-week rewet

tween Klitter and Clitter treatments for a given soil moisture treatment. ND

l and 4-week rewet treatments. Inset: Relationship between soil moisture and

ewets relative to CO2 release predicted by controls.

Rewet treatment

4% 8% 16% 60%

4% 8% 16% 60%

µg C

g-s

oil–1

0

50

100

150

200

250

300

4 week 2 week

4 week 2 week

–litter+litter

Control - Microbial C Rewet

***

(a)

P<0.001

Control - Microbial N

µg N

g-s

oil–1

0

20

40

60

80Rewet

*

(b)

N.S.

***

**

ND ND

ND ND

Fig. 3. Microbial (chloroform-labile) C (a) and N (b) in control and rewet soils following the 16-week incubation (meansG1 SE). Treatment differences are

denoted as in Fig. 2.

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–2204 2199

litter addition in control soils. Rewetting frequency

impacted both net N mineralization and net nitrification

rates, as 2-week rewet soils showed enhanced (Fig. 4b) or

equivalent rates (Fig. 4c) of N cycling, relative to 16%

controls, and significantly greater rates of N turnover than

soils wetted at 4-week intervals (Fig. 4b and c). Litter

addition reduced net nitrification rates by 75–88% across

soil moisture treatments, and to a lesser extent also impacted

net N mineralization (Fig. 4b and c).

3.3. Leaching experiment: 25 week harvest

3.3.1. Water-extractable C and N

Soil drying–rewetting increased extractable NHC4 , DOC

and DON concentrations from soils at 25 weeks, relative to

soils maintained at equivalent mean soil moisture (Table 1).

Extractable NOK3 was not affected by rewetting but was

reduced by litter addition in most soils. In 2-week rewets,

NOK3 accounted for approximately 50% of total soluble N in

soils without litter addition (Klitter), but only 10% of total

N when litter was added (Clitter). As with inorganic N

concentrations, extractable DON and DOC increased with

rewetting frequency; however, DON and DOC were also

high in 4% WHC controls, particularly in the Clitter

treatment. Extractable C:N (w23) exceeded soil C:N (w20)

only in the 4% WHC, Clitter soils, indicating that C and N

losses from litter strongly controlled leaching patterns from

dry soils.

Given the difference in DOC concentrations between

Klitter and Clitter treatments in the 4% WHC soils

(Table 1), and that approximately 18% of litter C was

available for leaching (see Section 3.1, Methods), we

estimate that 67% of soluble C was released from these

soils in a single wetting event. In contrast, the only slight

increase in total soluble N (DINCDON) with litter

addition suggests that !10% of the soluble N pool was

released from the 4% WHC soils at the same time

(Table 1).

Control - Nitrification potential

0

1

2

3

4

–litter+litter

Rewet

**

*

N. S.

ND NDb

a

b

(a)

Control - Net N mineralization

–0.1

0.0

0.1

0.2

0.3

0.4

0.5Rewet

Control - Net nitrification

4% 8% 16% 60%

µg N

g-s

oil–1

d–1

µg N

g-s

oil–1

d–1

µg N

g-s

oil–1

d–1

–0.1

0.0

0.1

0.2

0.3

0.4

0.5Rewet

4 week 2 week

***

*

(b)

(c)

**

ND

ND ND

ND

a

b

a

b

a

b

a

b

a

b

a b

ab

Rewet treatment

bND

a

b b

ND

**

*

P<0.01

P<0.0 1

N.S.

Fig. 4. Nitrification potential (a), net N mineralization (b) and net nitrification (c) measured in control and rewet treatment soils following the 16-week

incubation (untransformed meansG1 SE). Treatment differences for nitrification potential were determined on log-transformed data. Treatment effects are

denoted as in Fig. 2.

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–22042200

3.3.2. Carbon and nitrogen budgets

Carbon release (CO2CDOC) in rewet soils ranged from

12–18% of total soil C, with greater than 90% of C loss

attributed to soil respiration (Table 1). Relative to the

chloroform-labile C pool, these CO2 losses were large,

ranging from approximately 1.4–16 times greater than the

microbial C pool in control, Klitter soils, and from 16 to 28

times greater in the rewet, Klitter soils. DOC release was

negligible in all treatments but the 4% WHC, Clitter soils,

where it accounted for a two-fold greater release of C than

respiration (1.5 vs. 0.7% of total soil C, respectively; Table 1).

Total dissolved N (TDN) release from soils was not as

great as carbon release and ranged from 5–10% of total soil

N in rewet soils. With the exception of 4-week rewet soils,

which showed low nitrification rates after 16 weeks, net

nitrate production extrapolated over the 25-week incubation

accounted for 67–90% of NOK3 release. TDN losses in soils

without litter ranged from 1.0 to 2.4 times greater than

Table 1

Soluble C and N concentrations in water extracts of soil at 25 weeks (meansG1 SE)

Treatment NHC4 (mg N/g) NOK

3 (mg N/g) DON (mg N/g) TDN (mg N/g) DOC (mg C/g) CO2K25 weeks

(mg C/g)

Controls

4% WHC, Klitter 1.9 (0.3) 3.6 (0.2)a 7.7 (0.2)a 13.2 (0.5) 96.7 (8.4)a 154.9 (1.6)

4% WHC, Clitter 1.4 (0.3) 0.1 (0.1)b 13.1 (1.1)b 14.6 (1.3) 344.3 (52.7)b 156.0 (4.5)

8% WHC, Klitter 2.5 (0.4) 16.0 (0.9) 6.1 (0.4) 24.6 (1.1) 85.1 (3.8) 489.1 (I30.5)

16% WHC, Klitter 1.4 (0.5) 38.8 (7.2)a 4.8 (0.5) 45.0 (7.9)a 47.2 (5.8) 995.5 (99.4)

16% WHC, Clitter 0.5 (0.0) 10.9 (3.4)b 3.6 (0.4) 15.0 (3.6)b 50.7 (15.1) 1509.1 (85.8)

60% WHC, Klitter 0.6 (0.1) 48.3 (13.1) 1.9 (0.4) 50.8 (13.1) 22.4 (3.6) 2389.4 (63.9)a

60% WHC, Clitter 0.6 (0.4) 48.0 (7.7) 1.2 (1.3) 49.8 (7.7) 34.8 (2.9) 3638.6 (123.4)b

Rewets

4-week, Klitter 12.7 (1.3)*** 35.4 (2.3)*** 11.1 (1.4)** 59.1 (4.8)*** 114.7 (11.5)* 2779.5 (219.2)***

2-week, Klitter 21.4 (1.0)*** 52.2 (1.3)a 27.1 (1.1)*** 100.7 (1.2)***a 192.4 (6.3)*** 2622.6 (122.5)***a

2-week, Clitter 26.6 (2.0)*** 5.6 (1.8)b 19.5 (0.3)*** 51.7 (2.8)***b 210.1 (2.4)*** 4167.7 (183.8)***b

Lower case letters denote effects of litter addition significant at aZ0.05 within soils of a given moisture content (e.g. 4% WHC or 2-week rewet). Asterisks

denote significant effects of rewetting, relative to controls maintained at equivalent mean water content (*P!0.05, **P!0.01, ***P!0.001). Cumulative

CO2 release at 25 weeks is shown for comparison with DOC release.

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–2204 2201

chloroform-labile N pools in controls, and from 2.3 to 3.4

times greater in rewet soils.

4. Discussion

The response of soils to repeated drying and rewetting

could not be predicted from control soils maintained at

equivalent mean soil moisture, or even from soils

maintained at 60% WHC. This finding can be explained

by several factors: (1) soil drying and rewetting increased C

release, and to a lesser extent N release, relative to soils

maintained at a constant soil moisture; (2) rewetting

frequency affected the magnitude of the CO2 pulse during

each rewetting event, but not the cumulative release of CO2;

and (3) variation in soil moisture was more important than

litter inputs in controlling soil C and N dynamics.

4.1. Effect of soil drying and rewetting on C release

Carbon release, dominated by soil respiration, was

significantly greater in soils that experienced repeated

drying and rewetting than in soils maintained at either

equivalent mean soil moisture (8 and 16% WHC), or at 60%

WHC. Respiration rates following rewetting events were

approximately twice as great in the 4-week rewet treatment

as they were in the 2-week rewet treatment, in agreement

with the idea that rewetting history influences the magnitude

of the CO2 pulse (e.g. Soulides and Allison, 1961; Fierer and

Schimel, 2002). As a result, doubling the number of rewet

events over the 16-week incubation period had no effect on

cumulative CO2 release. Soluble C release was also high in

the rewet soils and may have derived from reduced

microbial utilization of DOC during dry periods (Lundquist

et al., 1999), increased turnover of microbial biomass and

microbial products with rewetting (Roberson and Firestone,

1992; Fierer and Schimel, 2003), and/or the release of

sequestered C from soil aggregates. While it seems likely

that microbial utilization of DOC was low during dry

intervals, we see little evidence that the large losses of C

from rewet soils could likewise be explained by microbial

turnover.

First, we found little change in the size of the microbial C

pool with rewetting (145 mg C g-soilK1 in the 60% control

vs. 100 mg C g-soilK1 in the 2-week rewet) relative to the

magnitude of CO2 release (1540 mg C g-soilK1 vs.

2295 mg C g-soilK1). If we assume that microbes in the

60% WHC treatment had access to at least as much C as

rewet soils (approximately 2300 mg C g-soilK1), then after

the release of roughly 1550 mg C g-soilK1 via respiration,

approximately 750 mg C g-soilK1 must have been returned

to the soil carbon pool, presumably recycled through the

biomass as SOM. The efficiency of C return in such a

scenario would be approximately 33%, and even greater if

the actual available C pool were larger. This would

represent a substantial recycling of the initial C pool back

into restabilized organic forms, and at least a 500% turnover

of the entire microbial biomass, accounting for the fact that

chloroform-labile C is only a portion of the biomass. A

substrate use efficiency of 33%, though plausible, would be

high even in short-term turnover studies (e.g. Sugai and

Schimel, 1993; Devevre and Horwath, 2000), since long-

term studies of 14C decomposition indicate that on average,

only 15–30% of added 14C is retained in the total soil carbon

pool (e.g. Paul and McGill, 1977; Paul and Voroney, 1980;

Van Veen et al., 1984). How, then, was so much labile C

generated within the rewet treatments if not through

microbial turnover? How do we account for the differences

in cumulative CO2 release between the rewet and 60%

WHC treatments when the differences in microbial C pools

are roughly an order of magnitude lower?

One possible explanation is that cumulative CO2 and

DOC release were enhanced in rewet soils through a

combination of physical disruption that could expose

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–22042202

recalcitrant SOM-C (Degens and Sparling, 1995; Lundquist

et al., 1999; Magid et al., 1999), and extracellular enzyme

activity that could transform this SOM-C into a stress-labile

C pool. Soils maintained at 60% WHC, in contrast, would

have been subjected to only one wet-up at the beginning of

the incubation, and thus would have experienced only a one-

time disruption and release of this slow-cycling SOM-C.

This mechanism of paired SOM disruption and exoen-

zyme activity could explain both the two-fold difference in

respiration rates between the 2- and 4-week rewet

treatments, and the insensitivity of long-term CO2 release

to rewetting frequency. Although exoenzyme activity is

correlated with soil moisture (e.g. Fioretto et al., 2001), low

levels of activity, and hence labile C production, are still

possible during prolonged dry periods. The regeneration of

labile C during these periods of low biological demand, and

the diffusion of accumulated C to microbes upon rewetting

could together produce the large fluxes of CO2 that we

observed. If the active lifetime of the exoenzyme pool

equaled or exceeded the longest dry interval between

rewetting events (here, approximately 3.2 weeks in the

4-week rewet treatment), such a flow of organic material

could result in equivalent fluxes of carbon from soils

experiencing different rewetting frequencies.

4.2. Effect of drying and rewetting on soil N dynamics

A second question that arises from our work is why N

release, but not C release, was sensitive to rewetting

frequency. While increasing the number of rewetting events

had no effect on CO2 release or on microbial C, it resulted in

slight to significant increases in microbial N, net N

mineralization, and nitrification rates that suggested

enhanced breakdown of SOM. This apparent lack of

correspondence between C and N dynamics in response to

multiple rewetting events indicates that the mechanisms

controlling each may differ, but is not inconsistent with our

findings for C release. A single rewetting pulse may

generate carbon from multiple sources: a stress-labile pool

comprised of a fast cycling, light fraction that could

stimulate net N mineralization, and a slower cycling,

heavy fraction that could promote N immobilization (Parton

et al., 1998). Based upon the results from this experiment, it

appears that a change in the frequency of the rewetting pulse

could alter the balance between N mineralization and

immobilization, as controlled by N incorporation into fast-

and slow-cycling SOM pools, while having no net effect on

C release. As such, it would appear that increasing the

frequency of rewetting could enhance the release of more

labile, fast-cycling SOM, consistent with a scenario of more

frequent aggregate disruption. Model (DAYCENT) simu-

lations run in parallel with our experimental work produced

a similar response for N mineralization (X. Li, unpublished

data), suggesting that the complex N dynamics we observed

may be due to the shifting utilization of these fast- and slow-

cycling SOM pools over time.

In spite of reduced nitrification rates with infrequent

(4-week) rewetting, NOK3 losses from 4-week rewet soils

were high. Long-term increases in microbial biomass and

nitrifier activity observed following rewetting (e.g. Fierer

and Schimel, 2002) could explain the shift from NOK3

immobilization to NOK3 production over time. Assuming

that infrequent rewetting was less stressful to nitrifiers than

frequent rewetting, and that the intervening dry period

between the cessation of rewetting events (16 weeks) and

the final leaching (25 weeks) was sufficient to allow

population recovery, it appears that a dry interval following

repeated pulse events could ultimately enhance NOK3 release

from soils. In this sense, our findings indicate that the wet

season prior to the summer drought may control the

magnitude of N loss observed during the current year’s

wet-up.

4.3. Effect of litter addition on soil N dynamics

Litter addition increased microbial biomass N and

generally enhanced NOK3 retention across soils maintained

at constant water-holding capacity. We used the litter

additions to infer mechanisms of C release, i.e. via soil

respiration or leaching of DOC, and to infer the potential

source of the released C in soils of differing moisture

content. We found that with initial wetting, C release in

undisturbed, dry soils was significantly greater than from

both rewet soils and soils that had been maintained at higher

soil moisture. The substantial mobilization of litter C during

a single rewetting event may explain the strong effect that

rewetting treatments had on net N mineralization, nitrifica-

tion, and water-extractable NOK3 pools in our study, and the

absence of this litter-derived ‘pulse’ of DOC from soils

maintained at higher average soil moisture. Indeed, total

carbon release per g C from Clitter, rewet soils was 60%

greater than from Klitter soils, in spite of an only 10%

increase in soil C with the litter amendment. Such a

difference between the Clitter and Klitter rewet treatments

suggests that the addition of potentially labile litter C may

have enhanced the initial breakdown of existing soil C,

effectively ‘priming’ the system (Schimel and Weintraub,

2003).

4.4. Conclusions

In many Mediterranean-type ecosystems, the short-term

drying and wetting pulses that characterize the transition

from summer drought to winter rains may have a

disproportionate effect on long-term C and N fluxes, the

magnitude of which may be strongly controlled by soil

carbon pools. These seasonal transitions mark a period of

potentially large losses of carbon and nitrogen from

environments that are already nutrient-limited and have

little capacity for nutrient retention. The finding that total N

and DOC release, but not CO2 release, may be determined

by rewetting frequency suggests that the mechanisms

A.E. Miller et al. / Soil Biology & Biochemistry 37 (2005) 2195–2204 2203

responsible for these losses are independent. While nitrogen

dynamics may rely primarily on the shifting balance

between utilization of active and slow SOM pools, the

relationship between rewetting history and decomposition

rates appears to be different for soil CO2. The mechanism

for the overall enhancement of respiration due to repeated

drying/rewetting appears to involve a physical process that

exposes additional organic matter to attack, likely through

the disruption of aggregates by rewetting, although such a

process remains to be shown conclusively. The loss of up to

18 and 10% of total soil C and N, respectively, with repeated

soil drying and rewetting indicates the potential importance

of these pulse events to annual nutrient budgets. Elucidation

of the mechanism(s) promoting carbon turnover in these

soils will therefore be critical to our understanding of the

biogeochemical interactions that occur during seasonal

transitions, and the processes that control nutrient retention

and loss over the longer term.

Acknowledgements

We thank Allen Doyle, University of California, Santa

Barbara, and Kendra McLaughlin, University of Minnesota,

for sample analyses. Kevin Skeen and Annie Esperanza

assisted with field collections and logistics, and Aimee

Davingnon, Shelly Cole, and Julie Butera assisted in the lab.

Joe Anderson and two anonymous reviewers provided

helpful comments on an earlier draft. Funding for this

project was provided by the National Science Foundation

Ecosystems Program (DEB-008939), and logistical support

by Sequoia-Kings Canyon National Park.

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