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Soil available phosphorus status determines indigenous

mycorrhizal colonization of field and glasshouse-grown

spring wheat from Argentina

Fernanda Covacevich a,*, Hernan E. Echeverrıa a,b,Luis A.N. Aguirrezabal a

a Facultad de Ciencias Agrarias (UNMP), INTA Balcarce, C.C. 276, Balcarce, Buenos Aires 7620, Argentinab Estacion Experimental Agropecuaria INTA Balcarce, C.C. 276, 7620, Balcarce, Buenos Aires 7620, Argentina

Received 22 February 2006; received in revised form 29 April 2006; accepted 8 June 2006

Abstract

Wheat production (Triticum aestivum L.) has increased across the world during last century with the intensification of

agriculture. Phosphorus (P) fertilization is a common practice to improve wheat growth in Argentina. We investigate whether

indigenous arbuscular mycorrhizal colonization (AMC) of hard red spring wheat is controlled by shoot P content (SPc) or by

available soil P in an agricultural soil from the southeastern Argentine Pampas. In the field, AMC was monitored four times during

two growing seasons of a conventional wheat crop. Treatments were: without P supply, annual supply of 11 and 22 kg P ha�1 during

the last 5 years, and 164 kg P ha�1 applied once 5 years before the experiment. In the glasshouse, AMC was assessed three times in

wheat growing in pots filled with the soil from unfertilized plots; treatments were: P (0 and 20 mg P pot�1), and nitrogen (N)

fertilization (0 and 150 mg N pot�1). A range of soil P between 6 and 60 mg P kg�1 was obtained and the AMC ranged from 1% to

67% of root length colonized under both field and glasshouse conditions. P supplied annually increased growth and SPc but

decreased AMC. N fertilization did not affect growth or AMC. Variations in SPc did not account for AMC. Variability in AMC was

best accounted for local current soil available P content (r2 = 0.59). A linear-plateau relationship between soil P and indigenous

AMC was established in wheat plants growing under contrasting environmental and experimental (field and glasshouse) conditions.

Indigenous AMC was depressed by available soil P in the range 0–27 mg P kg�1 (a decrease of 2.8% mg P�1 kg�1). Above

27 mg P kg soil�1, AMC was stabilized at about 10%. Grain yield increased with fertilization and the highest relative shoot dry

matter in field was obtained at 15.5 mg P kg soil�1. The soil P range that ensures high wheat production without deterring

indigenous AMC is discussed.

# 2006 Elsevier B.V. All rights reserved.

Keywords: Arbuscular mycorrhizal colonization; Wheat; Soil available phosphorus; Shoot P concentration

1. Introduction

During the 20th century global wheat (Triticum

aestivum L.) yield increased considerably, from <1 to

ca. 2.5 Mg ha�1. This increase in yield was the most

remarkable feature of the intensification of the wheat

cropping system of the last century. Two factors were

largely responsible for the production levels achieved

by farmers: better management practices (the techno-

logical component of yield gains) and new improved

genotypes (the genetic component of yield gains)

(Veron et al., 2004).

www.elsevier.com/locate/apsoil

Applied Soil Ecology 35 (2007) 1–9

* Corresponding author. Fax: +54 266 439101.

E-mail addresses: covac@mdp.edu.ar (F. Covacevich),

hecheverr@inta.gov.ar (H.E. Echeverrıa).

0929-1393/$ – see front matter # 2006 Elsevier B.V. All rights reserved.

doi:10.1016/j.apsoil.2006.06.001

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Wheat is one of the most important crops in

Argentina, with average production of 14.9 millon -

tonnes over the last 5 years. Thirty percent of this

amount is produced in the southeastern Humid Pampa

region of the Argentine Pampas. The southern Buenos

Aires province, where this study was carried out, does

not have a long cropping history. The soils of this region

are moderately acid, usually contain 60–80 g kg�1

organic matter content and have low concentrations (6–

8 mg kg�1) of native available phosphorus (P) (Eche-

verrıa and Ferrari, 1993). Wheat has high P and N

requirements (Baker and Tucher, 1973; Karlen and

Whitney, 1980), thus, fertilization with P (15–

25 kg P ha�1) and N (120 kg N ha�1) are common

practices for farmers in order to increase wheat yield.

Soils of the southern Buenos Aires province contain

indigenous arbuscular mycorrhizal fungi (AMF) (Cov-

acevich et al., in press) that colonize wheat (Covacevich

et al., 1995). The growth enhancement and P uptake of

plants colonized by AMF is a well-known process

(Pfleger and Linderman, 1996; Schweiger and Jakob-

sen, 1999; Jeffries et al., 2003). Although wheat is not

highly dependent on mycorrhiza (Azcon and Ocampo,

1981; Trouvelot et al., 1982; Hetrick et al., 1993), some

increases in wheat yield following inoculation with

AMF have been reported (Jakobsen and Nielsen, 1983;

Baon et al., 1992; Talukdar and Germida, 1994; Xavier

and Germida, 1997; Al-Karaki et al., 1998) particularly

in low-P soils (Thompson, 1990; Rubio et al., 2003).

With the current tendency for a reduced use of

agrochemicals, research is currently aimed at crop

yield improvement and at yield sustainability. Thus,

microbial-based approaches have been proposed to

improve crop yield.

Plant responses to agronomic practices and to

environmental stresses have been widely studied.

However, reports of the effects of soil P fertilization

on indigenous microorganisms in the rhizosphere have

been, in general, contradictory. Research on the

mycorrhizal symbiosis is often conflicted because while

soil P fertilization increases soil available P, plant P

concentration, and improves plant growth, high levels of

P may depress AMC (Jensen and Jakobsen, 1980; Baon

et al., 1992; Sylvia and Williams, 1992; Pfleger and

Linderman, 1996; Covacevich et al., in press). Attempts

have been made to establish whether AMC are more

affected by the soil or plant P status. The literature is not

clear on this point: Kurle and Pfleger (1996) concluded

that plant P status modulates AMC, whereas Miranda and

Harris (1994) demonstrated that soil P might have a direct

effect on AM external hyphal growth. Quantitative

relationships between soil or plant P levels and AMC are

needed in order to adjust the fertilizer dose to obtain

acceptable yield production without depressing the

symbiosis. This study included field and glasshouse

experiments to investigate whether the indigenous AMC

in spring wheat is mainly regulated by soil available P or

by shoot P content.

2. Materials and methods

2.1. Study site

Experiments were conducted at the Experimental

Station Unidad Integrada Facultad de Ciencias Agrar-

ias (UNMdP), Estacion Experimental Agropecuaria

(INTA) Balcarce, Argentina (378450S lat, 588180Wlong; 138 m a.s.l.). The climate of the region is humid

subhumid mesothermal. The annual mean temperature

is 14.6 8C and the annual average rainfall is 954 mm

with 80% of rainfall during spring-summer. The soil is

a moderately well drained Chernozemic loam (FAO

soil classification), a Petrocalcic Paleudoll (series

Balcarce), fine, mixed, thermic (USDA soil classifica-

tion). It has a petrocalcic horizon at a depth of 1.2 m and

a clay horizon at 33–74 cm depth. Mean soil

temperature at 20 cm depth is 8.5 8C in winter and

14.8 8C in spring. The soil has a mycorrhizal potential

of 0.70 (0.21–2.30) AM propagules g�1 dry soil (most

probable number method, average of six replications,

test plant: wheat; Porter, 1979). A brief description of

the soil properties at the start of the experiment is

shown in Table 1.

2.2. Field study

Before the beginning of the study, the experimental

site had been under conventional wheat monoculture for

5 years. Hard red spring wheat (T. aestivum L., cv.

ProINTA Federal) was sown (450 seed m�2) at the end

F. Covacevich et al. / Applied Soil Ecology 35 (2007) 1–92

Table 1

Soil properties in the Ap horizon (0 � 20 cm) of the experimental site

Soil parameter Experimental site

Organic matter (g kg�1) 62

Sand (g kg�1) (2 � 0.02 mm) 360

Silt (g kg�1) (0.02 � 0.002 mm) 390

Clay (g kg�1) (<0.002 mm) 250

P (Bray) (mg kg�1) 6.5

N–NO3 (mg kg�1) 15

pH (H2O) (1:2.5) 5.7

CEC (cmol kg�1) 33.9

Slope (%) 3.5

Bulk density (Mg m�3) 1.31

Particle density (Mg m�3) 2.58

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of July and grown under rain-fed conditions. Four

treatments in a randomized complete block design with

three replications were assessed: control without P

supply (P0), annual supply of 11 and 22 kg P ha�1 (P11

and P22, respectively) during previous 5 years, and

164 kg P ha�1 (P164) applied once 5 years before the

experiment. Plot size was 400 m2 and P was applied at

sowing as commercial triple-calcium superphosphate

(TSP). All plots were fertilized annually with

120 kg N ha�1 as commercial urea.

Shoot and root samples were taken four times

(Table 2) during two consecutive field-growing seasons

(Experiment 1 and Experiment 2, respectively). At each

harvest, one quadrate (0.25 m2 of plant material) per

plot was randomly cut to ground level. At the following

harvest, the plant material was cut from the same plot

but in a different quadrate than the former harvest.

Shoot dry matter (SDM), and grain yield (GY, 120 m2 of

each plot) were determined after oven-drying samples

(60 8C). Total P concentrations (SPc) of shoot or grains

were measured (Walinga et al., 1995).

At each harvest, roots and soil samples were collected

at random in each plot (ten cores of 10 cm diameter and

20 cm depth). At the following harvest, the root–soil

samples were extracted from the same plot but in a site

different than the former harvest. Roots were washed to

remove soil particles and collected by sieve (0.5 mm).

Roots were cut (1 cm), thoroughly mixed and stained

according to Phillips and Hayman (1970): roots were

cleared with KOH (10%, 30 min, 90 8C), acidified (HCl,

0.1N), and stained with trypan blue (0.05%, 5 min,

100 8C). Presence of mycorrhizal infection was assessed

by microscopic examination (40� and 100�) of the

stained roots system. A segment was considered infected

if contained arbuscules or coils plus hyphae and/or

vesicles. Colonization was assessed using the Trouvelot

et al. (1986) method that allows the simultaneous

evaluation of frequency of colonization, intensity of

arbuscular mycorrhizal colonization (AMC), and the

proportion of arbuscules (A) present in the roots.

Composite soil samples (0–20 cm depth) were used also

for measuring inorganic P (Bray I: Bray and Kurtz, 1945).

Climatic data was obtained from a meteorological

station located 400 m from the experimental site.

Precipitation was greater during Experiment 2 than

Experiment 1 (995 and 860 mm, respectively). Soil

water deficit occurred during anthesis of Experiment 1.

Mean air and soil temperatures were similar in both

years; however, frequency of freezing was higher during

Experiment 1 than Experiment 2 (data not shown).

At the end of the experiments, indigenous arbuscular

mycorrhizal spores were collected from topsoil of

unfertilized (P0) plots (5 kg, 0–20 cm). The rhizosphere

soil samples were wet-sieved and decanted according to

the methodology described by Siverding (1991). Each

spore type was mounted sequentially in water lacto-

phenol, PVA (polyvinyl lactic acid) and Melzer’s

reagent for identification. Mycorrhizal identification

was based on spore colour, size, surface ornamentation

and wall structure, with reference to the descriptions

provided by the International collection of vesicular and

arbuscular mycorrhizal fungi (Morton and Benny, 1990;

http://invam.caf.wvu.edu).

2.3.Glasshouse study

Soil was collected in July from the P0 plots of the

field study site. Soil from the surface (0–20 cm, Ap

horizon) or subsurface (20–40 cm, B1 horizon) was

ground though a 1 cm sieve. In the Ap horizon soil

properties were similar to those described in Table 1; the

B1 soil had an organic matter content of 37 g kg�1, pH

5.9, and 4 mg Bray P kg�1. Experimental units

consisted of PVC pots of 30, 50 and 100 cm depth,

and 10 cm diameter. Pots were filled with the soil (3, 5

and 10 kg soil pot�1 within 30, 50 and 100 cm depth

pots, respectively). In order to simulate the soil profile,

the upper 20 cm of pots were filled with the Ap horizon,

and the remaining depth (below 20 cm of pots) with soil

from the field B1 horizon. Three sets of four treatments

in three randomized replications were designed for

three sampling dates: two P levels: 0, and 20 mg P pot�1

(P0 and P20, respectively); and two N levels: 0, and

150 mg N pot�1 (N0 and N150, respectively). P and N

F. Covacevich et al. / Applied Soil Ecology 35 (2007) 1–9 3

Table 2

Phenological stages, days after planting (DAP), and Zadoks et al. (1974) code (ZC) into field-grown spring wheat (field experiment 1 and 2,

respectively), and into glasshouse-conditions (Glasshouse)

Field experiment 1 Field experiment 2 Glasshouse

Early tillering (DAP 44; ZC 21) End tillering (DAP 59; ZC 25) End tillering (DAP 35)

Stem elongation (DAP 52)

Anthesis (DAP 88; ZC 65) Anthesis (DAP 93; ZC 67) Anthesis (DAP 60)

Milk development (DAP 109; ZC 70) Milk development (DAP 119; ZC 72)

Ripening (DAP 138; ZC 90) Ripening (DAP 147; ZC 90)

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were added in water solution at sowing as TSP and urea,

respectively.

Seeds of wheat (T. aestivum L., cv. ProINTA federal)

were surface disinfected (30 min; 7% calcium hypo-

chlorite solution), cold-treated (4 8C, 2 days), germinated

and maintained in a constant environment room (12-h

photoperiod provided by fluorescent lighting, 21/24 8Cmean temperature, 55/65% mean relative humidity,

320 mE m�2 s�1 irradiance) until 3 days after emergence.

Three germinated homogeneous seedlings of 3 days age

were transplanted per pot (equivalent to 380 plants m�2).

Plants were grown in a glasshouse (14/30 8C day/night

mean air temperature; 40/100% day/night mean air

relative humidity; 7.8� 1.1 MJ m�2 day�1 mean solar

radiation at the top of plants), watered with distilled water

and maintained at 60% (w/w) of the soil’s moisture-

holding capacity.

Plants within 30, 50 and 100 cm depth pots were

harvested at 35, 52 and 60 days after planting (DAP),

respectively (Table 2). SDM and SPc were measured as

above. Pots were opened, soil and roots of 0–20 cm and

>20 cm depth in pots were removed and analyzed

separately. Roots were washed and collected by sieve

(0.5 mm). Root dry matter was measured (based on a sub-

sample of 50% of the total root fresh weight). The

remaining roots were used to estimate mycorrhizal infec-

tion intensity and arbuscule content as described above.

2.3. Statistical analysis

Data were analysed by ANOVA (SAS Institute Inc.,

1988). Means among treatments were compared with

the least significant difference (LSD) test (P � 0.05).

All presented data are means of untransformed values.

We explored the relationship between SDM, GY, SPc,

AMC and available soil P. For that, SDM and GY were

normalized by the maximum value obtained at each

sampling date for each field experiment. Linear-plateau

F. Covacevich et al. / Applied Soil Ecology 35 (2007) 1–94

Table 3

The effect of phosphorus (P) fertilization on soil Bray-P (Soil-P), shoot dry matter (SDM), shoot P concentration (SPc) and arbuscular mycorrhizal

colonization (AMC) at 0–20 cm of spring wheat growing in the field during 2 years (field experiment 1 and 2, respectively)

Field experiment 1 Field experiment 2

Treatment DAP Soil-P

(mg kg�1)

SDM

(kg ha-1)

SPc

(g kg�1)

AMC

(%)

DAP Soil-P

(mg kg�1)

SDM

(kg ha�1)

SPc

(g kg�1)

AMC

(%)

P0 0 6.5 c 0 6.8 c

P11 11.0 b 7.6 c

P22 16.9 a 24.5 a

P164 16.0 a 15.9 b

LSD 1.81 3.50

P0 44 190 b 0.013 c 28.5 a 59 10.4 c 300 b 0.014 b 22.5 a

P11 244 ab 0.027 a 15.2 b 29.8 b 650 a 0.024 a 7.1 bc

P22 282 a 0.029 a 5.4 c 60.3 a 769 a 0.027 a 0.1 c

P164 215 ab 0.017 b 11.0 bc 20.8 bc 550 b 0.018 b 17.7 ab

LSD 69 0.003 9.52 15.10 154 0.004 13.20

P0 88 2166 b 0.010 b 37.7 a 93 9.2 c 3071 c 0.019 b 56.7 a

P11 2450 ab 0.011 a 21.3 ab 17.6 b 4911 ab 0.020 ab 39.8 ab

P22 3398 a 0.012 a 12.7 b 30.7 a 5862 a 0.022 a 24.9 b

P164 2390 ab 0.012 a 25.2 ab 15.1 b 4661 b 0.023 a 58.2 a

LSD 1037 0.002 21.26 5.41 967 0.003 28.94

P0 109 3314 b 0.007 b 49.6 a 119 10.6 c 8031 b 0.012 b 63.4 a

P11 5090 a 0.009 a 33.7 b 19.8 b 11262 a 0.014 ab 36.0 b

P22 5594 a 0.009 a 17.2 c 37.5 a 11644 a 0.014 ab 28.3 b

P164 5545 a 0.009 a 41.4 ab 15.8 bc 9678 ab 0.016 a 61.1 a

LSD 2264 0.002 12.07 6.70 2028 0.002 24.68

P0 138 9.7 b 5308 b 0.006 b 53.1 a 147 7.5 c 9206 b 0.007 b 57.3 a

P11 15.2 a 6615 a 0.007 b 25.0 b 12.4 bc 11614 ab 0.010 a 34.2 c

P22 23.5 a 7539 a 0.012 a 9.0 c 23.8 a 12160 ab 0.011 a 20.7 d

P164 20.7 a 7147 a 0.011 a 29.7 b 13.4 b 13108 a 0.010 a 45.0 b

LSD 7.42 2148 0.002 15.51 5.11 3407 0.002 11.46

Treatments: P0, control without P application; P11 and P22, annual application of 11 and 22 kg P ha�1 during the last 5 years, and P164,

164 kg P ha�1 applied once 5 years ago the experiment. DAP, days after planting. At each DAP, different letters in the same column show significant

differences at P < 0.05.

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models were fitted using the Table Curve program

(Jandel TBLcurve, 1992).

3. Results

3.1. Field experiments

A wide range of soil P levels (6–60 mg P kg�1) were

obtained by applying different P fertilizer rates in plots

with different initial soil P levels (Table 3). The highest

levels of soil P were obtained in plots annually fertilized

with 22 kg P ha�1. Due to a moderately high soil residual

P in the P164 soil (Berardo and Grattone, 1997), soil P at

sowing and at ripening were higher in P164 treatments

than inP11ones, in spite ofhave not receivedany P supply

since 5 years before the experiment. In general, P fertili-

zation increased SDM, SPc and GY (Tables 3 and 4).

Indigenous mycorrhizal endophytes isolated were:

Acaulospora laevis Gerdemann & Trappe, Glomus

clarum Nicol. & Schenck, Glomus etunicatum Becker

& Gerdemann, Glomus microaggregatum Koske,

Gemma & Olexia, Glomus mosseae (Nicol. & Gerd.)

Gerdeman and Trappe and Gigaspora margarita Becker

& Hall.

Mycorrhizal infection intensity and arbuscule con-

tent were closely correlated (r2 = 0.89). Thus, we

present mycorrhizal infection intensity as indicator of

AMC. AMC was established during both years before

tillering, increased to a peak after anthesis for all

treatments, and tended to decrease by ripening,

particularly in P-supplied treatments (Table 3). The

highest AMC was obtained in plants with no annual P

supply. Plants that received 22 kg P ha�1 annually

showed the lowest AMC.

3.2. Glasshouse experiment

The highest soil P content, RDM and AMC were

found in the upper 20 cm depth of pots of all treatments.

These response parameters were affected by P fertiliza-

tion in the 0–20 cm depth, but there was no effect of P

below 20 cm depth (P-Bray = 8.28 � 1.1 mg kg�1;

AMC = 28.7 � 2%; mean values of all samplings and

treatments). Available soil P ranged from 10 to

16 mg P kg�1 (Table 5) in the upper 20 cm depth of

pots, and it was highest in P-supplied pots. The AMC in

the entire root system ranged from 11% to approximately

52%. P fertilization increased (in most cases) SDM,

F. Covacevich et al. / Applied Soil Ecology 35 (2007) 1–9 5

Table 4

The effect of phosphorus (P) fertilization on grain yield (GY) of spring

wheat growing in the field during 2 years (field experiment 1 and 2,

respectively)

Treatment Grain yield (kg ha�1)

Field experiment 1 Field experiment 2

P0 1732 c 3214 c

P11 2638 b 4486 b

P22 3478 a 5173 a

P164 2837 ab 3941 b

LSD 651 567

Different letters in the same column show significant differences at

P < 0.05. For abbreviations, see Table 3.

Table 5

The effects of phosphorus (P) and nitrogen (N) application treatments on soil Bray-P (soil-P, 0–20 cm depth), shoot dry matter (SDM), shoot P

concentration (SPc), root dry matter (RDM), arbuscular mycorrhizal colonization (AMC) and arbuscule content (A) at the entire root system of

glasshouse-grown spring wheat. DAP: days after planting

DAP Treatment Soil-P (mg kg�1) SDM (g pot�1) SPc (g kg�1) RDM (%) AMC A

35 P0 N0 11.4 b 0.29 b 0.013 c 0.07 b 42.90 a 41.15 a

P0 N150 11.6 b 0.38 b 0.013 c 0.09 b 51.60 a 37.30 a

P20 N0 15.6 a 0.94 a 0.021 b 0.20 ab 18.10 b 4.55 b

P20 N150 15.1 a 1.41 a 0.030 a 0.28 a 11.10 b 3.95 b

LSD 1.58 0.50 0.004 0.14 20.64 12.27

52 P0 N0 10.6 b 2.58 b 0.024 a 0.18 bc 42.20 a 22.40 a

P0 N150 11.0 b 2.13 b 0.028 a 0.13 c 41.55 a 23.35 a

P20 N0 15.8 a 4.29 a 0.021 b 0.29 a 20.80 b 11.85 b

P20 N150 15.4 a 4.74 a 0.028 a 0.26 ab 26.75 b 7.80 b

LSD 1.97 0.81 0.006 0.08 13.46 7.74

60 P0 N0 11.1 b 3.86 b 0.015 b 0.14 ab 27.96 ab 16.50 ab

P0 N150 12.2 b 2.66 b 0.017 ab 0.10 b 31.11 a 19.50 a

P20 N0 15.2 a 6.46 a 0.017 ab 0.22 a 19.85 b 7.50 c

P20 N150 15.9 a 6.50 a 0.022 a 0.20 a 22.71 ab 12.00 bc

LSD 2.43 1.48 0.005 0.09 10.13 7.43

At each DAP, different letters in the same column show significant differences at P < 0.05

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RDM and SPc (Table 5), but depressed AMC. N

fertilization did not increase growth or affect AMC.

3.3. Relationships between soil and plant P status,

growth and mycorrhizal colonization

There were significant and positive correlations

between SDM, GY and available soil P at field and

glasshouse experiments (r2 up to 0.58). Relative SDM

obtained in the field experiments on the different

sampling dates correlated well with soil P at 0–20 cm

depth of soil profile but was not associated significantly

with available soil P below 20 cm depth. The relationship

was positive up to a threshold of 15.5 mg P kg soil�1,

where the highest relative SDM was reached (Fig. 1).

After that threshold, soil available P did not increase

SDM. In addition, relative GYincreased with soil P (field

experiment 1, r = 0.76 P = 0.004; field experiment 2,

r = 0.81 P: 0.002).

Available soil P and AMC obtained both in the field

and glasshouse experiments at all sampling dates were

analyzed together. As we noted above, P fertilization did

not affect soil P or AMC below 20 cm depth of pots at the

glasshouse experiment. The AMC at 0–20 cm depth of

soil was negatively correlated with available soil P at the

same depth (r = �0.82, P = 0.01). The AMC was well

related to local (0–20 cm depth) current available soil P

through a unique linear-plateau function (Fit Std. Error

11.94, F-value: 23.95) for fertilized and unfertilized

plants grown both in field and in glasshouse (Fig. 2A) and

in all phenological stages tested (Fig. 2B). Over the range

of 0–27 mg P kg soil�1, an increase of 1 mg P kg soil�1

lead in AMC decreases of 2.6%. Above

27 mg P kg soil�1 (Std. Error = 3.48, T-value = 7.64),

soil P did not affect AMC, which remained stable at an

average value of 10%. Residuals from the relationship

between soil P and AMC were not accounted by SPc

(r = 0.08, P = 0.45). When P content of shoots and AMC

obtained both in the field and glasshouse experiments at

all sampling dates were analyzed together, no association

between SPc and AMC was detected (Insert in Fig. 2A,

P = 0.21).

4. Discussion

The combination of field and glasshouse experi-

ments allowed us to grow wheat plants in a wide range

of soil available P and to investigate AMC across a wide

range of soil and plant P status. The glasshouse

experiment demonstrated the local effect of P fertiliza-

tion on soil P and AMC dynamics. Fertilization

increased soil available P, plant growth and SPc.

Moreover, the levels of SPc we observed were in

accordance with those reported by Karlen and Whitney

(1980) for wheat.

Seasonal patterns of mycorrhizal colonization

observed in this study were similar to those reported

by Dodd and Jeffries (1986) and Mohammad et al.

(1998). Low infection was present following sowing in

the fall months, which are characterized by low soil

temperatures. During the spring AMC increased

F. Covacevich et al. / Applied Soil Ecology 35 (2007) 1–96

Fig. 1. Relationship between relative shoot dry matter (RSDM) and

soil Bray-P in field-grown spring wheat in relation to P fertilization

levels. See Table 3 for abbreviations.

Fig. 2. Relationship between intensity of arbuscular mycorrhizal

colonization (AMC) and soil Bray-P into field and glasshouse-grown

spring wheat in relation to P fertilization levels (A) and in relation to

phenological stages (B). Empty symbols in (A) correspond to field

conditions and filled symbols correspond to glasshouse conditions.

Insert in (A): effect of shoot P content on mycorrhizal intensity in P

levels. See Tables 3 and 5 for abbreviations.

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gradually and then tended to decrease by the time of

ripening. Abbott and Robson (1991) also reported a

rapid increase in root colonization in annual crops

concurrent with active root growth, followed by a

decline when roots senesced.

Our results clearly show that the AMC of wheat roots

is modulated by current local soil available P but not by

plant P status: (i) the relationship between soil available

P and AMC at 0–20 cm depth was significant, and no

clear relationships between SPc and AMC was found;

(ii) mycorrhizal intensity below 20 cm depth was

unaffected by P fertilization. There were differences in

AMC (up to 60%) at 0–20 cm depth between

unfertilized and fertilized pots; however, plants showed

similar AMC below 20 cm depth, where soil P was

unchanged by fertilizer application, and (iii) the

relationship with soil P at 0–20 cm depth accounted

for most of the variation in AMC at this depth, and at

any crop phenological stage for plants grown in the field

or in pots in the glasshouse. The same relationship

accounted for most of the variability in AMC, in spite of

quite different climatic conditions under the two

different years that resulted in varied wheat growth

and yield between field experiments.

Our results also show that the primary effect of P on

mycorrhizal symbiosis seems to be a reduction in AMC

caused by high local soil P concentration. Two

mechanisms have been suggested to explain the

modulation of AMC by soil P: (i) by decreasing root

exudation (Kurle and Pfleger, 1996) and consequently

the spread of the symbiosis, and (ii) by a direct effect of

high P concentration on development of external AM

hyphae (Miranda and Harris, 1994). The effect of high P

fertilization level as depressor of AMC has been widely

reported in wheat (Sparling and Tinker, 1978; Buwalda

et al., 1985; Cade-Menun et al., 1991; Rubio et al.,

2003). In agreement with these reports, P fertilization

reduced AMC in our experiments. In addition, we

established a relationship between soil P and indigenous

AMC, which accounted for most of variation in

mycorrhizal intensity in wheat plants growing under

contrasting environmental and experimental conditions.

This kind of relationship was established with plants

and AM fungi growing under field and glasshouse

conditions and was not previously available in the

literature.

Available soil P content depressed indigenous AMC

in wheat until a threshold of 27 mg P kg�1. Above this

threshold AMC was stabilized at a minimum of 10%.

Both the soil P threshold and the minimum AMC values

must to be considered in planning the fertilization

regime to be applied in order to increase growth of

wheat without depressing mycorrhizae. However, the

importance of the mycorrhiza symbiosis not only

includes benefits to the plant. In spite of increases in

host plant nutrition by mycorrhiza, hyphae of AMF are

considered to be primary soil aggregators, because the

extraradical hyphae of AMF have significant biomass in

most soils (Rillig et al., 2002). A positive correlation

between AMF hyphae and aggregate stabilization in

natural systems has been described by Jastrow et al.

(1998). Piotrowski et al. (2004) concluded that soil

aggregation is a function of both the fungus and its host,

because hyphal characteristics may contribute to

aggregation ability and plants with dense, fibrous root

systems (such as grasses) assist in the aggregate

formation.

Finally, we observed that at 15.5 mg P kg�1 we

obtained the highest relative SDM, at least under field

conditions. At this soil P level, AMC was predicted to

average 39%. Future experiments must evaluate if the

15.5 mg P kg�1 level should be considered as the

highest soil P value that ensures high wheat yield with

an indigenous mycorrhizal formation that efficiently

improves wheat growth and soil stability. Simulation

models of P dynamic in soil could be used to predict the

exact fertilizer dose required to obtain a soil P level that

increases growth without depressing the indigenous

AMC. Thus, the efficient use of AMF may allow the

attainment of acceptable yield levels with the minimum

fertilizer dose, while also reducing costs and environ-

mental pollution risk. This is a promising way for

obtaining high wheat yields with low fertilizer inputs in

order to support a sustainable agriculture.

Acknowledgements

This study was made possible with financial support

of EEA INTA Balcarce-FCA UNMP (15/A160). We are

grateful to A. Berardo for the use of the experimental

field. This research was completed while F. Covacevich

was supported by a studentship from the Consejo

Nacional de Investigaciones Cientıficas y Tecnicas

(CONICET) and the results were taken from her Thesis

of Magister Scientiae (FCA-UNMP). L.A. Aguirrezabal

is member of the CONICET.

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