Agroforestry Systems

, Volume 78, Issue 2, pp 139–150

Influence of improved fallow systems and phosphorus application on arbuscular mycorrhizal fungi symbiosis in maize grown in western Kenya

Authors

    • Botany DepartmentNational Museums of Kenya
  • Bashir Jama
    • World Agroforestry Centre (ICRAF)
  • Caleb Othieno
    • Department of Soil ScienceMoi University
  • Robert Okalebo
    • Department of Soil ScienceMoi University
  • David Odee
    • Kenya Forestry Research Institute (KEFRI)
  • Joseph Machua
    • Kenya Forestry Research Institute (KEFRI)
  • Jan Jansa
    • Swiss Federal Institute of Technology (ETH) Zurich
Article

DOI: 10.1007/s10457-009-9249-3

Cite this article as:
Muchane, M.N., Jama, B., Othieno, C. et al. Agroforest Syst (2010) 78: 139. doi:10.1007/s10457-009-9249-3

Abstract

A field study was carried out on a six-year-old on-farm field trial during long-rains season (April–August) 2003 to investigate the effect of improved fallow systems and phosphorus application on arbuscular mycorrhiza fungi (AMF) symbiosis in maize. The trial comprised of maize rotated with a fast growing leguminous Crotalariagrahamiana fallow and a non-leguminous Tithonia diversifolia fallow for 3 years followed by continuous maize. The experiment was randomized complete block design with three cropping (continuous maize, Crotalaria fallow and Tithonia fallow) systems and two phosphorus levels (0 and 50 kg P/ha). AMF colonization in maize roots, maize yield and macro-nutrients uptake were recorded. Phosphorus applications improved (P < 0.05) early (<8 weeks old maize) AMF colonization, nutrient uptake and maize yield in improved fallow systems. Greater differences due to phosphorus application were noted in maize in Tithonia fallow than in Crotalaria fallow. Following phosphorus application, a positive relationship existed between early AMF colonization and maize yield (r = 0.38), and phosphorus and nitrogen uptake (r = 0.40 and r = 0.43, respectively), demonstrating the importance of phosphorus fertilization in enhancing low-input technologies (improved fallows systems) in phosphorus deficient and acidic soils of western Kenya.

Keywords

Arbuscular mycorrhiza fungiCrotalaria fallowContinuous maizePhosphorusTithonia fallow

Introduction

Phosphorus (P) and nitrogen (N) deficiencies, and acidic soil conditions are common and widespread in western Kenya, where maize is grown as the main staple crop (Sanchez et al. 1997). Besides inherent soil properties and limited organic matter inputs, low fertilizer inputs cause early apparition of deficiency of P and other nutrients in this region. Due to soil acidity, fine texture, and high amounts of reactive iron, a great portion of added P is strongly sorbed and not available to the crops. This is further aggravated by extreme poverty widespread in this area. To replenish nutrients taken off the soil by crops, improved fallow systems, a low cost technology consisting of planted legumes or other (non-leguminous) species, are currently being adopted for rapid replenishment of soil fertility by majority of small-scale farmers (Sanchez 1999). Improved fallow systems have considerable potential to alleviate N deficiencies through biological nitrogen fixation and the retrieval of inorganic N from subsoil layers (Harteminkv et al. 1996; Mekonnen et al. 1997; Gathumbi et al. 2002), and can also increase the availability of soil P. However they cannot fully eliminate P constraints to crop production especially on severely P deficient and acidic soils (Buresh 1999). The amount of P in soil organic matter fractions has been shown to decrease during subsequent cultivation of maize (Maroko et al. 1999), necessitating the regular inclusion of one season fallow into the crop rotation. Phosphorus recapitulation is therefore vital to improve plant growth and crop yield especially where several seasons of crop cultivation are expected after the last improved fallow systems.

Management of arbuscular mycorrhizal fungi (AMF) and replenishment of P stocks via addition of inorganic fertilizer is considered necessary if soils are seriously depleted after decades of cropping without P inputs (Borie et al. 2002). AMF play an important role in P uptake and growth of many cereals, legumes, and other crop plants (Sieverding 1990; George et al. 1995). This process of enhancing P absorption by plants is particularly important in highly weathered, fine textured, and acid tropical soils, where great proportions of applied P fertilizer are not available to plants due to strong fixation of P on iron and aluminium oxides (Jama et al. 1997; Bünemann et al. 2004c). However, the formation and function of AMF symbiosis in crops has been shown to be affected by agricultural practices such crop rotation and P fertilization through the ability of proceeding crops to associate with indigenous AMF, and these effects are usually magnified under conditions of P limitation (Thompson 1991; Thompson 1994; Singer and Cox 1998; Karasawa et al. 2000; Karasawa et al. 2001). This is particularly important in P nutrition and growth at early developmental stages of maize through their effects on mycorrhiza development and functioning (Miller 2000; Evans and Miller 1998; McGonigle et al. 1990; Miller et al. 1995).

Recent studies in a Kenyan ferrasol soils reported a shift in AMF community composition in maize-crotalaria fallow rotation compared to maize monocropping, with higher spore abundances of Acaulospora and Scutellospora species recorded in the maize-crotalaria fallow rotation (Mathimaran et al. 2007). It is hypothesized that greater AMF diversity would result in more AMF function to crops, since the mycorrhizal community will span broader range of functions (Koide 2000). However, there is limited information to show whether a shift in AMF community would influence AMF symbiosis in maize, and how the latter relates to maize nutrient uptake, growth and productivity in the context of tropical agriculture in sub-Saharan Africa, and how P application would influence the AMF symbiosis in P deficient soil. This study was undertaken to assess (1) the influences of improved fallow systems on AMF communities and colonization in maize, (2) the influences of P application on the development of AMF colonization, and (3) importance of AMF in mineral nutrition and productivity of maize. We hypothesized that improved fallow systems (Crotalaria and Tithonia fallow) and P applications would modify the AMF communities, hence improve AMF colonization in maize and subsequent maize productivity and nutrient uptake.

Materials and methods

Study site

The study was conducted during long rains season (April–August) 2003 in Central Kisa, Butere-Mumias District, western Kenya (0°09′N, 34°33′E) at an elevation of 1,485 m. Mean annual rainfall in the area is estimated at 1,705 mm with an average monthly potential evaporation above 150 mm (Jaetzold and Schmidt 1982). There are two growing seasons, the long rainy seasons (LR) between March and August (with rainfall peaks between March and May) and the less reliable short rainy season (SR) from September through February (with rainfall peaks between October and November). The soil at this site is classified as a kaolinitic, isohyperthermic Kandiudalfic Eutrudox (USDA classification) or a Ferralsol (FAO) with 39% clay and 37% sand in the top 15 cm layer. The region has high agricultural potential, but soil nutrient depletion (particularly N and P) is a major limiting factor for crop production (Shepherd et al. 1996). In addition, the area is highly populated, the farm size per household is on average less than 0.25 ha, and the soils are continually cropped with little or no inputs.

Site history and experimental design

The study was conducted on a 6-year old on-farm field experiment established in 1997. The experiment was laid out as a randomized complete block design with four replicate blocks in March 1997 with three crop rotations (continuous maize, maize-crotalaria fallow rotation with Crotalaria grahamiana as improved fallow species and maize-tithonia fallow rotation with Tithonia diversifolia as improved fallow species) and two levels of P fertilization (0 and 50 kg P ha−1 yr−1), applied as triple super-phosphate (TSP). Plot size was 6 × 6 m. Prior to this study, maize was grown in rotation with crotalaria and tithonia fallow systems for 3 years (April 1997 to March 2000) at two levels of P fertilization (0 and 50 kg P ha−1 yr−1), applied as TSP at the beginning of the LR seasons (Table 1). Thereafter, the land was cropped continuously with maize with no inputs for 3 years (March 2000–2003) in all cropping systems to test the residual effect of the fallow species with or without P applications (Table 1). The current study was carried out during LR season (March–August, 2003). The soil was manually tilled down to 15 cm at the beginning of the LR seasons. During this season 50 kg P/ha of TSP was incorporated into the top 2 cm in all the plots which had received P fertilizers earlier. At the same time, all plots received potassium chloride (KCl) at the level of 100 kg K/ha. Maize was sown mid April 2003 at 0.75 × 0.25 m spacing, and harvested from all plots end of August 2003.
Table 1

Overview of fallow and fertilization treatments applied in the field experiment between 1997 and 2003

Year

1997

1998

1999

2000

2001

2002

2003

Season

LR

SR

LR

SR

LR

SR

LR

SR

LR

SR

LR

SR

LR

Plant cover

M

M/F

M

M/F

M

M/F

M

M

M

M

M

M

M

P input (TSP)

±P

−P

±P

−P

±P

−P

±P

−P

−P

−P

−P

−P

±P

In 1997 through 1999, maize was grown in rotation with the improved fallows and phosphorus (P) applications were made in all long rains (LR) during this period. In LR 2000, the last fallow was incorporated and last P application was made. From short rains (SR) 2000, no inputs were made and maize was grown continuously till the onset of this study in LR 2003, when P was applied again after six season of continuous maize growth

M, maize; F, improved fallow; TSP, triple super-phosphate

Soil sampling and isolation of AMF spores

To quantify the size and composition of AMF spore populations in the experimental site, soil was sampled at the beginning of and at the end of LR 2003 (in April and August 2003, respectively). Ten individual soil cores were taken from the upper 30 cm layer randomly from each replicate plot, pooled and mixed to obtain a representative sample per plot. Spores were isolated from 50 g of soil using wet sieving and decanting method (Mason and Ingleby 1998). Fifty grams of soil sample was weighed and then mixed in water by stirring thoroughly before decanting through 710- and 45-μm sieves. This process was repeated four times. Later, the sediments collected on the 45 μm sieve were washed into 50 ml centrifuge tubes and centrifuged for 5 min at 1,750 rpm. Water from the tubes was decanted to discard floating debris. Sucrose (48%, w:v) was added to the tubes, mixed thoroughly before centrifuging for 15 s at 1,750 rpm. Immediately after centrifugation, sucrose solution was decanted through 45 μm sieve. The spores retained on the sieve were rinsed thoroughly with water to wash out the sucrose and later transferred into a petri dish. Healthy spores were counted under microscope at ×400 magnification. The spores were examined microscopically and identified down to the genus level according to the description by Morton (1988).

Soil sampling and analysis of nutrient levels in soil

Ten individual soil cores were taken from the upper 15 cm layer randomly from each replicate plot, pooled and mixed to obtain a representative sample per plot in the beginning of the LR season before planting the maize. A composite sample from ten randomly selected grid points was taken for analysis. Thereafter, the samples were air dried, ground and sieved through 2 mm sieve.

Total N, extractable P, exchangeable K, Ca and Mg in oven dried soil (40°C for 48 h) was determined by elemental analysis described in ICRAF Laboratory Manual (ICRAF 2000). Total N was extracted by digestion and estimated by colorimetry. Extractable soil P and exchangeable K were estimated using Olsen extraction method. P concentration in the extracts was estimated colorimetrically, and the K concentration was estimated by flame photometry. Exchangeable Ca and Mg were extracted using 1N KCl and determined by atomic absorption spectroscopy. Soil pH was measured in aqueous suspension (1:2.5 w:v).

Maize root sampling and assessment of AMF colonization

Ten individual soil cores containing soil and roots were collected randomly from each plot every 2 weeks from the uppermost 30 cm layer, 5 cm away from the maize plant by using a soil auger (ø3 cm). The samples from each plot were then pooled and mixed thoroughly to obtain a composite sample. Maize roots obtained from the soil cores were washed free of soil by first soaking in a bucket of water, and later wet sieving (one mm mesh size) with tap water. Roots were separated from organic debris by hand. A sub-sample of maize roots from the field soil were cut into one cm segments, and stained using the modified procedure for staining roots to detect AMF structures as described by Mason and Ingleby (1998). Briefly, the roots were cleared in 2.5% KOH in autoclave for 15 min at a temperature of 121°C and were later bleached in a mixture of 30% hydrogen peroxide and 30% ammonium solution (1:1 v:v) for 30 min to remove phenolic compounds from the roots. The roots were then acidified for 2 h with 1% HCl and stained with 0.05% acidified trypan blue dissolved in glycerol–water (1:1 v:v) by autoclaving the roots in this solution for 3 min at 121°C.

Estimation of the AMF colonization in the roots was done according to Trouvelot et al. (1986). Thirty root fragments were mounted on two slides each containing 15 root fragments. The fragments were observed under the microscope (magnification 160–400×) for the presence of hyphae, arbuscules and vesicles and rated from 1 to 5 for hyphal colonization and from 1 to 3 for arbuscular colonization. Mycocalc program (www.dijon.inra.fr/mychintec/Mycocalc-prg/download.html) was then used to calculate root length colonized by AMF (% RLC) as well as root length colonized by arbuscules.

Sampling of maize and analysis of nutrients in plant biomass

At harvest (5 months after sowing), maize ears and stovers from each plot were weighed to get fresh weights. Total aboveground dry weights (ears, grains and stover) were determined after drying the sub-samples at 70°C till constant weight. The dry materials were ground to pass through 20 mm mesh and analyzed for nutrient (total P, N, K, Ca and Mg) contents using standard methods described in ICRAF Laboratory Manual (ICRAF 2000).

Gain/losses due to 50 kg P/ha applications

To separate the effect of previous P application and 50 kg P/ha applications made during LR season we calculated maize yield, mineral nutrition and AMF colonization gain or loss using short rains (2002; September 2002 to February 2003) data when no additional fertilization was made to previous P plots. We assumed that, if the effect of previous P observed during SR season was to be observed in LR season, hence the gains/losses recorded over the control plots (50 kg P/ha plot minus 0 kg P/ha) after removing the effect of previous P application in SR season, is the gain/losses resulting from 50 kg P/ha application during LR.

Data analysis

All data were tested for normality and analyzed by two- and one-way analyses of variance (ANOVAs) using GenStat (9th edition). Percentage data and count data were arcsine and square-root transformed respectively prior to analysis. Treatment means were separated by the Duncan multiple range test at P < 0.05. Data on root colonization by AMF, plant nutrient concentrations and maize yield were subjected to correlation analysis. Mean ± standard errors of means (SE) are reported unless specified otherwise.

Results

Soil site characteristics

Soil pH in the study site prior to planting of the experimental maize crop was low (pH in water ranging from 4.9 to 5.0) with slightly but significantly lower pH (and higher exchangeable acidity) in soils under continuous maize cropping than in those previously under fallows. Soil pH was affected neither by P fertilization nor by the interaction of cropping system with P fertilization (Table 2). Available P levels in this soil were low, values ranging between 0.28 and 5.6 mg P kg−1.
Table 2

Residual effect of fallow systems (maize rotated with Tithonia fallow, TD; maize with crotalaria fallow, CG) as compared to continuous maize (CM) and P fertilization (+P) as compared to unfertilized soil (−P) on soil pH, exchangeable acidity (EA), exchangeable cation concentrations (Ca, Mg and K), Olsen P, and total C and N levels at the depth of 0–15 cm in April 2003. Treatment means are shown along with results of two-way ANOVA

Treatment

Nutrient in the soil

PH (Cmol/kg)

EA (Cmol/kg)

Ca (Cmol/kg)

Mg (Cmol/kg)

K (Cmol/kg)

P (mg/kg)

C (%)

N (%)

Cropping systems

    CM

4.90a

18.72a

15.98b

4.88b

0.13b

4.03a

2.19b

0.22b

    CG

5.03a

15.08a

21.76a

6.58a

0.16a

4.16a

2.40a

0.24a

    TD

5.04a

14.70a

19.79ab

6.86a

0.17a

4.25a

2.36a

0.23ab

    SED

0.06

2.22

2.12

0.75

0.01

0.54

0.04

0.01

P fertilizer

    50 kg P/ha

4.99a

16.22a

19.73a

6.19a

0.14b

5.60a

2.32a

0.23a

    0 kg P/ha

4.98a

16.11a

18.63a

6.02a

0.17a

2.69b

2.31a

0.23a

    SED

0.02

0.69

1.08

0.37

0.01

0.39

0.04

0.003

Cropping system (A)

NS

NS

*

*

***

NS

***

**

P fertilization (B)

NS

NS

NS

NS

***

***

NS

NS

A × B

NS

NS

NS

NS

NS

NS

NS

NS

+P, 50 kg/ha applied annually as triple super-phosphate between 1997 and 2000

*** P < 0.001; ** 0.001 ≤ P < 0.01; * 0.01 ≤ P < 0.05; NS, P ≥ 0.05. Means followed by the same letters in the same parameters are not significantly different at P < 0.05

Plots previously fertilized with P recorded higher levels of available P compared to those without previous P applications in the all the three cropping systems (P < 0.001, Table 2). Levels of total C in the soil were significantly higher in both crotalaria and tithonia fallows than in continuous maize treatment, whereas only the soil previously under crotalaria fallow showed higher total N levels than soil under continuous maize. The soil previously under tithonia showed no significant differences in N levels from the other cropping systems. Levels of exchangeable Ca and Mg were roughly the same in all cropping systems (continuous maize and both rotation fallows) with or without P application, with lower levels under continuous maize as compared to the fallow treatments (Table 2), which was consistent with previous results from 2000 (Bünemann et al. 2004a). Likewise, the levels of exchangeable K were higher in soils previously under fallow.

AMF spore communities

AMF communities were surveyed directly on spores isolated from the field soil, and identification of the AMF was based on observation of spore morphologies. Total number of spores per unit weight of soil was affected neither by cropping system nor by P fertilization nor by their interaction before the LR 2003, but the total number of spores was marginally lower (P = 0.05) in P fertilized soils after the LR 2003. Four AMF genera (Scutellospora, Gigaspora, Acaulospora, and Glomus) were recorded in all cropping system, with the Glomus and Scutellospora being most abundant spore types. Fewer Scutellospora spores were found in P-fertilized than unfertilized soils after the LR 2003 (P = 0.020), with some marginally significant variation between the cropping systems (interaction between cropping systems and P fertilization P = 0.05). The decrease in Scutellospora abundance was significant in continuous maize and in previous crotalaria fallow, but was not seen in the previous Tithonia fallow treatment. In addition, marginally fewer spores of Glomus were recorded in P fertilized soils after the LR 2003 as compared to the unfertilized soils (P = 0.074). Significantly more Gigaspora spores were also found in soils previously under crotalaria fallow than under continuous maize at the beginning of LR 2003 (P = 0.017), whereas no differences in Gigaspora spore abundance due to any treatments were recorded at the end LR 2003. No differences due to any factor were observed for Acaulospora spore abundances.

AMF colonization

In this study we divided the maize growth period into three stages i.e. early (till 8 weeks after sowing), mid (8–12 weeks after sowing) and late (12–16 weeks after sowing) depending on the AMF colonization differences. Two way ANOVA indicated strong positive effect of P fertilization on AMF colonization of roots by both hyphae and arbuscules (P = 0.028 and P = 0.002, respectively) during early growth of maize (Fig. 1). No significant effects could be attributed, however, to either the cropping systems or the interaction of the two main factors. However, a trend was observed towards somehow higher root colonization in previously fallowed and fertilized soil compared to continuous maize fertilized treatment (Fig. 2). During mid period of maize growth (8–12 weeks after sowing), the proportion of maize roots colonized by AMF stagnated which resulted in disappearance of the P fertilization effect observed during early maize growth period (Figs. 1, 2). There was a general AMF colonization trend peaking in week 12, and declining thereafter in all the cropping systems at later stages of maize growth (Fig. 2).
https://static-content.springer.com/image/art%3A10.1007%2Fs10457-009-9249-3/MediaObjects/10457_2009_9249_Fig1_HTML.gif
Fig. 1

The trends of AMF colonization; total AMF colonization (a) and colonization by arbuscule (b) of maize roots between April and August 2003 under different phosphorus levels (50 and 0 kg P/ha) annual addition of 50 kg P/ha in Butere Mumias, Western Kenya. Errors bars represent ±1 standard error of mean difference. Where, % RLA is percentage root length with arbuscules, % RLC is total percentage AMF colonization, 50 P—50 kg P/ha and 0 P—0 kg P/ha

https://static-content.springer.com/image/art%3A10.1007%2Fs10457-009-9249-3/MediaObjects/10457_2009_9249_Fig2_HTML.gif
Fig. 2

The trends of AMF colonization of maize roots between April and August 2003 under different cropping systems (continuous maize, CM; Crotalaria fallow, CG and Tithonia fallow, TD) with (+P) or without (−P) annual addition of 50 kg P/ha. Errors bars represent ±1 standard error of mean difference

Maize yield and acquisition of nutrients

Two-way ANOVA indicated significant (P < 0.05) influences of both cropping system and P fertilization, as well as of their interaction, on maize grain yield. Phosphorus (50 kg P/ha) applications dramatically increased (P < 0.01) the grain yield of maize (Fig. 3) and uptake of P, N, K, Ca and Mg in all the three cropping systems (Table 3), with exception of Ca, whose uptake was not different (P = 0.12) among P-fertilized plots previously under the three different cropping systems. The content of Ca was only influenced by P fertilization of soil (P = 0.006), but not by cropping system (P = 0.082) in LR season. P and N uptakes were significantly (P < 0.05) higher in P fertilized plots previously under improved fallow systems compared to plots under continuous maize.
https://static-content.springer.com/image/art%3A10.1007%2Fs10457-009-9249-3/MediaObjects/10457_2009_9249_Fig3_HTML.gif
Fig. 3

Effects of cropping systems (continuous maize, CM; Crotalaria fallow, CG and Tithonia fallow, TD) with (+P) or without (−P) P applications on maize yield in long rainy season (April and August) 2003. Error bar represents the standard error of means

Table 3

Effects of cropping systems (Continuous maize, CM; Crotalaria fallow, CG and Tithonia fallow, TD) with (+P) or without (−P) P applications on uptake of N, P, K, Ca and Mg in maize crops between April and August 2003

Treatment

Nutrient uptake (kg/ha)

N

P

K

Ca

Mg

CM+P

11.25c

1.97c

8.96b

2.99b

1.30b

CG+P

29.50b

4.76b

26.76a

8.25ab

4.81a

TD+P

41.38a

7.33a

29.94a

14.50a

5.39a

CM−P

3.82c

0.52d

3.14b

1.63b

0.44b

CG−P

9.54c

1.22d

5.61b

2.40b

0.78b

TD−P

9.74c

1.30d

7.89b

2.54b

1.44b

SED

3.69

0.70

3.83

3.58

0.88

Sources of variation

    Cropping systems (A)

***

**

**

*

***

    P Fertilization (B)

***

***

***

**

***

    A × B

***

***

**

*

*

Where CM, Continuous maize; CG, Crotalaria fallow; TD, Tithonia fallow; SED, Standard error of the difference of means

Means followed by the same letters in the same parameter are not significantly different at P < 0.05

* Significant at P < 0.05; ** Significant at P  0.01; *** Significant at P < 0.001

Residual effect versus gain/losses due to 50 kg P/ha applications

Previous P did not have any significant effect on AMF communities and AMF colonization as well as mineral nutrition (N, P, K Ca and Mg) in maize under the three cropping systems. However, previous P applications improved maize grain yield in the fallowed (Crotalaria and Tithonia) plots than in the continuous maize systems, but no effect of previous P applications was observed on maize grain yield under the two fallows (data taken during short rains 2002 and not shown). When the effect of previously P applications was removed, an increase in early AMF colonization was observed in both Tithonia and Crotalaria fallow while a decrease in early AMF colonization was recorded in the continuous maize systems (Fig. 4a). On the other hand maize grain yield increased in all the three cropping systems with Tithonia fallow systems recording the highest gain (Fig. 4b). There was an increase in mineral nutrition (P, N, Ca, and K) in Tithonia fallow system while the opposite was recorded for continuous maize. In Crotalaria fallow system, there was an increase only for Ca and K uptake (Fig. 4c). Of the 50 kg P/ha added in the soil, only 6.03 kg/ha P was harvested in maize grown in Tithonia fallow compared to Crotalaria fallow (approximately 3.54 kg/ha P). Similarly, only 1.45 kg/ha P was harvested in maize grown in continuous maize. Phosphorus fertilizer use efficiency was highest (approximately 12%) in the Tithonia fallow system, intermediate in Crotalaria fallow systems (approximately 7%) and lowest in continuous maize (1%).
https://static-content.springer.com/image/art%3A10.1007%2Fs10457-009-9249-3/MediaObjects/10457_2009_9249_Fig4_HTML.gif
Fig. 4

AMF colonization (a), grain yield (b) and nutrient uptake (c) gain/loss in maize under different cropping systems (continuous maize, CM; Crotalaria fallow, CG and Tithonia fallow, TD) following 50 kg P/ha application during LR 2003 in Butere Mumias, Western Kenya. % RLC is total percentage AMF colonization

Relationships

To determine the importance of AMF in mineral nutrition and productivity of maize, correlation analysis between AMF colonization and yield and mineral nutrition were performed. Only early AMF colonization (8 weeks after sowing) showed a statistically significantly positive relationship with maize yield and mineral nutrition. However, early AMF colonization was poorly correlated with yield (r = 0.38, P = 0.007), P (r = 0.40, P = 0.005) and N (r = 0.43, P = 0.002) uptake.

Discussion

The study investigated the impact of fallow management and P-application on AMF symbiosis (communities and colonization) and maize performance. Because of the design of the previous fallow, the P-treatment includes both residual P-effects and actual P-effects, however, the effect of P observed in this study is attributed to 50 kg P/ha applied given that no significant effect of previous P was observed on AMF communities, AMF colonization as well as on the yield and nutrient uptake in maize under the two fallows.

We hypothesized that cultivation of maize in rotation with improved fallow systems (Crotalaria and Tithonia fallow) and P application would modify the AMF communities, hence improve AMF colonization in maize, nutrient uptake and productivity of maize. Although our results revealed a trend showing that the total AMF spore abundance as well as abundance of Gigaspora before the LR 2003 was higher in soil previously under Crotalaria fallow than under continuous maize, the study, however, did not show a significant effect of fallow systems (Crotalaria and Tithonia fallows) on AMF communities and colonization in maize, 3 years after the fallow rotations ceased most likely because the study tested residual effects of fallow after several years suggesting that impact of fallow (lower disturbance) on mycorrhizal inoculum potential may not be visible after 3 years of maize cropping. Please note that Mathimaran et al. (2007) found that crotalaria-maize rotation significantly affected AMF composition, but not the density and species of AMF communities in the same site 1 year before our study. Taken together, these studies may indicate that the effect of maize-fallow rotations on AMF may persist for several cropping seasons but decline gradually with time following maize monocropping. Similarly, we did not record significant effects of the fallows on soil chemical properties except for C and N levels (where N levels were only higher in soils in the Crotalaria fallows systems), suggesting a similar declining residue effect of fallows on soil chemical properties.

Our study found no strong effect of P application on AMF communities. Only fewer Scutellospora spores were found in P-fertilized than unfertilized soils. Lack of a strong effect of P application on AMF communities was in accordance with a previous study by Mathimaran et al. (2007) in the same site. However, the generally lower Scutellospora spores in P fertilized plots than unfertilized soils in this study could have indicated the negative impact of P fertilization on AMF communities. Phosphorus fertilization have been shown to cause a shift in AMF species composition, supporting more Glomaraceae species while reducing Gigasporaceae especially Scutellospora (Treseder and Allen 2002). In spite of limited effect of P application on AMF communities, we observed strong effect of P fertilizer addition on early AMF colonization of maize. The effect of P fertilization on early AMF colonization in maize in this study was surprising. Over the past, it has repeatedly been shown that P fertilization results in low levels of root colonization by AMF (Miller et al. 1995; Kahiluoto et al. 2001; Allison and Goldberg 2002). Increases of P in soil or in plant tissues may be associated with increases, decreases or no changes in the AMF colonization in roots and, the sensitivity and the direction of change may depend on the initial level of available P (and consequently more % P in tissue) for plant growth. Treseder and Allen (2002) reported that initial nutrient status of ecosystems influences responses of AMF to fertilization, with deficient site raising the AMF biomass. If available P is deficient (or highly deficient) for plant growth (could be even greater in P-retentive soils), increases of P in plant tissue could not be sufficient to alleviate the deficiency of P, and the AMF colonization may increase (Bolan et al. 1984; Mendoza and Pagani 1997; García and Mendoza 2007). Our soil was, however, acidic and strongly P deficient. The soils prior to planting of the experimental maize crop recorded a low pH in water (ranging from 4.9 to 5.0) and available P levels generally below 6 mg P kg−1 in all the three cropping systems, which was far below the critical P level of 10 mg P kg−1 recommended by Okalebo et al. (2002). In addition previous studies (e.g. see Bünemann et al. 2004b) on the same site reported a dramatic increase of P availability and doubling in maize yields following P fertilization at levels of 50 kg P ha−1 yr−1. It is likely that under these specific conditions the potential negative P fertilization effect on AMF root colonization could have been offset by increased C supply to the fungi by more vigorously growing crops (Mathimaran et al. 2007). It has indeed been shown that application of moderate amounts of P fertilizer into P deficient soils may stimulate the levels of mycorrhizal colonization of roots and increase the magnitude of mycorrhizal benefits for the plants, following a bell-shaped curve with a maximum under low-to-moderate, but not extremely low P availabilities (Picone 2002). Since soils in the tropics are commonly depleted in available P pools (Smithson and Giller 2002), continuing removal of P with crop export (e.g. through harvesting without incorporating them back to the soil) may further aggravates the problems associated with P deficiency in our study system. Our result strongly advocates the necessity of application of moderate P fertilization levels in these soils to offset loss of fertility as earlier suggested (Buerkert et al. 2001; Bünemann et al. 2004c).

This study also found a strong effect of P fertilizations on maize grain yield and nutrient uptake (N, K, and Mg) in all the three cropping systems, with more nutrient uptake in fallowed soil. This implies that improvement of P nutrition in our study site resulted in improved acquisition of other nutrients (N, K, and Mg) in maize consequently improving maize grain yields. Further studies are required to test and elucidate why such effect was not observed for Ca.

Phosphorus fertilization improved nutrient uptake (N, P, K, and Mg) and maize grain yield more in fallowed plots than in continuous maize systems. It was also amazing to note that, when the effect of previous P was removed 50 kg P/ha applications did not improve uptake of P and other nutrients (N, K, Ca and Mg) in the maize under continuous maize systems. In a previous study on the same site (Bünemann et al. 2004b), maize-Crotalaria fallow rotations supported higher soil microbial activities resulting in more carbon, N, and P being bound in microbial biomass. Higher microbial activities in previously fallowed soil compared to the soils under continuous maize might explain the improved uptake of nutrients and higher yield of maize in fallowed soils observed in our study. If such beneficial microbial communities were in the soil previously under fallow, addition of P could have improved uptake of other (N, K and Mg) nutrients. Additionally the studied soil was acidic (pH in water ranging between 4 and 5), therefore, applied P could have been fixed and unavailable to plants in continuous maize fertilized treatment in comparison to plots previously under fallows, where the fixation would be partly overcome by changing soil organic matter amounts (we recorded high C levels in fallowed plots) and composition, as well as by changing microbial activities (Bünemann et al. 2004a). Application of crop residues is known to reduce P sorption by producing organic acids during decomposition that compete with P for soil sorption sites (Nziguheba et al. 1998), and has been shown to increase the available P in the soil (Gichuru et al. 2003). Our results (see Table 2) also suggests that organic matter levels were lower in continuous maize without P than in the other cropping systems. Organic matter levels could both increase mycorrhizal activity and increase P availability not only through organic acids, but also through humic acids (Weng et al. 2008) and possibly through glomalin (Cardoso and Kuyper 2006), which again would then link to mycorrhizal associations.

This study also revealed that the two improved fallow species (Tithonia—non N-fixing and Crotalaria—N-fixing tree species) significantly differed in mineral (N and P) acquisition and yield of maize following moderate P fertilization. One striking result is that of 50 kg P/ha added, only a very small portion (e.g. 157 kg/ha in continuous maize) was additionally harvested, suggesting that only 2% of P-fertilizer was used, and the rest fixed to the soil. In contrast, 12% of P-fertilizer was used in Tithonia fallow compared to 7% in Crotalaria fallow, suggesting that fallow systems enhanced uptake non-available (fixed) form of P. Given that the soil in our study area is acidic (Sanchez et al. 1997), great proportions of applied P fertilizer were not available to plants due to strong fixation of P on iron and aluminium oxides (see Bünemann et al. 2004c). It is therefore likely that the efficiency of P-fertilizer use was highest in the Tithonia fallow system. We attribute this to specific physico-chemical conditions or due to induced changes in microbial and mycorrhizal communities under the two improved fallow systems. The differences observed in early AMF colonization of roots in soil in the Tithonia fallow-fertilized compared to the continuous maize-fertilized plots, indicates potentially long-term persisting differences between the two fallow systems, which could include functional properties of indigenous AMF communities such as capacity to colonize maize roots early in the season. It has been suggested that AMF could be useful in Fe-fixing soils (ferralsols) by increasing fertilizer P use efficiency through better mining of the soil (Cardoso and Kuyper 2006). Cardoso et al. (2006) observed increased availability of P from non-labile sources by AMF in P fixing soils. Similarly, Li et al. (2006) using 32P observed over 50% of P uptake by plants being absorbed via AMF even when P was added in P fixing soils. Considering positive correlation between initial mycorrhizal colonization and P uptake and gains in mycorrhizal colonization (Fig. 4), it seems plausible that mycorrhizal management could affect the efficiency with which fertilizer-P is used. We propose future studies on monitoring the effect of different improved fallow systems on the composition of AMF communities, AMF colonization and functioning in maize nutrition at different times after incorporation of the fallows, including the seasons immediately following a fallow period. This would be a promising result for redesigning successful cropping systems for severely P-deficient soils of western Kenya, where maize is frequently grown.

The observed significant though weak positive relationship between early AMF colonization and mineral nutrition and maize yield indicates that it is probably the early AMF colonization (in addition to other factors) which can play a significant role in P and N uptakes and productivity of maize. Maize plant has high demand for P particularly at early stages of growth. Small amounts of P application were reported to result in greater P absorption during early growth period (Miller 2000). In addition, AMF develops very early in the growth of maize, and contributes significantly to early P nutrition (Gavito and Miller 1998; Miller 2000) and other less mobile nutrients such as Cu and Zn. This suggest that, maize which completes its life cycle early, can thus benefit more from early AMF colonization than delayed AMF colonization. Taking into considerations that maize yield is affected by shoot P concentration at four- to five-leaf stage, timing of the AMF colonization and P absorption in relation to the P requirements of the plants (Miller 2000), would be a prerequisite for maize growth in our study area. Contrary to the standard view that P suppresses AMF functioning, our results show beneficial effects of small doses of P fertilizer (50 kg P/ha or below) on early AMF colonization in P-fixing soils and a subsequent significant contribution to maize productivity and nutrient uptake. This result is in accordance with other studies (Bolan et al. 1984; Gavito and Miller 1998; Miller 2000).

Acknowledgments

The financial support for this study was provided by the ETH Zurich, secured through its collaboration with ICRAF. We wish thank the technical assistance from staff, colleagues and friends from the International Centre for Research in Agroforestry (ICRAF), Moi University, Kenya Forestry Research Institute (KEFRI), ETH, and National Museums of Kenya (NMK). In particular, Dr. Joyce Jefwa (NMK) for technical advice during the study, Milton (KEFRI) who assisted in generating AMF data and Mathimaran (ETH) for assisting in nutrient analysis of maize samples. We are also grateful to Richard Coe (ICRAF) for guiding in data analysis, and Omondi and his family for permission to use their farm. Lastly we are grateful to Dr. Muchane Muchai (NMK) for critically commenting on earlier version of the manuscript.

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© Springer Science+Business Media B.V. 2009