Agricultural Management Effects On Soil Microbial Dynamic Under Peanut Cropping Systems in Semi

Agricultural Management Effects on Soil Microbial Dynamic Under Peanut Cropping
Systems in Semi-Arid Soil
By
Brandon Melester, B.S.
A Thesis
In
MICROBIOLOGY
Submitted to the Graduate Faculty

of Texas Tech University in
Partial Fulfillment of
the Requirements for
the Degree of
MASTER OF SCIENCE
Approved
Dr. John C. Zak

Chair of Committee
Dr. Veronica Acosta-Martinez
Dr. Michael San Francisco
Dr. Fred Hartmeister

Dean of the Graduate School
August, 2010
Copyright 2010, Brandon Melester
Texas Tech University, Brandon Melester, August 2010
TABLE OF CONTENTS
ABSTRACT .......................................................................................................................... iv
LIST OF TABLES ............................................................................................................... vi
LIST OF FIGURES ............................................................................................................ vii
I. INTRODUCTION TO MICROBIAL DYNAMICS, MICROBIAL

EVALUATION AND AGRICULTURAL PRACTICES .......................................... 1
Introduction ......................................................................................................................... 1
Arbuscular Mycorrhiza ................................................................................................. 1
Microbial Communities ................................................................................................ 4
Microbial Biomass ........................................................................................................ 5
FAME ........................................................................................................................... 5
Soil Enzymes ................................................................................................................ 6
Tillage ........................................................................................................................... 7
Objectives and Hypothesis ............................................................................................ 8
References ......................................................................................................................... 11
II. EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID

AGROECOSYSTEM ON SOIL MICROBIAL BIOMASS AND MICROBIAL
COMPOSITION ...................................................................................................... 16
Introduction ....................................................................................................................... 16
Methods............................................................................................................................. 18
Plot Design ........................................................................................................................................ 18
Crop Management Treatments .................................................................................... 20
Sampling ..................................................................................................................... 20
Microbial Biomass Carbon ......................................................................................... 20
Microbial Community Structure ................................................................................. 21
Data Analysis .............................................................................................................. 22
ii
Results ............................................................................................................................... 23
Effects of Tillage on MBC.......................................................................................... 23
Effects of Irrigation on MBC ...................................................................................... 23
Effects of Nitrogen on MBC ....................................................................................... 24
Changes in field conditions ......................................................................................... 24
Influences between MBC, nutrients, and enzymatic activities .......................................... 25
Temporal changes in MBC ......................................................................................... 25
FAME ......................................................................................................................... 26
Discussion ......................................................................................................................... 28
Nitrogen ...................................................................................................................... 28
Tillage ......................................................................................................................... 29
Irrigation ..................................................................................................................... 30
Soil characteristics and seasonal variation .................................................................. 30
References ......................................................................................................................... 32
III. EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID

AGROECOSYSTEM ON SOIL MICROBIAL ENZYMATIC
ACTIVITY ........................................................................................................ 50
Introduction ....................................................................................................................... 50
Methods............................................................................................................................. 54
Plot Design ........................................................................................................................................ 54
Crop Management Treatments .................................................................................... 56
Sampling ..................................................................................................................... 56
Enzyme Assays ........................................................................................................... 57
Data Analysis .............................................................................................................. 57
Results ............................................................................................................................... 58
Effect of Irrigation ...................................................................................................... 58
Effects of Nitrogen ...................................................................................................... 60
Tillage Effects ............................................................................................................ 62
Combined enzymatic and nutrient effect .................................................................... 64
Effect of nutrients ....................................................................................................... 65
Correlations between enzyme activity levels .............................................................. 65
Changes in key nutrient levels .................................................................................... 65
Discussion ......................................................................................................................... 67
Effects of Irrigation ..................................................................................................... 67
Response to nitrogen application ................................................................................ 68
Response to tillage type .............................................................................................. 69
Influence of nutrient and other enzymes ..................................................................... 70
Conclusion ........................................................................................................................ 70
References ......................................................................................................................... 73
iii
ABSTRACT
Recent agricultural practices have been shifting towards low-input sustainable
systems as it’s becoming more evident that the cost and negative side effects associated
with current agricultural practices are becoming increasingly too great. As much of our
current agriculture occurs in semi-arid regions of the United States increasing attention is
being focused on management practices that decrease water consumption along with
decreasing overall energy inputs.
West Texas produces more peanuts than any other area of the Unite States outside
of Georgia. Currently, over 320,000 tons are produced annually in the south plains. In
order to improve those numbers without increasing input, it will be imperative to
understand how agricultural management affects soil quality and microbial dynamics.
The soil microbial biota has tremendous influence on the overall soil quality. Soil quality
and the sustainability of agroecosystems are greatly reliant on the activity and diversity of
the soil microbial community and the processes they facilitate. Understanding the
interactions between management practices and the soil microbiota will be a key
component in developing the best system to maximize crop productivity while decreasing
soil inputs.
The effects of tillage, nitrogen, and irrigation management within a peanut
cropping system in West Texas were examined over a two year study to determine the
effects they had on structural and functional microbial dynamics. Microbial Biomass
Carbon and Fatty Acid methyl Ester analysis were used to examine how these
managements practices affected the soil microbe structure and composition. Enzyme
iv
assays of phosphodiesterase, β-glucosaminidase, β-glucosidase, alkaline phosphatase, and
α-galactosidase were made over the two year period to determine the effect of
agricultural management on soil microbe functioning.
Tillage was not shown to have any effect on the structural or functional microbial
dynamics. It was concluded that strip-tillage would be ideal for these systems due to
lower cost and less disturbance while producing the same benefit. Full Irrigation
significantly increases β-glucosidase activity, bacterial abundance, and MBC. Irrigation
reductions were shown to produce “hangover effects” in which microbial activity,
biomass and abundance not only decreased during the irrigation reduction but were also
lower in subsequent seasons. Nitrogen had differential effects on microbial dynamics.
The results indicated that while there was no significant effect on microbial biomass
carbon, nitrogen addition did have differential effects on enzymatic activity. Reduction
in bacterial abundance was shown to be associated with nitrogen addition. These results
indicate that crop management decisions will have consequences that either maintain the
activity and diversity of the soil microbial community or alter composition and activity
such that great inputs of energy are needed to compensate for the loss of microbial
dynamics.
v
LIST OF TABLES
2.1. Correlation matrix comparing soil extractable nutrients to Microbial Biomass

Carbon ......................................................................................................................... 35
2.2. Correlation matrix comparing soil extracellular enzymes to Microbial Biomass

Carbon ......................................................................................................................... 35
3.1. P-values for comparisons of the three different irrigation treatments effects on
soil extracellular enzyme activity ............................................................................. 76
3.2. Correlation matrix between interactions of soil enzymes and soil nutrients ............... 77
3.3. Correlation matrix between all extracellular soil enzymes effects on each other ........ 78
vi
LIST OF FIGURES
2.1. Plot design of study site (2007) ....................................................................................................36
2.2. The effect of strip tillage compared to conventional tillage on Microbial Biomass
Carbon (a) 2007-2008 (b) Seasons ............................................................................. 37
2.3. The effect of late season drought compared to full irrigation on Microbial Biomass
Carbon (a) 2007-2008 (b) Seasons ............................................................................ 38
2.4. The effect of all irrigation levels on Microbial Biomass Carbon

(a) 2008 (b) Seasons................................................................................................... 39
2.5. The effect of nitrogen on Microbial Biomass Carbon

(a) 2007 (b) Seasons................................................................................................... 40
2.6. Monthly changes in nutrient dynamics and microbial functionality

(a) Discriminate function analysis .............................................................................. 41
2.6. Monthly changes in nutrient dynamics and microbial functionality

(b) Vector correlations ................................................................................................ 42
2.7. Changes in Microbial Biomass Carbon across growing seasons 2007-2008

(a) Seasonal (b) Monthly (c) Extended Months ......................................................... 43
2.8. Relative bacterial and fungal abundances

(a) Seasonal totals for bacteria and fungi abundance (b) Seasonal Fungi:Bacteria
ratio ............................................................................................................................. 44
2.9. Relative bacteria and fungi groups abundances

(a) Fungal groups (b) Bacterial groups ...................................................................... 45
2.10. Effects of nitrogen on relative bacteria abundances

(a) 2007 (b) Seasonal ................................................................................................. 46
2.11. Effects of nitrogen on relative fungi abundance

(a) 2007 (b) Seasonal ................................................................................................. 47
2.12. Effects of irrigation on relative Gram-positive bacteria abundance

(a) 2007-2008 (b) Seasonal ........................................................................................ 48
2.13. Effects of irrigation on relative Gram-negative bacteria abundance

(a) 2007-2008 (b) Seasonal ....................................................................................... 49
vii
3.1. Study site (2007) ...............................................................................................................................79
3.2. Effects of irrigation of phosphodiesterase activity

(a) 2007-2008 (b) Seasonal ..........................................................................................................80
3.3. Effect of irrigation on β-glucosaminidase activity

(a) 2007-2008 (b) Seasonal .................................................................................................................. 81
3.4. Effects of irrigation on β-glucosidase activity

(a) 2007-2008 (b) Seasonal .................................................................................................................. 82
3.5. Effects of irrigation on alkaline phosphatase activity

(a) 2007-2008 (b) Seasonal ..........................................................................................................83
3.6. Effects of irrigation on α-galactosidase activity

(a) 2007-2008 (b) Seasonal ................................................................................................................... 84
3.7. Comparison of 3 irrigation levels on collective enzyme activity

(a) Discriminate function analysis (b) Vector correlations .................................................85
3.8. Discriminate function analysis on effect of nitrogen on collective enzyme activity.....86
3.9. Effect of nitrogen on phosphodiesterase activity

(a) 2007 (b) Seasonal .....................................................................................................................87
3.10. Effect of nitrogen on β-glucosaminidase activity

(a) 2007 (b) Seasonal ................................................................................................. 88
3.11. Effect of nitrogen on β-glucosidase activity

(a) 2007 (b) Seasonal ................................................................................................. 89
3.12. Effect of nitrogen on alkaline phosphatase activity

(a) 2007 (b) Seasonal ................................................................................................. 90
3.13. Effect of nitrogen on α-galactosidase activity

(a) 2007 (b) Seasonal ................................................................................................. 91
3.14. Seasonal nitrogen effect on collective enzyme activity

(a) Discriminate function analysis ...............................................................................................92
3.14. Seasonal nitrogen effect on collective enzyme activity

(b) Vector correlations ....................................................................................................................93
viii
3.15. Effect of tillage on phosphodiesterase activity

(a) 2007-2008 (b) Seasonal ..........................................................................................................94
3.16. Effect of tillage on β-glucosaminidase activity

(a) 2007-2008 (b) Seasonal ................................................................................................................... 95
3.17. Effect of tillage on β-glucosidase activity

(a) 2007-2008 (b) Seasonal ................................................................................................................... 96
3.18. Effect of tillage on alkaline phosphatase activity

(a) 2007-2008 (b) Seasonal ..........................................................................................................97
3.19. Effect of tillage on α-galactosidase activity

(a) 2007-2008 (b) Seasonal ................................................................................................................... 98
3.20. Yearly tillage effect on collective enzyme activity and nutrient dynamics
(a) Discriminate function analysis ...............................................................................................99
3.20. Yearly tillage effect on collective enzyme activity and nutrient dynamics
(b) Vector correlations .................................................................................................................. 100
3.21. Season variation in soil extractable nitrate .............................................................. 101
3.22. Seasonal variation in soil extractable ammonium ................................................. 102
3.23. Season variation in soil phosphorus ........................................................................ 103
3.24. Seasonal variation in soil organic matter .............................................................. 104
ix
CHAPTER 1
INTRODUCTION TO SOIL MICROBIAL DYNAMICS, MICROBIAL

EVALUATION METHODS AND AGRICULTURAL PRACTICES
Introduction
The interactions between management practices and the soil biota and subsequent
effects on the performance of crops are of great agricultural importance. Moreover
there is a large potential for increased productivity in understanding these interactions
and managing them effectively (Watt et al. 2006). However, despite the important roles
of the soil microbiota in agroecosystem functioning (e.g.,Verhoef and Brussaard 1990)
very little is known of their activities, diversities, composition, and abundance under
semi-arid conditions in a peanut-based cropping systems (e.g., Acosta-Martinez et al.
2003). Soil quality and the sustainability of agroecosystems are reliant on the activity
and diversity of the soil microbial community and the processes they facilitate (Pankhurst
et al. 1996). Soil microbes are sensitive to disturbances from management decisions and
will alter activity rates, biomass, and community structure accordingly to the severity and
duration of the disturbance (e.g., Bossio et al. 1998, Schloter et al. 2003). Understanding
how to manipulate environmental conditions to fully utilize the microbial potential will
be invaluable in moving forward in developing more sustainable agroecosystems.
Arbuscular Mycorrhiza
Microorganisms are vital to all terrestrial ecosystems by performing essential
functions such as decomposition, mineralization and nutrient transfer to plants (Prescott
1
et al. 2005). Nitrogen and phosphorous are two essential macro-nutrients that
microorganisms use to complete key steps in cycling and uptake (Cardon and Gage
2006). Plant productivity in natural and managed ecosystems is often limited by nitrogen
and phosphorous (Chapin et al. 1986), and interactions between the soil microflora and
plants to increase nutrient availability become crucial (Richardson 2001, Biro et al.
2000).
Arbuscular mycorrhizae (AM) are the largest group of mycorrhizae (Wright 2005)
and are one of the most important symbioses in terrestrial ecosystems (Smith and Read,
1997). AM fungi (AMF) influence soil carbon fluxes as well as nutrient dynamics of
plants, soils, and the atmosphere (Treseder and Cross 2006). There are some 150 known
AMF species (Vandenkoornhuyse 2002) that form symbiotic relationships with
approximately 80-85% of all terrestrial plant species (Vandenkoornhuyse 2002) including
various cropping systems (Zak and McMichael 2001). AM fungi are key mediators
between above-ground plant biomass and available soil nutrients (Read et al. 1992).
Arbuscular mycorrhizae are intracellular symbiontes that form arbusculars, which
are intricately branched haustoria (hyphal tips) in the cortex cells of a plant root (Smith
and Read, 2006).Arbuscules are the site of nutrient exchange between the plant and the
fungus. Arbuscular mycorrhiza benefit the host plant by increasing uptake of phosphorus
(P), zinc (Zn), copper (Cu), and nickel (Ni) and may increase resistance to soil-borne
pathogens, insect herbivores, and drought (Smith and Read 1997, Wright 2005).
The primary benefit of forming arbuscular mycorrhiza for most plants occurs
through the increased P uptake as phosphorus is one of the most limiting nutrients of
2
plants (Tressier and Raynal 2003). The necessity to form this symbiotic relationship with
the AM fungi arises from the morphological and physiological characteristics of root
systems combined with low levels of soil P and the immobility of P in the soil matrix
(Wright 2005). Many plant roots exhibit slow growth rates that make it extremely
difficult to react to nutrient changes in the environment, and a large number of the roots
also have insufficient branching, lateral roots, and root hairs to fulfill their P demands
(Wright 2005). Plants that form the symbiotic relationship with AM fungi depend on the
AM hyphae as an extension of the roots to access a greater nutrient pool (Leyval et al.
2002, Wright 2005). Furthermore, research has shown that mycorrhizal plants are more
effective at using organic P than nonmycorrhizal plants (Wright 2005). The AMF
mycelium receives 3-20% net photosynthate produced by host and 37-47% of carbon (C)
delivered below ground (Treseder and Cross 2006). This substantial allocation of C
demonstrates how valuable this symbiosis is to plant nutrient uptake; otherwise the plant
would not invest such a large percentage of net photosynthate to sustain the arbuscular
mycorrhizal relationship. AM fungi also increase the aggregate stability of soil reducing
wind and soil erosion (Liang et al. 2008, Wright 2005). The production of the
hydrophobic, insoluble, recalcitrant C glomalin protein by the AMF is the means by
which they are able to aggregate soil particles.
AM fungi can be increasingly important in agroecosystems to compensate for
lower overall microbial activity and nutrient cycling resulting from high input of nutrients
and disturbance from plowing (Dick 1994). Most agricultural plants form arbuscular
mycorrhizae and as a result there has been a greater effort made to understand how AM
3
fungi can improve crop productivity and reduce fertilizer inputs (Smith and Read 1997,
Zak et. al. 1998). However, there is been very little research into the establishment and
maintenance of AMF within annual agricultural ecosystems under arid and semi-arid
conditions (Smith and Read 1997, Zak and McMichael 2001).
Microbial Communities
Bacteria are estimated to be the most numerous group of microorganisms found in
soil even though they account for less than half of the microbial biomass due to their
small size (Kennedy 1999). Bacterial community composition and diversity in
agricultural systems has a key role in the cycling of nutrients within these systems. These
influences include key roles in the cycling of N, S, C, methane and other nutrients
decomposition, and compound transformation (Zak et al. 1994, Kennedy 1999). Soil
bacteria also play a large role in the formation of soil structure (Tisdall 1991). A vast
array of bacteria is present and affects agroecosytems and has the potential to be altered
due to environmental stresses and management practices (Kennedy 1999).
Soil fungi comprise the largest portion of the soil microbial biomass and in many
terrestrial systems are responsible for the majority of the ecosystem processes.
Saprophytic fungi are extremely important in soil dynamics because of their roles in
decomposition, C and N storage and sources, soil stabilization, and biochemical
processes (Bills et al. 2004, Went and Stark 1968). Fungi have been shown to be
biological control agents in some agricultural systems (Hall and Papierok 1982) and
4
important decomposers and nutrient supplier to agroecosystems (Lodge 1997, Miller and
Lodge 2007, Hietschmidt et al. 1996). Fungal abundance and diversity have been shown
to be susceptible to changes in agricultural management practices such as tillage and in
turn these changes can influence the nutrient cycles of that system (Miller and Lodge
1997).
Microbial Biomass
The microbial biomass is an important regulator of nutrient transformation and
storage of many vital nutrients including N, P, and S and is also a key component of
organic C mineralization (Nunan et. al. 1998, Horwarth and Paul 1994). Previous
research has demonstrated that microbial biomass is a sensitive indicator of differences in
sustainable cropping systems, including examining effects of tillage, crop rotations, and
soil type on nutrient turnover and organic C (Anderson and Domsch 1989, Horwath and
Paul 1994) and also considered to be the most reliable indicator of soil quality (Gil-Sotres
et al. 2004).
FAME
Fatty Acid Methyl Ester (FAME) analysis is a technique that relies on the
different fatty acids among organisms to indentify species or groups present using gas
chromatography (GC) of the volatile fatty acid methyl esters extracted from
microorganisms (Kunitsky et. al. 2006, Sasser 1990). The MIDI Sherlock Microbial
Identification System developed procedure is a cheap, quick, and reliable system that has
5
helped create libraries with over 1,500 bacterial species and over 200 species of yeast for
pure systems (Kunitsky et al. 2006). The FAME analysis has been shown to be an
effective tool in estimating of AM fungi biomass, as well as monitoring the degree to
which microbes are affected by management practices in agricultural soils (Olsson et al.
1999). FAME analysis has many advantages in describing microbial community
structure over culture-based methods in that FAME avoids selectivity toward fast
growing organisms on media. However, the technique can extract fatty acids from clay-
organic matter complexes and may include fatty acid profiles that are no longer present in
the soils in the community structure assessment (Acosta-Martinez et al. 2004). To date
16:1ω5c has been identified as an indicative fatty acid marker for AM fungi (Madan et al.
2001, Olsson et al. 1999) while 18:1ω9c, 20:1ω9c, 20:2ω6c, and 22:1ω9c have the
potential to be additional AMF fatty acid markers (Madan et al. 2001). Pankhurst et al.
1997 showed that FAME profiles can be used as an effective indicator to detecting
changes microbial compositions in response to agricultural management practices.
Soil Enzymes
Microorganisms are the main source of enzymes in the soils and thus the
composition and abundance of the soil microbial community controls soil processes (e.g.,
Acosta-Martinez et al. 2004). These microbial enzymes play vital roles in critical soil
processes such as decomposition, nutrient cycling, decomposition of xenobiotics, and soil
aggregation (Barea et al. 1996, Reynolds et al. 2003). The products from the reactions
6
from the microbial enzymes are extremely important in driving soil nutrient dynamics
(Acosta-Martinez et al. 2004).
Previous research has shown that soils from the rhizosphere region have greater
enzymatic activity than that of non-rhizosphere soils (Dick 1994) and this can be
explained by the greater species diversity associated with the rhizosphere. Roots can
excretes carbon that can stimulate microbial activity increasing nutrient cycling within
the root –region and benefitting the plant directly (Dick 1994). Different agricultural
management practices can increase or decrease the soil enzyme production (Bandick and
Dick 1999) and thereby can have significant effects on nutrient availability and soil
organic matter levels. A decrease in microbial enzyme production will result in a
decrease in mineralization and cycling of nitrogen and phosphorous necessary to sustain
productivity. (Mina et al 2001). Understanding the specific interaction between
management practices and microbial dynamics in the rhizosphere of annual cropping
systems will be important in developing techniques to maximize efficiency of
agroecosystems while reducing inputs.
Tillage
Strip-tillage (or minimal tillage) combines the benefits of conventional tillage and
no-till agriculture. The process disturbs the row but leaves the furrow undisturbed with a
complete residue cover. The undisturbed soil will retain moisture more effectively than
the disturbed soils with this moisture available to the crop plants (Musick et al. 1977). In
a semi-arid system conventional tillage provides advantages over no-till agriculture
7
through reduction in wind erosion but also can lead to soil compaction, loss of organic
matter and disruption of soil aggregates (Deng and Tabatabai 1996, Dick 1984, Doran
1980). Strip-tillage is a management technique that combines the benefits associated
with conventional tillage while reducing the negative effects. The effects of tillage will
have strong impacts on microbial diversity (Kennedy 1999). The minimal tillage
approach used in agricultural systems has been shown to increase the soil microbial
biomass and soil enzymatic activity (Dalal 1989, Spedding et al. 2004, Follett and
Schimel 1989). As agriculture shifts towards minimal tillage it will be important to know
exactly what affect it will have on the microbial dynamics of the soil.
Objectives and Hypotheses
Objective1:
Evaluate the response of the microbial biomass to tillage, irrigation, and nitrogen under
a peanut cropping system in semi-arid soils.
Hypotheses
1) Tillage, irrigation, and nitrogen fertilizer will each have significant negative
effects on the size of the microbial biomass.
As water is limiting in semi-arid environments differences in irrigation schedules
should have the largest negative impact on the size of the microbial biomass. An increase
in the microbial biomass is expected to be correlated with increases in irrigation.
2) Traditional tillage during the growing season will negatively impact the
amount of microbial biomass that develops.
Strip-tillage should have a measureable positive effect on soil microbial biomass
due to the residue cover in the furrows and a decrease in disturbance. It was expected
8
that the decrease in disturbance of soil aggregates in strip-tillage practices as compared to
conventional tillage would contribute to increased microbial biomass carbon C as the
result of keeping the fungal mycelia network intact.
3) Nitrogen application should decrease the microbial biomass by changing soil
microbial community structure. Increased soil nitrogen will allow fast growing microbes
to become dominant during period of optimum soil moisture.
Objective 2:
Evaluate the response of the microbial community structure to tillage, irrigation, and
nitrogen of a peanut cropping system in semi-arid soils.
Hypotheses
1) Irrigation will increase the bacterial contributions to microbial community
structure.
As fungi have the ability to translocate water through their mycelium and survive
at lower water potentials, irrigation should favor a shift towards a bacterial dominated
system.
2) Tillage will change the microbial community structure to a bacterial dominated
system. Soils under strip-tillage are expected to have greater relative fungi abundance
associated with them. Conventional tillage should result in more disruption on mycelial
networks and negatively affect the fungal abundances.
3) Nitrogen fertilization will alter the structure of the soil microbial community
and reduce complexity.
9
Objective 3:
Evaluate the response of microbial enzyme production to tillage, irrigation, and nitrogen
of a peanut cropping system in semi-arid soils.
Hypotheses
1) Reduction in irrigation should result in reduced microbial enzyme production
for measured enzymes.
2) As irrigation will increase microbial activity, those enzymes associated with
the bacterial assemblages should increase accordingly.
3) Conventional-tillage should have a negative effect on the enzyme activity
levels as this practice is most disruptive to the soil microbial community particularly
fungi populations through disruption of hyphae.
4) Nitrogen was expected to have negatives effect on phosphodiesterase and
alkaline phosphatase activity levels and to positively affect β-glucosamindase, β-
glucosidase, and α-galactosidase.
Phosphodiesterase and alkaline phosphatase activity levels are expected to decline
with nitrogen inputs because these enzymes are associated with P cycling and are
expected to have an inverse relationship with N. The other enzymes are involved in C
cycling and would be expected to increase as a limiting nutrient such as nitrogen is
added.
10
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15
CHAPTER 2
EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID AGROECOSYSTEM

ON SOIL MICROBIAL BIOMASS & MICROBIAL COMPOSITION
Introduction
Agro-management practices such as tillage, irrigation, herbicide and pesticide
applications, and fertilizer applications can have substantial negative impacts on
agricultural systems (Welbaum et al. 2004). These production decisions directly affect
soil quality by influencing the chemical and physical make-up of the soil, the activity and
diversity of the soil microflora and the soil macro and micro fauna and their activity
(Watt et al. 2006, Bossio et al. 1998, Yeates et al. 1997, Lupwayi et al. 1998). Greater
understandings of the negative impacts of traditional agricultural practices are needed if
there is to be a shift towards developing more sustainable agricultural systems (Pankhurst
et al. 1996). In order for this to occur, it is important to understand the affects of the
different management practices used and to determine which of these practices supports a
sustainable agro ecosystem.
Watt et al. (2006) emphasized that there is a large potential for increasing
productivity by managing properly the interactions between management practices and
the soil microflora Moreover understanding the interactions between microbial activity
and management practices may be key to developing sustainable agricultural systems that
maximize crop productivity while reducing inputs.
Saprophytic and symbiotic soil bacteria and fungi have large impacts on the
functioning of agroecosystems through their contributions to decomposition by regulating
16
nutrient cycles (Miller and Lodge 2007). The inputs of carbon and nutrients from plants
influences the abundance of fungi and bacteria in the rhizosphere by driving nutrient
cycles (Acosta-Martinez et al. 2004, Zak et al. 1994, Kennedy 1999). In highly managed
agroecosystems, however, the most common response to management decisions is a
reduced responses in microbial activity resulting from high input of nutrients and
disturbance from plowing (Dick 1994).
A key indicator to how the agricultural soil management practices are affecting
microbial dynamics is through the evaluation of microbial biomass. The microbial
biomass is an important regulator of nutrient transformation and storage of many vital
nutrients including N, P, and S and is also a key component of organic C mineralization
(e,g., Nunan et. al. 1998, Horwarth and Paul, 1994). Previous research has demonstrated
that microbial biomass is a sensitive indicator of differences in sustainable cropping
systems, including the effects of tillage, crop rotations, and soil type on nutrient turnover
and organic C (e,g., Anderson and Domsch 1989, Horwath and Paul 1994). Moreover
the amount and dynamics of soil microbial biomass also considered to be the most
reliable indicator of soil quality (Gil-Sotres et al. 2004).
For a semi- arid agroecosystem, the dynamics of the soil microbial biomass is
regulated in part by the seasonal distribution of precipitation, which can impose a major
stress on the soil microbial community in addition that as a result of management
practices. To maximize sustainability of cropping system in semi-arid environments and
to reduce costs associated with irrigation, tillage practices and fertilizer application this
17
study was conducted within a peanut cropping system to evaluate key production impacts
on soil microbial biomass.
Methods
Plot Design
Field plots for this two year study (2007 and 2008) were established at the USDA-
ARS Cropping System Research Laboratory in Lubbock, Texas USA (101⁰ 47’ west
longitude; 33⁰ 45’ north latitude; 993m elevation. The field soil is an Olton clay loam
(mixed, superactive, thermic and aridic soil) and are classified as Paleustolls (Bronson et
al. 2004). Previous to the initiation of this study, the field was under conventional cotton
production for 6 growing seasons. In November 2006, the whole circular field was
portioned into 36 concentric rows. Approximately 1.3 hectares (3.3 acres), one half of a
circular field was prepared for peanut production to assess the impacts of irrigation,
nitrogen, and tillage on soil microbial dynamics and microbial functionality.
The sample site was divided into six equally sized triangular plots that are labeled
plots 1-6. Every two plots receive the same imposed conditions so that there were two
replicates for each treatment. Within each of the plots, different tillage techniques were
applied alternating between conventional tillage and strip tillage zones. In the
conventional tillage rows there was rye planted in the furrow only and in the strip tillage
portions rye was planted across the bed and the furrow. Within the plots zones of strip
and conventional tillage rows alternated respectively starting from the outside of the
plots. There were two strip and two conventional tillage sections within each zone. Two
18
alternating conventional and strip tillage sections of six rows each were sampled in each
plot.
Plots 1 and 2 served as the control and did not receive nitrogen fertilizer in year
2007. The remaining Plots 3, 4, 5, and 6 received 30lb/acre (33.6 kg/Hectare) of URAN
(Urea Ammonium Nitrate) on July 25th, August 9th, August 27th, September 24th and
September 24th in 2007. In 2008, Plots 1 through 6 received URAN 35 lb/acre (39.2
Kg/Ha) on July 8th, August 1st and September 2nd. Thus, all Plots received nitrogen in
2008.
There were 2 levels of irrigation applied in 2007. Plots 1, 2, 5 and 6 received full
irrigation (100-100-100% EvapoTranspiration (ET)), where 100% ET irrigation refers to
the amount of water supplemented to the crop that equals the total loss of water through
evaporation from the soil and the total transpiration form the aerial parts of the plant. For
2007 full irrigation was determined to be 1.5 inches of water. The late growing seasons
last from beginning of August till the end of October. Plots 3 and 4 were kept under late
deficit treatment (100-100-50%ET), applying only 0.75 inch of irrigation (half) during
the last 90 days while maintain full irrigation prior to. There were 3 levels of irrigation in
year 2008. Plots 1 and 2 received full irrigation (100-100-100%ET). Plots 3 and 4 were
kept under late deficit treatment (100-100-50%ET). Plots 5 and 6 were under early deficit
irrigation. They were irrigated with half of the regular irrigation 90.75 inch quantity in
initial 45 days (beginning of May to mid-June).
19
Crop Management Treatments
In 2007, the treatments applied to the field were: 1) Full Irrigation and no
Nitrogen (plots 1 & 2), 2) Nitrogen and a Late Season Drought (Plots 3 &4), and 3) Full
Irrigation and added Nitrogen (Plots 5 & 6). In 2008, the applied treatments were: 1) Full
Irrigation and added Nitrogen (Plots 1 & 2), 2) Late Season Drought and added Nitrogen
(Plots 3 & 4), and 3) an Early Season Drought and added Nitrogen (Plots 5 & 6). As with
2007 there were alternating conventional and strip tillage rows that were sampled (Figure
2.1).
Sampling
Four soil samples were taken within each plot; two from conventional tillage rows
and two strip tillage rows. For each sample, soil was collected from three different
locations across the plot to account for any variations that may be present within each
section of the zone and combined per row. Soil samples were collected from a depth of
15cm, with approximately 330g of samples taken for each replicate sample. Once
samples were taken, they were kept in a cooler and stored at 4⁰ C until they were
processed for enzymatic analysis.
Microbial Biomass Carbon
The chloroform fumigation and extraction method (Nunan et al. 1998, Vance et
al. 1987) was used to assess microbial biomass carbon (MBC) in response to
management practices. For each sample collection a 10 g dry weight equivalent was
20
measured out for each soil sample. A 10 g dry-weight equivalent is determined by drying
5.0 g soil for 48 hrs at 60°C and measuring the % soil moisture. Two 10 g dry-weight
equivalent sub-samples of each field sample were placed in a dessicator with a moist
paper towel at the bottom to impede the desiccation of soil samples during the
fumigation. Twenty five ml of ethanol-free chloroform with anti-bumping granules will
be added along with the moist paper towel in a dessicator for the fumigation. A second
sub-set of samples were placed within a dessicator without chloroform to serve as a
control. All samples were fumigated for 48 hrs. Soil samples were then extracted using
50ml of a 0.5 M K2SO4 solution, shaken for 1 hours and then filter through Whatman #32
150mm filter paper. To determine the microbial biomass present in each sample, a
subsample of the filtered K2SO4 was read at 280nm according to the procedure described
by Nunan et. al. (1998). To obtain the amount of microbial biomass carbon, the control
absorbance values are subtracted from the fumigated absorbance values and multiplied by
the KEC.
Microbial Community Structure
To evaluate the impacts of management decisions on the structure of the soil microbial
community fatty Acid Methyl Ester (FAME) analyses were conducted (referenced
needed) To conduct the FAME analyses a 3 gram dry weight equivalents were collected
from each treatment plot. The soil samples were first subjected to a saponification step
(45g sodium hydroxide, 150ml methanol, and 150ml distilled water) and boiled in a
100ºC water bath for 30 minutes. Samples were then methylated using a 325ml 6.0N
21
hydrochloric acid, and 275ml methyl alcohol reagent. Subsequently, 200ml hexane,
200ml methyl tert-butyl ether reagent were used to extract the methyl esters into the
organic phase. Lastly, samples were run through a gas chromatography (6890 GC Series
II Hewlett Packard, Wilmington, DE, USA) with a temperature program that increased
the temperature 5ºC per minute starting at 170ºC and ending at 270ºC. A GC detector
measured the peaks from the volatile methyl esters and the Sherlock MIS Software
(MIDI, Inc. Newark, DE) was used to obtain the types and proportional abundances of
each peak.
Data Analysis
Changes in microbial biomass levels in response to different management
practices across seasons and across years were evaluated using one-way ANOVA. The
overall impacts of management affects on microbial were evaluated using mANOVA.
Influences of soil nutrients levels on microbial biomass and the influence of the enzyme
levels on each other were evaluated using bivariate correlations. Statistical analyses
including one-way ANOVA, mANOVA, bivariate correlations were performed using
SPSS 14.0 for windows. Each data set underwent the Least Significant Difference (LSD)
post hoc test when significance (p ≤ 0.05) was indicated on the one-way ANOVA and
mANOVA.
Discriminant function analysis (DFA) was employed to visualize microbial
responses to managanement practices in multivariate space, using MATLAB (Natick,
Massachusetts). DFA analyzed microbial responses to 2007 and 2008 separately. For
22
each DFA, 1000 randomized (bootstrap) iterations were performed in order to build null
distributions from which to compare actual sampling distribution.
Results
Effects of Tillage on MBC
There was no significant difference (p= 0.732) between the overall MBC levels
between the conventional and strip tillage treatments across seasons and the two years of
the study (Figure 2.2a). There was also no significant (p ≤ 0.05) difference seen in MBC
levels between the two treatments within any of the seasons. The overall MBC was
greater in 2008 than in 2007 for both tillage types (Figure 2.2b).
Effects of Irrigation on MBC
Full irrigation vs. late season drought 2007-2008
There was a significant difference in irrigation’s affect on MBC. Overall MBC
was highest across both years and across seasons under full irrigation (p = 0.01) than that
of reducing irrigation resulting in a late season drought (LSD) across both 2007 and
2008(Figure 2.3a). MBC was greater in the full irrigation treatments in every season
compared to LSD. MBC was statistically (p ≤ 0.05) greater fall 2007 and summer 2008
and partially separated in fall 2008 (Figure 2.3b).
23
Full irrigation vs. late season drought vs. early season drought
During 2008, full irrigation produced greater MBC than LSD and early season
drought (ESD) but difference were only significantly different (p ≤ 0.05) from MBC
associated with the “late season drought.” While MBC from the ESD plots appeared
greater than MBC from LSD treatment plots, the values were not significance (p = 0.858)
(Figure 2.4a). Full irrigation produced a consistent pattern of greater MBC than LSD and
ESD within seasons but was significantly greater than LSD in summer 2008 only. The
greatest difference in MBC between LSD and ESD was seen in summer 2008, although
the difference was not considered significant (Figure 2.4b).
Effects of Nitrogen on MBC
Nitrogen fertilization of a peanut cropping system did not have an overall effect
on MBC across years and seasons (Figure 2.5a). There were statistical differences
between fertilized and no fertilizer plots within seasons. There was a trend of the MBC
being higher in both treatments for the summer season compared to the fall, thought this
pattern was only significant (p ≤ 0.05) for the no-nitrogen treatments (Figure 2.5b).
Changes in field conditions
There was a clear separation in microbial structure among all sample dates and
October 2008 among (Figure 2.6a). Note that 1 is also different from 6, 5, 3 and 2. The
mANOVA confirmed that there was a significant (p ≤ 0.05) difference in what between
this month from the rest of what –expand your comparisons. The DFA also showed that
24
microbial community structure in July 2007 separated out from all of the other months
other than October 2007 and that October 2007 separated out from all other months
except July 2007 (Figure 2.6a). Microbial community structure for July and October
2007 was shown to be different from all other months by mANOVA (p ≤ 0.05). Vector
analysis demonstrated that nutrient dynamics and soil organic matter were the driving
force for this temporal separation (Figure 2.6b).
Influences between MBC, nutrients, and enzymatic activity
All significant correlations between MBC and soil nutrients across both growing
seasons were negative (Table 2.1). Some of these negative correlations with MBC
included phosphorus (-28%), potassium (-36%), magnesium (-35%), Boron (-31%) and
manganese (-37%). There were no overall significant relationships between MBC and
levels of extractable NO3 and NH4. No strong correlations were detected between MBC
and any of the measured microbial enzymes with the highest being 17% with α-
galactosidase (Table 2.2).
Temporal changes in MBC
Seasonally, MBC did not change significantly as there was only a significant
difference in MBC between fall 2007 and fall 2008 with MBC in 2008 being significantly
greater at that time period (p ≤ 0.05). There were no differences in MBC levels within
either year (Figure 2.7a). levels of MBC were the highest in October of 2008 and it’s the
lowest in October 2007 (Figure 2.7b & 2.7c).
25
FAME
The relative abundances of fungal FAMES were greater than the relative bacteria
abundances % within every season (Figure 2.8a). The fungi:bacteria ratio of relative
abundance was the highest in the fall season of 2007 and the lowest in the summer of
2008 (Figure 2.8b). In both sampling periods the changes in the fungi:bacterial ratios
were the result of fluctuations in the relative fungal abundances and not those of the soil
bacteriaFungal abundance was significantly greater in fall 2007 and decreased
significantly in summer 2008. Both years showed a significant increase in the
fungi:bacteria raito from the summer to fall (Figure 2.8a & 2.8b).
The relative abundance of Arbuscular Mycorrhizal Fungal FAME profiles and
saprophytic fungi FAME profiles remained consistent and almost identical in all seasons
except during summer 2008. The relative abundance of saprophytic fungi FAMEs
decreased significantly during this season (Figure 2.9a). There was a significant and
positive correlation between MBC levels and relative abundance of saprophytic fungal
FAME levels (R=.361).
The relative abundances of Gram-Positive bacteria FAME profiles remained
consistent throughout all the growing seasons and were significantly higher than the
relative abundances for Gram-Negative bacteria and actinomycetes (Figure 2.9b).
Nitrogen Effects
Nitrogen addition had a significant negative effect (p ≤ 0.05) on the relative
bacteria abundances across 2007 when compared to those plots that received no
26
additional nitrogen (Figure 2.10a). There were seasonal effects with no significant
difference between the relative bacterial abundances in the summer. However, the level
of bacterial FAMES within nitrogen application was significantly less in the fall season
that year.
Nitrogen did not have a significant effect on the relative fungi abundances for the
2007 data combined (Figure 2.11a). However, when looking within the seasons fungal
abundances increased positively to nitrogen addition in the fall season 2007. There were
no differences in the relative fungi abundance % between the nitrogen and no nitrogen
plots in summer 2007 (Figure 2.11b).
Effects of Irrigation
Overall Gram -Positive FAME levels were lower under late season drought
compared to full irrigation (p ≤ 0.05). The overall Gram -Positive relative abundance
levels decreased across both years under late Season Drought when compared to Full
Irrigation (Figure 2.12a). The relative abundances Gram-Positive FAME levels were
greater under full irrigation in every season but the differences were only significant for
fall 2007 (2.12b). Gram-Negative relative abundances exhibited a positive response to the
LSD treatment when compared to Full irrigation (Figure 2.13a). However, this response
to LSD by the Gram-Negative bacteria was not seen in every season. The relative
abundance was greater under LSD in summer 2007 and fall 2008 but only significantly
greater in fall 2008. In summer 2008, the relative Gram-Negative abundance was greater
under full irrigation and no response was able to be measured for fall 2007 as a result of
insufficient amounts of relative Gram-Negative abundance in that season (Figure 2.13b)
27
Discussion
Nitrogen
One of the goals of this study was to understand the impact that nitrogen
application had on the soil microbiological dynamics in a semi-arid agroecosystem.
Previous research examining nitrogen inputs effect on MBC have produced contradictory
results of both positive and negative responses of soil MBC to nitrogen (Moore et al.
2000). The results from this study indicated that nitrogen did not have any significant
effect on the soil MBC.
Nitrogen did elicit a significant response from the soil bacteria associated with the
peanut cropping system. The relative bacteria abundances of FAME profiles responded
negatively to short-term nitrogen application. It is hard to speculate to the true
relationship between added N and relative bacteria abundance due to short-term data
collected. This response could be the result of N inhibition on soil microbial
communities through repression of soil enzyme activity (Bardgett et al. 1999).
Alternatively, the increased N could be reducing the need for N-fixing bacteria in the soil.
Previous research yielding similar results concluded increased N inputs could promote
the growth of certain nematodes which in turn decrease a certain population of the
sustained bacteria (Sarathchandra et al. 2001).
The relative fungal abundance showed an opposite but lesser response to soil
nitrogen addition. The N application effect was not considered significant for the
duration of the study but was considered significant in the fall season when the relative
fungi abundance was at its greatest. The relatively small amount of data collected in the
28
short-term portion of this study makes it is difficult to speculate whether N did have an
effect on fungal abundance or not. Because N fertilization has been shown to cause
chances in the abundance of fungal species (Sarathchandra et al. 2001) and the result was
near significant we feel comfortable to at least suggest that there is a small effect.
The effects seen as result of N application reflect short term results. The data
obtained is only valuable when evaluating short-term management strategies. The effects
of long-term N application can have dramatically different effects than that of short-term
application (Marchner et al. 2003) and such long-term evaluation of N application and its
effect on relative microbial abundance are needed to fully understand the consequences
of this management practice.
Tillage
The two tillage methods compared in this study did not differ in result from one
another. The level of MBC remained almost exactly the same, as did all the relative
microbial abundance within the conventional and strip tillage samples. MBC has often
been shown as a sensitive indicator to changes in tillage practices (Anderson and
Domsch, 1989, Horwath and Paul 1994, Jackson and Calderon 2003) but it has also been
shown to be non-responsive to tillage management changes (Calderon and Jackson 2002,
Aslam et al. 1999). The lack of difference between the two tillage treatments is probably
a result of the similarity between them as strip tillage is a reduction of conventional
tillage. These results indicate that when strategizing between tillage treatments, effects
other than microbial responses such as soil aggregation or erosion should be considered
29
when trying to determine which method will be more beneficial for that particular
agroecosystem.
Irrigation
Full irrigation had a significantly positive effect on the soil MBC and Gram-
Positive bacteria abundance. These results are not surprising as water is necessary for
any successful agroecosystem. These results further demonstrate the necessity of water
for crop productivity. Further research will be needed to determine ways to minimize
irrigation without reducing microbial dynamics. One short-term method evaluated in this
study was timing of irrigation. The effects of late season irrigation and early season
irrigation were inclusive in this study but the results suggest that further investigation
may lead to promising results. Although there were no significant differences between
the LSD and ESD irrigation treatments, variables measured all followed a pattern of
higher levels associated with ESD indicating that irrigation is more influential in the later
part of the growing season. Although this should be evaluated over more than one
growing season, it would appear if water is limited irrigating in the later months will help
to maximize the soil microflora potential.
Soil characteristics and seasonal variation
FAME analysis indicated that the soil in this study was fungal dominated, with
the fungi:bacteria ratios being significantly greater in the fall seasons compared to the
summers. The relative fungi abundance were almost exactly the same between
30
saprophytic fungi and mycorrhizal fungi in all seasons with the exception of the summer
2008 season in which saprophytic fungi displayed a significant decrease in abundance.
Gram-Positive bacteria made up the highest concentration of bacteria in the soil, a result
consistent with other research (Ibekwe and Kennedy 1999). Actinomycetes and Gram-
Negative bacteria abundance were both relatively low and did not display much variation.
This information can be used in future research to help focus on the microbial groups that
could have the most influence on agroecosystems due to their abundance.
DFAs showed strong monthly variation in the plots mainly as the result of
nutrient fluxes. The October 2008 month separate out most significantly followed by the
October 2007 month. Interestingly enough, these two months also represented the
extreme high and low in MBC months with October 2008 displaying the highest MBC
and October 2007 resulting in the lowest MBC. Although, there were no significant
correlations between individual nutrients and soil MBC levels, it would certainly appear
that there may be some interaction between the combined levels of soil nutrients and the
MBC. There results also indicated that there may be some key shifts going on between
the September and October months as each year the MBC between these two months
were the greatest. More detailed examination of trends in these months may provide
insight into soil nutrient shifts and their effects.
31
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34
Table 2.1. Correlation matrix comparing soil nutrients and edaphic characteristics to
Microbial Biomass Carbon from soil samples collected across the growing seasons of
2007-2008. All numbers represent r calculated using Pearson Correlation. The r values
range from 1 to -1 with being a 100% positive correlation and -1 a 100% negative
correlation. Significant r values are indicated in bold. Note [MBC= microbial biomass
C, NO3 = soil extractable nitrate, NH4= soil extractable ammonium, P= soil extractable
phosphorus, K= soil extractable potassium, Mg= soil extractable magnesium, Ca= soil
extractable calcium, S= soil extractable sulfur, B= soil extractable boron, Zn= soil
extractable zinc, Mn= soil extractable manganese, Fe= soil extractable iron, Cu= soil
extractable copper, CEC= cation exchange].
NO3 NH4 pHw P K Mg Ca S B Zn Mn Fe Cu CEC

MBC 0.12 0.04 0.03 0.28 0.36 0.35 0.09 0.03 0.31 0.09 0.37 0.27 0.25 0.03
Table 2.2. Correlation matrix comparing soil extractable nutrients to Microbial Biomass
Carbon from soil samples collect across the growing seasons of 2007-2008. All numbers
represent r calculated using pearson correlation. The r values range from 1 to -1 with
being a 100% positive correlation and -1 a 100% negative correlation. Significant r
values are indicated in bold. Note [MBC= microbial biomass, Phosph=
phosphodiesterase, Glucomini= β-glucosaminidase, Glucosidase= β-glucosidase,
Alkaline= alkaline phosphatase, Alpha= α-galactasidase].
Phospho Gucomini Glucosi Alk Alpha

MBC -0.03 0.07 0.04 0.13 0.17
35
Figure 2.1. Plot design established to study the effects of irrigation, nitrogen fertilization,
and tillage on soil microbial dynamics and community structure associated with a peanut
cropping system in semi-arid west-Texas in year 2007. The numbers indicated refer to the
different Plots. Tillage treatment and Plot alignment remained the same in 2008.
Irrigation and nitrogen treatments differed in Plots 1,2,5, and 6 in 2008.
36
Figure 2.2. The effect of strip tillage compared to conventional tillage treatments on
Microbial Biomass Carbon (MBC) associated with a peanut cropping system in semi-arid
West Texas. (a)Total MBC from 2007-2008. Values are means± S.E., n= 96. (b) MBC
in each of the summer and fall seasons in both 2007 and 2008. Values are means± S.E.,
n=24. Groups statistically (p ≤ 0.05) different are indicated with different letters above
mean bars.
Conventional
Microbial Biomass C (g/ g soil) 600 Strip
a a
500
400
300
200
100
a.
700
Conventional
Strip
a
Microbial Biomass C (g/ g soil)
600
ab ab
ab ab
b b b
500
400
300
200
100
Summer 07 Fall 07 Summer 08 Fall 08

b.
37
Figure 2.3. The effect of late season drought (LSD) compared to full irrigation (Full)
on Microbial Biomass Carbon (MBC). (a)Total MBC from 2007-2008. Values are
means± S.E., n= 64. (b) MBC in each of the summer and fall seasons in both 2007
and 2008. Values are means± S.E., n=16. Groups statistically different are indicated
with different letters above mean bars.
600 Full
Microbial Biomass C (g/ g soil) LSD a
500
b
400
300
200
100
a.
700
Full
a a
LSD
600
ab ab
bc bc bc
500
c
400
300
200
100

b.
38
Figure 2.4. The effect of late season drought (LSD) and early season drought (ESD)
compared to full irrigation (Full) on Microbial Biomass Carbon (MBC). (a)Total MBC
from 2007. Values are means± S.E., n= 32. (b) MBC in each of the summer and fall
seasons in both 2007 and 2008. Values are means± S.E., n=16. Groups statistically
different are indicated with different letters above mean bars.
700
Full
Microbial Biomass C (g/ g soil) LSD a
600 ESD
ab
b
500
400
300
200
100
a.
Full
700
LSD
ESD a a
600
ab ab
ab
500 b
400
300
200
100
Summer 08 Fall 08
b.
39
Figure 2.5. The effect of nitrogen application compared to nitrogen application

treatments on Microbial Biomass Carbon (MBC). (a)Total MBC from 2007-2008.
Values are means± S.E., n= 32. (b) MBC in each of the summer and fall seasons in both
2007 and 2008. Values are means± S.E., n=16. Groups statistically (p ≤ 0.05) different
are indicated with different letters above mean bars.
No nitrogen
Microbial Biomass C (g/ g soil) 600 nitrogen
a a
500
400
300
200
100
a.
700 No nitrogen
nitrogen
a
600
ab
500 bc
c
400
300
200
100
Summer 07 Fall 07
b.
40
Figure 2.6. (a) Discriminate function analysis (DFA) examining microbial and nutrient
dynamics associated with different management programs associated with a peanut
production system in semi-arid West Texas conditions in the various months in 2007 and
2008 using different enzyme activity levels, nutrient levels, and MBC as the parameters.
Note [1= June 07, 2= July 07, 3= Sept. 07, 4= Oct. 07, 5= June 08, 6=July 08, 7=Sept. 08,
8= Oct. 08, Phosph= phosphodiesterase, Glucomini= β-glucosaminidase, Glucosidase= β-
glucosidase, Alkaline= alkaline phosphatase, Algal= α-galactasidase, NO3 = soil
extractable nitrate, NH4= soil extractable ammonium, P= soil extractable phosphorus, K=
soil extractable potassium, Mg= soil extractable magnesium, Ca= soil extractable
calcium, S= soil extractable sulfur, B= soil extractable boron, Zn= soil extractable zinc,
Mn= soil extractable manganese, Fe= soil extractable iron, Cu= soil extractable copper,
CEC= cation exchange, MBC= microbial biomass C].
a.
41
Figure 2.6. (b) Vector correlations indicating correlations between the different nitrogen
treatments in summer and fall and enzyme activities. Correlations are indicated by the
smallest angle between two vectors. Ex 0°= 100% positive correlation, 90°= no
correlation, and 180°= 100% negative correlation. Note [1= June 07, 2= July 07, 3=
Sept. 07, 4= Oct. 07, 5= June 08, 6=July 08, 7=Sept. 08, 8= Oct. 08, Phosph=
Alkaline= alkaline phosphatase, Algal= α-galactasidase, NO3 = soil extractable nitrate,
NH4= soil extractable ammonium, P= soil extractable phosphorus, K= soil extractable
potassium, Mg= soil extractable magnesium, Ca= soil extractable calcium, S= soil
extractable sulfur, B= soil extractable boron, Zn= soil extractable zinc, Mn= soil
extractable manganese, Fe= soil extractable iron, Cu= soil extractable copper, CEC=
cation exchange, MBC= microbial biomass C].
b.
42
Figure 2.7. Changes in Microbial Biomass Carbon (MBC) during the growing
seasons from 2007-2008. (a)MBC during the summer and fall seasons of 2007-2208.
Values are means± S.E., n= 48. (b) MBC in months corresponding to enzyme
measurements. Values are means± S.E., n=24. (c) MBC in all measured months.
Values are means± S.E., n=24. Groups statistically (p ≤ 0.05) different are indicated
with different letters above mean bars.
600
a

ab
ab
500 b
400
300
200
100

a.
700
a
600
ab ab ab b
bc
500 bc
c
400
300
200
100
June 07 July 07 Sept 07 Oct 07 June 08 July 08 Sept 08 Oct 08

b.
700
ab a
600 ab
ab ab ab ab bc
bc bc
500
c c
400
300
200
100
07
08
07
07
07
07
07
08
08
08
08
08
ly
ly
ay
ay
ne
pt
ct
ne
pt
ct
Au
Au
Ju
Ju
O
O
Se
Se
M
M
Ju
Ju
c.
43
Figure 2.8. Relative bacteria and fungi abundances measured by fatty acid methyl ester
(FAME) analysis across two growing seasons in a peanut production system in West
Texas. (A) Relative FAME abundance (%) of bacteria and fungi. (B) Change in the
Fungal:Bacterial ratio during the growing seasons 2007-2008. Means were calculated by
relative fungi abundance (%) over relative bacteria abundance (%). Groups statistically
(p ≤ 0.05) different are indicated with different letters above mean bars.
Bacteria
30 Fungi
Relative Microbial Abundance (%)
25
20
15
10

a.
a
2.5
Relative Microbial Abundance (%)
Fungal:Bacterial Ratio
2.0
b b
1.5
c
1.0
0.5

b.
44
Figure 2.9. Relative bacteria and fungi abundances measured by fatty acid methyl ester
(FAME) analysis across two growing seasons in a peanut production system in West
Texas.. (A) Relative abundance (%) of FAMEs associated with AMF and Saprophytic
fungi. (B) Changes in the relative abundance (%) of FAMEs associated with Gram
Positive, Gram Neagitive, and Actinomycete bacteria.
16 AMF
Saprophytes
14
Relative Fungi Abundance (%)
12
10

a.
14 Gram +
Gram -
Relative Bacteria Abundance (%)
Actinomycetes
12
10

b.
45
Figure 2.10. Effects of nitrogen on relative bacteria abundances associated with a

peanut production system in West Texas. (a) Relative bacteria abundance % in year
2007. Values are means± S.E., n=48. (b) Relative bacteria abundance % in growing
seasons in year 2007. Values are means± S.E, n=24. Groups statistically (p ≤ 0.05)
16 No nitrogen
nitrogen
a
Relative Bacteria Abundance %
14
b
12
10
a.
16 No nitrogen
nitrogen
a a
Relative Bacteria Abundance (%)
14
ab
12 b
10
Summer 07 Fall 07
b.
46
Figure 2.11. Effects of nitrogen on relative fungi abundance associated with a peanut
production system in West Texas. (a) Relative fungi abundance % in year 2007. Values
are means± S.E., n=48. (b) Relative fungi abundance % in growing seasons in year 2007.
Values are means± S.E, n=24. Groups statistically (p ≤ 0.05) different are indicated with
different letters above mean bars.
30
No nitrogen
nitrogen a
a
25
20
15
10
a.
No nitrogen a
nitrogen
30
c c
20
10
Summer 07 Fall 07
b.
47
Figure 2.12. Effects of irrigation on relative Gram-Positive bacteria abundance

associated with a peanut production system in West Texas. (a) Relative gram-positive
bacteria abundance % in year 2007 & 2008. Values are means± S.E. (b) Relative Gram-
Positive abundance % in growing seasons in years 2007 & 2008. Values are means± S.E.
Groups statistically (p ≤ 0.05) different are indicated with different letters above mean
bars.
Relative Gram-Positive Bacteria Abundance (%) Full Irrigation

12
LSD
a
10 b
a.
Relative Gram-Positive Bacteria Abundance (%)
14 Full Irrigation
LSD
12 a
ab
abc ab
abc c abc
10 bc

b.
48
Figure 2.13. Effects of irrigation on relative Gram-Negative bacteria abundance %. (a)

Differences In relative gram-negative bacteria abundance % in year 2007 & 2008.
Values are means± S.E. (b) Differences in relative Gram-Negative abundance % in
growing seasons in years 2007 & 2008. Values are means± S.E. Groups statistically (p ≤
0.05) different are indicated with different letters above mean bars. * indicates values too
low to be displayed on graph.
Relative Gram-Negative Bacteria Abundace (%) 1.2

Full Irrigation
LSD a
1.0
b
0.8
0.6
0.4
0.2
a.
Relative Gram-Negative Bacteria Abundance (%)
2.5 Full Irrigation

LSD
a
2.0
1.5 b
b
bc
1.0
c
c
0.5

b.
49
CHAPTER 3
EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID AGROECOSYSTEM

ON SOIL MICROBIAL ENZYMATIC ACTIVITY
Introduction
Enzymes are biological catalysts that facilitate reactions without undergoing
permanent chemical changes and are necessary for any biochemical reaction to take place
under most natural conditions. In all terrestrial ecosystems, microbial enzymes are
essential to perform necessary ecosystem processes by breaking down organic and
inorganic compounds into useable forms for subsequent biomass production by plants
and soil microbes. While the production of enzymes have multiple levels of regulation,
from their synthesis to their reactivity, for microbial enzymes excreted into the soil,
activity is influenced by soil pH, ionic strength, soil temperature, and the presence and
absence of clay and organic matter that can bind soil enzymes (Dick and Tabatabai
1993).
For the soil ecosystem, enzymes can be found both in the form of intracellular
enzymes and extracellular enzymes. While soil enzymes originate mainly from plants
and microorganisms, the largest portion originates from the microbial biomass (Tabatabai
1994). As bacteria and fungi are the main source of enzymes in the soils the composition
and abundance of microbes that produce these degradative enzymes controls soil
processes (Acosta-Martinez et al. 2004). Decomposition rates, nutrient cycling,
decomposition of xenobiotics, and soil aggregation are critical soil processes that are
controlled by the microbial enzyme activities (Barea et al. 1996, Reynolds et al. 2003).
50
Once extracellular enzymes are produced the activity of these molecules can become
uncoupled from biological activity. While extracellular enzymes can be degraded over
time, a significant portion becomes stabilized by binding to clay particles and forming
complexes (Boyd and Mortland 1990). The enzymes in these enzyme-clay complexes
still have activity but their activity is significantly reduced, thus directly impacting rates
of critical ecosystem processes. Formation of these complexes with clay can also causes
the enzymes to be immobilized, thus completely decreasing activity (Dick and Tabatabai
1987). For most terrestrial ecosystems it is known that the activity and production of
extracellular enzymes are influenced by vegetation composition, agricultural practices,
such as tillage and chemical applications, and levels of organic matter (Tabatabai 1994).
It is not unreasonable to expect the management practices used in agriculture soils can
have affects on the activity of the enzymes in the soils (Bandick and Dick 1999).
Changes in concentration and activity of certain extracellular enzymes in the soil
can have strong impacts on the nutrient cycles of those soils. Substantial changes in the
dynamics of extracellular enzymes will often be seen in the rhizosphere region where
previous research has shown greater enzymatic activity than that of non-rhizosphere soils
(e.g. Dick 1994). This can be explained by the greater amount of plant material and
microorganisms and species diversity associated with the rhizosphere compared to the
bulk soil. The roots themselves excrete carbon to stimulate microbial activity that can
increase the production of microbial enzymes (Dick 1994). Soil enzymes can lead to
higher levels of crop yields when the byproducts of their reactions including increased
mineralization are utilized by the roots in the rhizosphere (Dick and Tabatabai 1993).
51
The activity of many soil enzymes has been found to correlate with the fertility status of
soils (Dick and Tabatabai 1993) and enzyme assays give an estimate of the microbial
activity of the soil in relationship to the management practices that are employed.
For agricultural systems, enumerating the seasonal levels of hydrolases,
phosphatases, phosphodiesterase and β-glucosidase (e.g., Acosta Martinez et al. 2009)
provide key insight to the soil quality of the system. Hydrolases are a class of enzymes
that catalyze the hydrolysis of proteins, nucleic acids, starches, fats, phosphoric acids,
and other macromolecule substances. Included in the hydrolases are the phosphatase
enzymes which catalyze the hydrolysis of esters and anhydrides of phosphoric acids.
Phosphatses play important roles in P and N cycles in soils making them valuable
enzymes to examine when trying to understand microbial dynamics (Dick and Tabatabai
1993). Acid phosphatase or alkaline phosphatase, phosphodiesterase, β-glucosidase, α-
galactosidase, and β-glucosaminidase all contribute to P and N dynamics in soils.
Acid phosphatase and alkaline phosphatase are two important
phosphomonoesterases that have been shown to be pivotal in soil organic P
mineralization and have been seen to be specifically involved in hydrolysis in soils of β-
glycerophosphate, phenylphosphate, β-napthyl phosphate, and p-nitrophenyl (Tabatabai
1994). The two enzymes differ in pH ranges of their optimum activity. Acid
phosphatases have optimum activity in the acidic pH ranges and are usually most active
in soils with a pH of around 6. Alkaline phosphatases have optimum activity in the
alkaline pH ranges and have been shown to be most active with a pH of around 8 (Dick
and Tabatabai 1993). The activity of acid and alkaline phosphatase has been shown to
52
become more active in certain cropping systems as organic P becomes more limited
(Baligar et al. 1988).
Phosphodiesterase has been the least studied phosphatase in soils and is believed
to potentially be an important component of the P cycle and contributes to the
degradation of nucleic acids in the soil (Tabatabai 1994). Moreover, Phosphodiesterase
has been suggested to be an extracellular enzyme utilized by Arbuscular Mycorrhizal
fungi in their symbiosis with plants (Lindahl et al. 2005).
β-glucosidase is an important extracellular soil enzyme that provides a major
energy source for soil microbes and has been positively correlated to crop yields (Dick
1997). β-glucosaminidase is a key extracellular enzyme involved in N mineralization
that hydrolyzes C-N bonds of organic sugars free NH3 that can be uptaken by plants
(Dick 1997, Ekenler and Tabatabai 2002). α-galactosidase is another enzyme involved in
C dynamics by hydrolyzing melibiose and lactose (Dick 1997).
Tillage and crop-residue management can have significant impacts on microbial
mediated biochemical reactions in the soil (Dalal 1989, Spedding et al. 2004). These
impacts are the result of the effects the tillage practices on altering the distribution, and
activities of soil microbial communities and in turn an effect on the soil microbial
enzymes (Dick 1984). Organic C and N have been shown to accumulate at the soil
surface (0-2. cm) under reduced tillage involving crop residue placement (Havlin et al.
1990). Extracellular soil enzymes have been shown to be significantly affected by tillage
(Deng and Tabatabai 1996) and in general have lower activity in no-tillage management
compared to conventional tillage (Doran 1980, Angers et al. 1993).
53
The objective of this component of the overall project was to evaluate the response of the
microbial activity (enzyme production) to tillage, irrigation, and nitrogen of a peanut
cropping system in semi-arid soils.
Methods
Plot Design
Field plots for this two year study (2007 and 2008) were established at the USDA-
ARS Cropping System Research Laboratory in Lubbock, Texas USA (101⁰ 47’ west
longitude; 33⁰ 45’ north latitude; 993m elevation. The field soil was a mixed superactive,
thermic and aridic soil (Olton clay loam) and are classified as Paleustolls (Bronson et al.
2004). Previous to the initiation of this study, the field was used for continuous cotton
production under conventional tillage for a duration of 6 growing seasons. The circular
field was portioned into 36 concentric rows prior to the initiation of this study in
November 2006. Approximately 1.3 hectares (3.3 acres), one half of a circular field was
prepared for peanut production to assess the impacts of irrigation, nitrogen, and tillage on
soil microbial dynamics and microbial functionality.
The sample site was divided into six equally sized triangular plots that are labeled
plots 1-6. Every two plots receive the same imposed conditions so that there were two
replicates for each treatment. Within each of the plots, different tillage techniques were
applied alternating between conventional tillage and strip tillage zones. In the
conventional tillage rows there is rye planted in the furrow only and in the strip tillage
portions rye is planted across the bed and the furrow. The plots alternate between zones
54
of strip and conventional tillage rows respectively starting from the outside of the plots.
There are two strips and two conventional tillage sections within each zone. Two
alternating conventional and strip tillage sections of six rows each were sampled in each
plot.
Zone 1 and 2 served as the control and did not receive nitrogen fertilizer in year
2007. The remaining zones 3, 4, 5, and 6 received 30lb/acre (33.6 kg/Hectare) of URAN
(Urea Ammonium Nitrate) on July 25th, August 9th, August 27th, September 24th and
September 24th in 2007. In 2008, zones 1 through 6 received URAN 35 lb/acre (39.2
Kg/Ha) on July 8th, August 1st and September 2nd. All 6 zones received nitrogen
fertilization in 2008.
In 2007, two levels of irrigation were implemented. Zone 1, 2, 5 and 6 served as
controls and received full irrigation (100-100-100% EvapoTranspiration (ET)). 100% ET
irrigation is defined as the amount of water supplemented to the crop equaling the total
loss of water through evaporation (water lost from the soil to the atmosphere) and the
total transpiration (subsequent water lost from aerial parts of the plant). For 2007 full
irrigation was determined to be 1.5 inches of water. The late growing seasons last from
beginning of August till the end of October. Zones 3 and 4 were kept under late deficit
treatment (100-100-50%ET), applying only 0.75 inch of irrigation (half) during the last
90 days while maintain full irrigation prior to. There were 3 levels of irrigation in year
2008. Zones 1 and 2 received full irrigation (100-100-100%ET). Zones 3 and 4 were
kept under late deficit treatment (100-100-50%ET). Zones 5 and 6 were under early
55
deficit irrigation. They were irrigated with half of the regular irrigation 90.75 inch
quantity in initial 45 days (beginning of May to mid-June).
Crop Management Treatments
In 2007, the treatments applied to the field were: 1) Full Irrigation and no
Nitrogen (plots 1 & 2), 2) Nitrogen and a Late Season Drought (plots 3 &4), and 3) Full
Irrigation and added Nitrogen (plots 5 & 6). In 2008, the applied treatments were: 1) Full
Irrigation and added Nitrogen (plots 1 & 2), 2) Late Season Drought and added Nitrogen
(plots 3 & 4), and 3) an Early Season Drought and added Nitrogen (plots 5 & 6). As with
2007 there were alternating conventional and strip tillage rows that were sampled (Figure
3.1).
Sampling
Four soil samples were taken within each plot; two from conventional tillage rows
and two strip tillage rows. For each sample, soil was collected from three different
locations across the plot to account for any variations that may be present within each
section of the zone and combined per row. Soil samples were collected from depth of
15cm, with approximately 330g of samples taken for each replicate sample. Once
samples were taken, they were kept in a cooler and stored at 4⁰ C until they were
processed for enzymatic analysis.
56
Enzyme Assays
Alkaline phosphatase, phosphodiesterase, β-glucosaminidase, β-glucosidase, and
α-galactosidase were measured for activity in all samples following the procedures
described by Tabatabai (1994) at their optimal pH value. Results were expressed in
milligrams of p –nitrophenol released per kilogram soil per hour (Acosta-Martinez et. al.
2003). Absorbance was measured at 400 nm, the optimal wavelength of p –nitrophenol
(Eril and Avci 2002).
Data Analysis
Differences in individual enzymatic activities in response to different
management practices were evaluated using one-way ANOVA. The overall impacts of
management affects on the total microbial enzymatic activities of the soil and total
nutrient influence of the soil were evaluated using mANOVA. Influences of soil
nutrients levels on enzymatic activities and the influence of the enzyme levels on each
other were evaluated using bivariate correlations. All analyses were performed using
SPSS 14.0 for windows. Each data set underwent the Least Significant Difference (LSD)
post hoc test when significance (p ≤ 0.05) was indicated on the one-way ANOVA and
mANOVA.
Discriminant function analysis (DFA) was employed to visualize microbial
responses to managanement practices in multivariate space, using MATLAB (Natick,
Massachusetts). DFA analyzed microbial responses to 2007 and 2008 separately. For
each DFA, 1000 randomized (bootstrap) iterations were performed in order to build null
distributions from which to compare actual sampling distribution.
57
Results
Effect of Irrigation
Full Irrigation vs. Late Season Drought (LSD) for the 2007-2008 growing seasons
Phosphodiesterase Activity
There was not a significant (p = 0.065) effect of the LSD treatment on
phosophodiesterase activity when compared to the full irrigation control (Figure 3.2a)
over the two year period. However, there did appear to be the trend of higher levels of
phosphodiesterase activity in the full irrigation treatment when compared to the levels
seen in the LSD treatment. This pattern was further revealed when the phosphodiesterase
levels between the two treatments were compared throughout the individual seasons.
Phosphodiesterase activities were consistently greater (p ≤ 0.05) under full irrigation
treatment than when exposed to late season drought in the fall months of 2007 (Figure
3.2b).
β-Glucosaminidase Activity
β-glucosaminidase activity did not differ in response to irrigation (p= 0.137)
(Figure 3.3a). There was a noticeable pattern of higher β-glucosaminidase under full
irrigation than LSD β-glucosaminidase levels. This was seen in the summer months of
2008 when the β-glucosaminidase levels of the Full Irrigation treatment were
significantly (p ≤ 0.05) higher than that of the β-glucosaminidase levels of the LSD
treatment (Figure 3.3b).
58
β-Glucosidase Activity
The effect of the full irrigation treatment was overall significantly (p ≤ 0.05)
greater than that of the LSD treatment on the levels of β-glucosidase activity across both
sampling seasons (Figure 3.4a). The effect of irrigation treatment resulted from the fall
months of 2007, in which the full irrigation treatment produced significantly (p ≤ 0.05)
higher β-glucosidase levels than the LSD treatment for this same period (Figure 3.4b).
Alkaline Phosphatase Activity
There was no significant (p= 0.44) difference seen between the full irrigation
treatment and the LSD treatment on the overall levels of alkaline phosphatase activity
across both sampling seasons (Figure 3.5a). This trend of no difference between the two
treatments was observed in each of the summer and fall seasons in 2007 and 2008
respectively (Figure 3.5b).
α-Galactosidase Activity
The activity levels of α-galactosidase did not exhibit any overall difference (p=
0.754) between the full and LSD irrigation treatments (Figure 3.6a). Furthermore, no
difference could be seen in the summer and fall months in both 2007 and 2008 (Figure
3.6b).
Full Irrigation vs. Late Season Drought (LSD) vs. Early Season Drought (ESD) during
growing season of 2008
The mANOVA showed that there was not a significant difference between the
three irrigation treatments when all enzymes are used. Furthermore, DFAs showed that
59
the irrigation treatment groups do not separate out for both years combined (Figure 3.7a).
However, if either phosphodiesterase or α-galactosidase data are removed as parameters
the mANOVA detected significant differences across irrigation treatments (p ≤ 0.05).
Phosphodiesterase and α-galactosidase activity levels were shown to be affected the least
by the three irrigation treatments (Figure 3.7b). One-way ANOVA showed that there
were significant (p ≤ 0.05) differences between the LSD and ESD treatments for β-
glucosaminidase and alkaline phosphatase activity levels with full higher levels in
response to ESD treatments. A significant difference between full vs. LSD for β-
glucosidase activity levels was seen with full irrigation producing higher enzymatic
activity levels (Tale 3.1).
Effects of Nitrogen
No nitrogen vs. Nitrogen Application: 2007
Effect on combined enzyme activity
The mANOVA indicated that the addition of nitrogen did not have a significant
effect on the combined enzyme activity compared to the no nitrogen treatment. There
were patterns seen on the levels of enzyme activities for the individual enzymes and the
effect of the two treatments varied depending on the Individual enzyme examined across
the growing season (Figure 3.8).
There were no significant (p = 0.263) effect on phosphodiesterase levels between
the nitrogen and no nitrogen treatment (Figure 3.9a). There were also no seasonal
60
differences seen between the phosphodiesterase activity levels in the two treatments
(Figure 3.9b).
There were no significant (p= 0.086) effects of nitrogen on the overall levels of β-
glucosaminidase activities. However there did appear to be a trend of lower levels
associated with the nitrogen treatments across the growing seasons (Figure 3.10a).
When looking at the summer and fall seasons, there are no statistical significant
differences between the two treatments (Figure 3.10b).
The nitrogen and no nitrogen treatments showed no significant (p= 0.685)
differences in the overall β-glucosidase activity levels (Figure 3.11a). The nitrogen
treatment did display significantly (p ≤ 0.05) greater β-glucosidase activity levels in the
fall than that of the no nitrogen treatment (Figure 3.11b).
The overall levels of alkaline phosphatase activity appeared to be greater with the
no nitrogen treatment compared to treatment with nitrogen (Figure 3.12a) but there
wasn’t a significant (p= 0.111) difference. The no nitrogen treatment was statistically no
greater than the nitrogen treatment in both the summer and fall season even though it
appeared to have a pattern of greater alkaline phosphates activity (Figure 3.12b).
61
There were no significant differences (p= 0.652) between the nitrogen and no
nitrogen treatment on the overall α-galactosidase activity levels (Figure 3.13a). The
activity of this enzyme did not differ across season irrespective of the addition of nitrogen
(Figure 3.13b).
Combined enzymatic effect across growing seasons in 2007
The mANOVA found that the there were no significant (p≤ 0.05) differences
between the no nitrogen and nitrogen treatments for all enzymes combined for either the
summer or fall seasons. However, enzyme activities under nitrogen addition during
summer sampling approached significant differences in comparison to no-nitrogen
summer (p= 0.092) and no nitrogen fall (p= 0.082). There was a significant seasonal
difference (p ≤ 0.001) difference seen between the summer and fall nitrogen treatments.
This was further seen in DFA which displayed a separation between the nitrogen summer
treatments from the rest of the treatments (Figure 3.14a). This separation was driven
most by the levels of β-glucosidase activity (Figure 3.14b).
Tillage Effects: Conventional vs. Strip (2007-2008)
Levels of phosphodiesterase activity were overall higher under conventional
tillage than activity levels across years under strip-tillage (Figure 3.15a) but the levels
were not significantly different (p= 0.77). Moreover, the enzyme activity levels between
the two treatments were not significantly different during any of the summer or fall
62
seasons in 2007 and 2008. However, the fall season of 2008 had significantly lower
levels of phosphodiesterase activity than all the other seasons for both tillage treatments
(Figure 3.15b).
There was no significant (p= 0.424) difference in β-glucosaminidase between the
conventional and tillage treatments on the overall levels of β-glucosaminidase activity
across all seasons (Figure 3.16a). Within seasons, β-glucosaminidase levels differed only
during the 2008 season when the summer levels were significantly higher for the
conventional summer and the strip fall of 2008 (Figure 3.16b).
There was no significant difference (p= 0.412) between the overall levels of β-
glucosidase activity between the conventional and strip tillage treatments across both
growing seasons (Figure 3.17a). There were no significant (p ≤ 0.05) differences in β-
glucosidase activity levels between the two tillage treatments within any of the seasons.
However, there was an observed seasonal difference as the summer seasons had
significantly lower levels of activity than the fall seasons for both the conventional and
strip tillage treatments (Figure 3.17b).
There was no significant difference (p= 0.736) between the overall levels of
alkaline phosphatase activity between the conventional and strip tillage treatments across
both years (Figure 3.18a). While there were no significant (p ≤ 0.05) difference observed
63
in alkaline phosphatase levels between the two treatments within any of the seasons,
alkaline phosphatase levels for the strip tillage treatments in 2007 were lower than those
observed under conventional tillage (Figure 3.18b). However, for the 2008 growing
season, the levels of alkaline phosphatase activities were higher during the fall under the
strip tillage treatment.
Across both growing seasons, α-galactosidase activity was not significantly
different between the conventional and strip tillage treatments (p= 0.373) (Figure 3.19a).
There were no significant (p ≤ 0.05) differences between the two tillage treatments within
any of the seasons, however the α-galactosidase levels were significantly less in 2007
than 2008 for both treatments (Figure 3. 19b).
Combined enzymatic and nutrient effect
The mANOVA indicated that tillage did not impact the total soil nutrient and
extracellular enzyme dynamics (p= 0.575). However, when examining the annual tillage
effect there was a separation of the different groups (p ≤ 0.001). Discriminate function
analysis (Figure 3.20) displayed the groups separating out by different years. There was
a complete separation among the nutrient and enzymatic effects of the conventional
treatment between 2007 and 2008. The DFA did not show a complete separation
between nutrient and enzyme dynamics between the strip 2007 treatment and strip 2008
treatment but the subsequent ANOVA indicated that overall nutrient and enzymatic
levels were significantly different between the two growing years (p ≤ 0.001).
64
Effect of nutrients
Correlation between nutrient levels and enzyme levels
Ammonium (NH4) and organic matter (OM) levels had the highest correlations
with individual enzyme. Phosphodiesterase activities showed correlated (-0.43) with
NH4 levels. OM was shown to have the strongest and largest number of significant
correlations with the most enzymes with including phosphodiesterase (-0.21), β-
glucosaminidase (0.33), alkaline phosphatase (0.29), along with α-galactosidase (0.43).
The levels of β-glucosaminidase was the least related to any of the soil nutrient
parameters measured (Table 3.2).
Correlations between enzyme activity levels
Alkaline phosphatase and α-galactosidase showed the strongest significant
correlation (36%) of activity levels followed by β-glucosaminidase and α-galactosidase
(33%). Activity of β-glucosaminidase had the most significant correlations with other
(Table 3.3).
Changes in key nutrient levels (growing seasons of 2007-2008)
Soil Extractable Nitrate
NO3 levels were highly variable across all treatments in 2007 compared to the
levels in 2008 (Figure 3.21). There was no statistical difference between NO3 levels in
the summer and fall of 2008. There was much more variability in the NO3 levels in the
different seasons during 2007. The levels in the summer of 2007 were statistically
65
greater than levels seen in any other season in any of the two years. Conversely, the
levels in the fall of 2007 were statistically lower than any other season in either of the
two years (Figure 3.21).
Soil Extractable Ammonium (NH4) Dynamics
NH4 levels across all treatments remained very consistent from the summer of
2007 through the summer of 2008 (Figure 3.22). In the fall 2008 the levels showed a
significant increase. NH4 levels in the fall of 2008 exhibited 15 fold increase when
compared to the NH4 levels from the summer 2008 growing season (Figure 3.22).
Soil Phosphorus (P) dynamics
P levels showed no statistical (p ≤ 0.05) differences between the seasons within an
individual year but exhibited significant differences between the two years irrespective of
season (Figure 3.23).
% Soil Organic Matter (OM) dynamics
OM levels remained almost completely consistent throughout the 2007 and 2008
growing season with one exception. The levels of soil organic matter (OM) were
significantly (p ≤ 0.05) greater in the summer of 2008 than either season of 2007 or the
fall of 2008 (Figure 3.24).
66
Discussion
The worlds’ population is continually proliferating and as a result so must the
worlds’ food supply. Increasing the efficiency of modern agriculture will be monumental
in helping to meet the ever rising food demands of a growing human population. Current
agricultural practices such as chemical inputs and irrigation are becoming more
expensive and are not as sustainable as low-input systems (Clark et al. 1998, Zak and
McMichael 2001). Furthermore, global climate changes may present even more
difficulties in maintaining and improving on current productivity (Adam et al. 1990).
Finding low input solutions to maximizing agricultural production will not only cut down
on cost and potentially increase productivity but will also help create more sustainable
agroecosystems even in a more unpredictable climate (Stainforth et al. 2005).
Understanding how agricultural practices affect the microbial dynamics of the soil will be
essential to developing the most effective and sustainable agroecosystems. Determining
which tillage technique, irrigation strategies, and fertilizer application are the best for a
particular agroecosystem will be the key to developing the most efficient system. The
semi-arid conditions in West Texas impose unique challenges to developing sustainable
production systems under increasing costs of fertilizers and greater demands for limited
water supplies.
Effects of Irrigation
As full irrigation maintained higher plant growth and provided consistently
optimum conditions for microbial activity, one would expect the highest levels of enzyme
67
activities under these conditions thin in either early of late season drought. The latter two
treatments were established to determine if a reduction in irrigation at specific periods in
crop growth would not have detrimental effects. Full irrigation only produced
significantly (p ≤ 0.05) greater activity levels for β-glucosidase suggesting that having
consistent moisture alone will not provide for maximum microbial activity.
β-glucosaminidase and alkaline phosphatase did not show any differences to the
overall response to the different irrigation treatments but the ANOVA confirmed that
they did show significant differences between the late season drought treatment and the
early season drought treatment within particular seasons. This seasonal pattern may
suggest that β-glucosaminidase and alkaline phosphatase production is highly dependent
on when adequate soil moisture available. Both enzymes displayed significantly greater
activity levels in the early season drought treatment than they did in the late season
drought treatment in 2008 suggesting that seasonal differences in either production or the
densities of microbes producing these enzymes may be contributing to the observed
pattern.
Response to nitrogen application
Levels of β-glucosaminidase showed the strongest response to nitrogen
significantly increasing with its input. This result was to be expected due to its role in N
cycling by breaking C-N bonds of amides and releasing NH3 in the process used in N
mineralization which provides a useable form of N for plants (Dick, 1997). Moreover β-
glucosaminidase can contribute 5-10% of the organic N in the soil surface by hydrolyzing
68
amino sugars (Ekenler and Tabatabai, 2002). This result suggest that β-glucosaminidase
is aiding in N cycling.
β-glucosidase is involved in the hydrolysis of glycosidic molecules releasing
glucose to be used for and energy source for the micro flora (Dick 1997) and would not
be expected to be strongly affected by N inputs. It indeed did not display an overall
response to the nitrogen application but did show significantly greater levels of activity
with nitrogen application in the fall season. One explanation for this observation is a lag
response to altered C:N ratios in the soil from plant growth during the summer. Soil
enzymes have been shown to respond to changing C:N ratios in soil (Sinsabaugh et al.
2005) and β-glucosidase is directly involved in soil C dynamics.
Alkaline phosphatase showed consistently lower levels of activity in the nitrogen
treated plots compared to the no nitrogen treatment. This trend may be a result of
nitrogen addition creating a competitive disadvantage to organisms that secrete alkaline
phosphatase.
Response to tillage type
Soil enzymes can be accurate and sensitive indicators to agricultural management
treatments (Deng & Tabatabai, 1996, Dick 1997, Burns and Dick 2002) including the
short-term effects of tillage practices (Kandeler et al. 1999). However, this hasn’t always
been shown to be the case. Conversely, studies have shown that soil enzymes do not
always respond to changes in tillage (Mina et al. 2008). In this study, tillage had no
impact on the activity of any of the extracellular soil enzymes measured within seasons or
69
overall in this study. The lack of a functional response to tillage treatments could have
been affected by the soil composition of the agricultural plots in this study or the
cropping system studied. Tietjen showed that clay-enzyme complexes can reduce the
activity of alkaline phosphatase but increased the activity of β-glucosidase. Clay
particles can also indirectly affect the activity of various soil enzymes forming bonds to
other cationic particles which can alter the pH affecting the optimal activity or interact
with the enzymes themselves (Boyd and Mortaland 1990). The affects of tillage
treatment on the functional responses in the soil measured by enzymatic activity may
indeed be there but outweighed by the effect of the formation of clay-enzyme complexes.
Influence of nutrients and other enzymes on enzymatic activity levels
Soil organic matter (OM) had the broadest correlation with enzyme activity
levels, significantly correlating with phosphodiesterase, β-glucosaminidase, alkaline
phosphatase, and α-galactosidase, the most notable being β-glucosaminidase (33%) and
α-galactosidase (43%). This result is consistent with other research which has shown
OM to be a driving force in N mineralization and C cycling (Fontaine et al. 2003),
processes directly affected by β-glucosaminidase and α-galactosidase (Dick 1997). This
indicates that OM levels may be an appropriate measure of microbial functionality
irrespective of cropping management.
Conclusion
Irrigation had an affect on enzymatic activity. phosphodiesterase and β-
glucosaminidase activity was greater under constant irrigation while others did not
70
exhibit any response to irrigation schedule. The differential response suggests that
composition of the soil microbial community changed in response to irrigation. Nitrogen
fertilization directly impacted the activity of β-glucosaminidase, an enzyme directly
involved in N mineralization and cycling dynamics. Nitrogen inputs significantly
increased short-term β-glucosidase activity in the fall seasons. This is mostly likely due
to a secondary effect due to the fact that β-glucosidase is involved in proving C for soil
microbes and not N cycling. Tillage was not found to significantly impact the activity
levels of the measured soil enzymes. This contradicts most of the previous research.
This result may be due to other variables having stronger affects on enzymatic activity
levels than tillage or that the impacts of tillage in this system were short-lived. As seen in
other studies, soil organic matter appears to be the best measure of the level of soil
functionality and microbial dynamics.
The outcome of this study was the determination of the importance of irrigation to
peanut cropping systems in a semi-arid environment. Full irrigation is needed to
maximize the activity and abundance of the soil microbial community. Furthermore,
attempts to conserve water and reduce water quantities will not only hinder the soil
microbial activity and abundance in that particular growing season but will also
negatively affect them in subsequent growing seasons even if full irrigation is re-
implemented. Strip-tillage was not shown to have any negative impacts on microbial
dynamics in comparison and therefore should be considered a better tillage option
because of the reduced soil erosion and energy required associated with this practice.
71
Nitrogen addition had minimal effects on the system and appears to be an unnecessary
expenditure when growing peanut crops in this particular system.
72
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75
Table 3.1. P-values for comparisons of the three different irrigation treatments imposed
in 2008 for a peanut cropping system. Values were considered significant at p ≤ 0.05.
Significant values are indicated in bold. Note [Phospho= phosphodiesterase,
Glucosamini= β-glucosaminidase, Glucosidase= β-glucosidase, Alkaline= alkaline
phosphatase, Alpha= α-galactosidase, Full= full irrigation, LSD= late season drought,
ESD= early season drought].
Phospho Glucosamini Glucosidase Alkaline Alpha

Full vs. LSD 0.55 0.07 0.08 0.44 0.91
Full vs. ESD 0.88 0.89 0.05 0.16 0.81
LSD vs.ESD 0.45 0.05 0.82 0.03 0.90
76
Table 3.2. Correlation matrix comparing soil extractable nutrients to extracellular soil
enzyme activity from soil samples collect under peanut cultivation for two growing
seasons of 2007-2008. All numbers represent r calculated using Pearson Correlations.
The r values range from 1 to -1 with being a 100% positive correlation and -1 a 100%
negative correlation. Significant r values are indicated in bold. Note [Phosph=
Alkaline= alkaline phosphatase, Alpha= α-galactasidase, NO3 = soil extractable nitrate,
NH4= soil extractable ammonium, P= soil extractable phosphorus, K= soil extractable
potassium, Mg= soil extractable magnesium, Ca= soil extractable calcium, S= soil
extractable sulfur, B= soil extractable boron, Zn= soil extractable zinc, Mn= soil
extractable manganese, Fe= soil extractable iron, Cu= soil extractable copper, CEC=
cation exchange, OM= soil organic matter].
NO NH CE
3 4 P K Mg Ca S B Zn Mn Fe Cu C OM
Phosph
o 0.04 0.43 0.02 0.13 0.19 0.29 0.24 0.24 0.04 0.22 0.13 0.18 0.26 0.21
Gucomi 0.02 0.11 0.03 0.10 0.02 0.10 0.11 0.01 0.06 0.05 0.01 0.10 0.08 0.33
Glucosi
d 0.16 0.13 0.04 0.01 0.07 0.08 0.18 0.25 0.09 0.03 0.01 0.08 0.04 0.01
Alkalin
e 0.06 0.18 0.09 0.13 0.15 0.08 0.08 0.10 0.16 0.11 0.22 0.16 0.03 0.29
Alpha 0.05 0.15 0.11 0.18 0.27 0.25 0.03 0.18 0.22 0.36 0.24 0.27 0.17 0.43
77
Table 3.3. Correlation matrix between all extracellular soil enzymes collected under
peanut cultivation throughout the growing seasons of 2007-2008. All numbers represent r
calculated using Pearson Correlations. The r values range from 1 to -1 with being a
100% positive correlation and -1 a 100% negative correlation. Significant r values are
indicated in bold. Note [Phosph= phosphodiesterase, Glucomini= β-glucominidase,
Glucosidase= β-glucosidase, Alkaline= alkaline phosphatase, Alpha= α-galactasidase].
Phospho Gucomini Glucosidase Alk Alpha

Phospho 1.00
Gucomini 0.03 1.00
Glucosidase 0.18 0.23 1.00
Alk -0.06 0.24 0.17 1.00
Alpha -0.16 0.34 0.16 0.36 1.00
78
Figure 3.1. Plot design for established to study the effects of irrigation, nitrogen
fertilization, and tillage on soil microbial dynamics and community structure associated
with a peanut cropping system in semi-arid west-Texas in year 2007. The numbers
indicated refer to the different zones. Tillage treatment and zone alignment remained the
same in 2008. Irrigation and nitrogen treatments differed in plots 1, 2, 5, and 6 in 2008.
79
Figure 3.2. The effect of late season drought (LSD) compared to full irrigation (Full) on
phosphodiesterase activity levels associated with a peanut cropping system. (a)Total
phosphodiesterase activity levels across two years, 2007-2008. Values are means± S.E.,
n=64. (b) Phosphodiesterase levels in each of the summer and fall seasons in both 2007
and 2008. Values are means ± S.E, n= 16. Groups statistically (p ≤ 0.05) different are
indicated with different letters above mean bars.
Full
a
LSD
80 a
(mg Pn kg soil h )
-1
60
-1
40
20
a.
Full
120 LSD
100 a
ab
(mg kg-1 soil h-1)
abc ab
bc
80 c
d
d
60
40
20
b.
80
β-glucosaminidase activity levels within a peanut cropping system. (a)Total β-
glucosaminidase activity across two growing seasons, 2007-2008. Values are means±
S.E., n= 64. (b) β-glucosaminidase levels in each of the summer and fall seasons in both
2007 and 2008. Values are means± S.E., n=16. Groups statistically different are
18 a
Full
16 LSD
a
-Glucosaminidase Activity
14
(mg PN kg soil h )
-1
12
-1
10
a.
Full
25 LSD a
ab
-Glucoscaminidase
(mg PN kg soil )
20
-1
ab
bc bc
-1
15 bc c
c
10

b.
81
β-glucosidase activity levels within a peanut cropping system. (a)Total β-glucosidase
activity levels across two growing seasons, 2007-2008. Values are means± S.E., n= 64.
(b) β-glucosidase levels in each of the summer and fall seasons in both 2007 and 2008.
Values are means± S.E., n=16. Groups statistically (p ≤ 0.05) different are indicated with
a
80 Full
LSD
b
-Glucosidase Activity
(mg PN kg soil h )
-1
60
-1
40
20
a.
Full
120 LSD
a
100
(mg PN kg soil h )
-1
ab
80 bc bc
bc
-1
c
c c
60
40
20

b.
82
alkaline phosphatase activity levels from a peanut cropping system. (a)Total alkaline
phosphatase activity levels from 2007-2008. Values are means± S.E., n= 64. (b)
Alkaline phosphatase levels in each of the summer and fall seasons in both 2007 and
2008. Values are means± S.E., n=16. Groups statistically (p ≤ 0.05) different are
a
Full a
140 LSD
120
(mg Pn kg soil h )
-1
100
-1
80
60
40
20
a.
210
Full
LSD
180
a a
a a a
(mg PN kg soil h )
150
-1
a a a
120
-1
90
60
30

b.
83
α-galactosidase activity levels from a peanut cropping system. (a) Total α-galactosidase
activity levels from 2007-2008. Values are means± S.E., n= 64. (b) α-galactosidase
levels in each of the summer and fall seasons in both 2007 and 2008. Values are means±
S.E., n=16. Groups statistically (p ≤ 0.05) different are indicated with different letters
above mean bars.
16 Full
LSD
14 a a
-Galactosidase Activity
-1)
12
(mg PN kg soil h
10
-1
a.
20 Full
LSD
a
a
a
(mg PN kg soil h )
ab
-1
15 abc
-Galactosidase
bc
bc
-1
10

b.
84
Figure 3.7. (a) Discriminate function analysis (DFA) comparing the differences of the
three different irrigation treatment imposed in 2008 on a peanut cropping system using
different enzyme activity (phosphodiesterase, β-glucosaminidase, β-glucosidase, alkaline
phosphatase, α-galactosidase) levels as the parameters. (b) Vector correlations indicating
correlations between irrigation treatments and enzyme activities. Correlations are
indicated by the smallest angle between two vectors. Ex 0°= 100% positive correlation,
90°= no correlation, and 180°= 100% negative correlation. Note [1= full irrigation, 2=
late season drought, 3= full season drought, Phos= phosphodiesterase, Gmini= β-
glucosaminidase, Gdase= β-glucosidase, AlkPho= alkaline phosphatase, Agal= α-
galactosidase].
a.
b.
85
Figure 3.8. Discriminate function analysis (DFA) comparing no nitrogen application and
nitrogen application to a peanut cropping system using phosphodiesterase, β-
glucosaminidase, β-glucosidase, alkaline phosphatase, and α-galactosidase activity levels
as the parameters.
86
Figure 3.9. The effects of nitrogen application on phosphodiesterase activity levels from
a peanut cropping system. (a)Total phosphodiesterase activity levels across two growing
years, 2007-2008. Values are means± S.E., n= 32. (b) Phosphodiesterase levels in each
of the summer and fall seasons in both 2007 and 2008. Values are means± S.E., n=16.
bars.
No nitrogen
nitrogen
100 a
a
(mg PN kg soil h )
-1
80
-1
60
40
20
a.
120 No nitrogen
nitrogen
100 a a
a
a
(mg Pn kg soil h )
-1
b
80
-1
60
40
20
Summer 07 Fall 07
b.
87
Figure 3.10. The effect of no nitrogen application compared to nitrogen application

treatments on β-glucosaminidase activity levels from a peanut cropping system. (a)Total
β-glucosaminidase activity levels across two growing years, 2007-2008. Values are
means± S.E., n= 32. (b) β-glucosaminidase levels in each of the summer and fall seasons
in both 2007 and 2008. Values are means± S.E., n=16. Groups statistically (p ≤ 0.05)
20 No nitrogen
nitrogen a
a
15
(mg PN kg soil h )
-1 -1
10
a.
25 No nitrogen
nitrogen
a
20
(mg Pn kg soil h )
-1
a
15 a a
-1
10
Summer 07 Fall 07
b.
88
Figure 3.11. The effect of nitrogen on β-glucosidase activity levels from a peanut
cropping system. (a)Total β-glucosidase activity levels from 2007-2008. Values are
means± S.E., n= 32. (b) β-glucosidase levels in each of the summer and fall seasons
across two growing seasons, 2007-2008. Values are means± S.E., n=16. Groups
statistically (p ≤ 0.05) different are indicated with different letters above mean bars.
No nitrogen
nitrogen
a
80 a
(mg PN kg soil h )
-1
60
-1
40
20
a.
a
100 No nitrogen
nitrogen
b
80
(mg PN kg soil h )
bc
-1
c
-1
60
40
20
Summer 07 Fall 07
b.
89
Figure 3.12. The effect of no nitrogen application compared to nitrogen application

treatments on alkaline phosphatase activity levels from a peanut cropping system.
(a)Total alkaline phosphatase activity levels from 2007-2008. Values are means± S.E.,
n= 32. (b) Alkaline phosphatase levels in each of the summer and fall seasons across
two growing years, 2007- 2008. Values are means± S.E., n=16. Groups statistically (p ≤
0.05) different are indicated with different letters above mean bars.
a
No nitrogen
160 nitrogen
a
140
(mg PN kg soil h )
-1
120
100
-1
80
60
40
20
a.
210 No nitrogen
nitrogen
a
180
a
a
(mg PN kg soil h )
-1
150 a
120
-1
90
60
30
Summer 07 Fall 07
b.
90
Figure 3.13. The effect of nitrogen application compared to nitrogen application

treatments on α-galactosidase activity levels in a peanut cropping system. (a)Total α-
galactosidase activity levels from 2007-2008. Values are means± S.E., n= 32. (b) α-
Galactosidase levels in each of the summer and fall seasons across two growing years,
2007 -2008. Values are means± S.E., n=16. Groups statistically (p ≤ 0.05) different are
14 No nitrogen
nitrogen
a
12 a
(mg PN kg soil h )
-1
10
-1
a.
18 No nitrogen
nitrogen
16
a
14 a
(mg PN kg soil h )
-1
12 a
a
-1
10
Summer 07 Fall 07
b.
91
Figure 3.14. (a) Discriminate function analysis (DFA) comparing the differences of
nitrogen and no nitrogen application in different seasons in 2007 using different enzyme
activity (phosphodiesterase, β-glucosaminidase, β-glucosidase, alkaline phosphatase, α-
galactosidase) levels as the parameters. Note [1= no nitrogen summer, 2= nitrogen
summer, 3= no nitrogen fall, 4= no nitrogen fall, Phos= phosphodiesterase, Gmini= β-
glucosaminidase, Gdase= β-glucosidase, AlkPho= alkaline phosphatase, Agal= α-
galactosidase].
a.
92
Figure 3.14. (b) Vector correlations indicating correlations between the different
nitrogen treatments in summer and fall and enzyme activities. Correlations are
90°= no correlation, and 180°= 100% negative correlation. Note [1= no nitrogen summer,
2= nitrogen summer, 3= no nitrogen fall, 4= no nitrogen fall, Phos= phosphodiesterase,
Gmini= β-glucosaminidase, Gdase= β-glucosidase, AlkPho= alkaline phosphatase, Agal=
α-galactosidase].
b.
93
Figure 3.15. The effects of strip tillage compared to conventional tillage on

phosphodiesterase activity levels in a peanut cropping system. (a) Total
phosphodiesterase activity levels from 2007-2008. Values are means± S.E., n= 96. (b)
Phosphodiesterase levels in each of the summer and fall seasons across two growing
years, 2007- 2008. Values are means± S.E., n=24. Groups statistically (p ≤ 0.05)
Conventional
Strip a
a
80
(mg PN kg soil h )
-1
60
-1
40
20
a.
Conventional
Strip
100 a
a a
a
a a
(mg PN kg soil h )
-1
80
b
b
-1
60
40
20

b.
94
Figure 3.16. The effect of strip tillage compared to conventional tillage treatments on β-
glucosaminidase activity levels in a peanut cropping system. (a) Total β-glucosaminidase
activity levels from 2007-2008. Values are means± S.E., n= 96. (b) β-glucosaminidase
levels in each of the summer and fall seasons across two growing years, 2007- 2008.
18 Conventional
Srip a
16 a
14
(mg PN kg soil h )
-1
12
-1
10
a.
22 Conventional
Strip a
20
ab
18 ab
ab ab
ab
(mg PN kg soil h )
16
-1
ab b
14
-1
12
10

b.
95
Figure 3.17. The effect of strip tillage compared to conventional tillage on β-glucosidase
activity levels in a peanut cropping system. (a) Total β-glucosidase activity levels from
2007-2008. Values are means± S.E., n= 96. (b) β-glucosidase levels in each of the
summer and fall seasons across two growing years, 2007- 2008. Values are means±
S.E., n=24. Groups statistically (p ≤ 0.05) different are indicated with different letters
above mean bars.
80 Conventional
a
Strip a
60
(mg PN kg soil h )
-1 -1
40
20
a.
Conventional a
Strip a a
80 ab
b
b
(mg PN kg soil h )
b b
-1
60
-1
40
20

b.
96
Figure 3.18. The effect of strip tillage compared to conventional tillage on alkaline
phosphatase activity levels in a peanut cropping system. (a) Total alkaline phosphatase
activity levels from 2007-2008. Values are means± S.E., n= 96. (b) Alkaline
phosphatase levels in each of the summer and fall seasons across two growing years,
2007- 2008. Values are means± S.E., n=24. Groups statistically (p ≤ 0.05) different are
Conventional
160
Strip a a
140
120
(mg PN kg soil h )
-1
100
-1
80
60
40
20
a.
200 Conventional
Strip
180 a
ab
Alkaline Phosphatae Activity
abc
160 abc
bc
(mg PN kg soil h )
bc bc
-1
140 c
120
-1
100
80
60
40
20

b.
97
Figure 3.19. The effect of strip tillage compared to conventional tillage on α-

galactosidase activity levels in a peanut cropping system. (a) Total α-galactosidase
activity levels from 2007-2008. Values are means± S.E., n= 96. (b) α-Galactosidase
levels in each of the summer and fall seasons across two growing years, 2007- 2008.
16 Conventional
Strip
14 a
a
12
(g PN kg soil h )
-1
10
-1
a.
18
Conventional
Strip a a
16 ab ab
14 bc
(mg PN kg soil h )
bc
-1
12
c c
-1
10

b.
98
Figure 3.20. (a) Discriminate function analysis (DFA) comparing the differences of
conventional and strip tillage in 2007 and 2008 using different enzyme activity levels
and nutrient levels as the parameters. Note [1= conventional 07, 2= strip 07, 3=
conventional 08, 4= strip 08, Phosph= phosphodiesterase, Glucomini= β-
glucosaminidase, Glucosidase= β-glucosidase, Alkaline= alkaline phosphatase, Alpha=
α-galactasidase, NO3 = soil extractable nitrate, NH4= soil extractable ammonium, P= soil
extractable phosphorus, K= soil extractable potassium, Mg= soil extractable magnesium,
Ca= soil extractable calcium, S= soil extractable sulfur, B= soil extractable boron, Zn=
soil extractable zinc, Mn= soil extractable manganese, Fe= soil extractable iron, Cu= soil
a.
99
Figure 3.20. (b) Vector correlations indicating correlations between the different
nitrogen treatments in summer and fall and enzyme activities. Correlations are
90°= no correlation, and 180°= 100% negative correlation. Note [1= conventional 07, 2=
strip 07, 3= conventional 08, 4= strip 08, Phosph= phosphodiesterase, Glucomini= β-
glucosaminidase, Glucosidase= β-glucosidase, Alkaline= alkaline phosphatase, Alpha=
α-galactasidase, NO3 = soil extractable nitrate, NH4= soil extractable ammonium, P= soil
extractable phosphorus, K= soil extractable potassium, Mg= soil extractable magnesium,
Ca= soil extractable calcium, S= soil extractable sulfur, B= soil extractable boron, Zn=
soil extractable zinc, Mn= soil extractable manganese, Fe= soil extractable iron, Cu= soil
b.
100
Figure 3.21. Season variation in soil extractable nitrate (NO3) levels across all
treatments associated with a peanut cropping system. Values are means± S.E., n= 48.
bars.
30 a
Soil extractable N03 (ppm)
25
20
b
15 b
10
c
101
Figure 3.22. Seasonal variation in soil extractable ammonium (NH4) levels across all
treatments associated with a peanut cropping system. Values are means± S.E., n= 48.
bars.
18.0
a
16.0
Soil extractable NH4 (ppm)
14.0
12.0
10.0
2.0
1.5 b
b
b
1.0
102
Figure 3.23. Seasonal variation in soil phosphorus (P) levels associated with a peanut
cropping system. Values are means± S.E., n= 48. Groups statistically (p ≤ 0.05)
70 a
a
60
b
50
Soil P (ppm)
40
30
20
10
103
Figure 3.23. Seasonal variation in soil organic matter (OM) levels associated with a
peanut cropping system. Values are means± S.E., n= 48. Groups statistically (p ≤ 0.05)
1.6
a
1.4
b b
% Soil Organic Matter
1.2 b
1.0
0.8
0.6
0.4
0.2
104

Agricultural Management Effects On Soil Microbial Dynamic Under Peanut Cropping Systems in Semi

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Agricultural Management Effects on Soil Microbial Dynamic Under Peanut Cropping

Systems in Semi-Arid Soil

Brandon Melester, B.S.

Submitted to the Graduate Faculty

Dr. John C. Zak

Dr. Veronica Acosta-Martinez

Dr. Michael San Francisco

Dr. Fred Hartmeister

LIST OF TABLES ............................................................................................................... vi

LIST OF FIGURES ............................................................................................................ vii

I. INTRODUCTION TO MICROBIAL DYNAMICS, MICROBIAL

II. EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID

III. EFFECTS OF MANAGEMENT PRACTICES IN A SEMI-ARID

Recent agricultural practices have been shifting towards low-input sustainable

decreasing overall energy inputs.

order to improve those numbers without increasing input, it will be imperative to

The effects of tillage, nitrogen, and irrigation management within a peanut

assays of phosphodiesterase, β-glucosaminidase, β-glucosidase, alkaline phosphatase, and

agricultural management on soil microbe functioning.

significantly increases β-glucosidase activity, bacterial abundance, and MBC. Irrigation

reductions were shown to produce “hangover effects” in which microbial activity,

lower in subsequent seasons. Nitrogen had differential effects on microbial dynamics.

2.1. Correlation matrix comparing soil extractable nutrients to Microbial Biomass

2.2. Correlation matrix comparing soil extracellular enzymes to Microbial Biomass

2.1. Plot design of study site (2007) ....................................................................................................36

2.4. The effect of all irrigation levels on Microbial Biomass Carbon

2.5. The effect of nitrogen on Microbial Biomass Carbon

2.6. Monthly changes in nutrient dynamics and microbial functionality

2.6. Monthly changes in nutrient dynamics and microbial functionality

2.7. Changes in Microbial Biomass Carbon across growing seasons 2007-2008

2.8. Relative bacterial and fungal abundances

2.9. Relative bacteria and fungi groups abundances

2.10. Effects of nitrogen on relative bacteria abundances

2.11. Effects of nitrogen on relative fungi abundance

2.12. Effects of irrigation on relative Gram-positive bacteria abundance

2.13. Effects of irrigation on relative Gram-negative bacteria abundance

3.1. Study site (2007) ...............................................................................................................................79

3.2. Effects of irrigation of phosphodiesterase activity

3.3. Effect of irrigation on β-glucosaminidase activity

3.4. Effects of irrigation on β-glucosidase activity

3.5. Effects of irrigation on alkaline phosphatase activity

3.6. Effects of irrigation on α-galactosidase activity

3.7. Comparison of 3 irrigation levels on collective enzyme activity

3.8. Discriminate function analysis on effect of nitrogen on collective enzyme activity.....86

3.9. Effect of nitrogen on phosphodiesterase activity

3.10. Effect of nitrogen on β-glucosaminidase activity

3.11. Effect of nitrogen on β-glucosidase activity

3.12. Effect of nitrogen on alkaline phosphatase activity

3.13. Effect of nitrogen on α-galactosidase activity

3.14. Seasonal nitrogen effect on collective enzyme activity

3.14. Seasonal nitrogen effect on collective enzyme activity

3.15. Effect of tillage on phosphodiesterase activity

3.16. Effect of tillage on β-glucosaminidase activity

3.17. Effect of tillage on β-glucosidase activity

3.18. Effect of tillage on alkaline phosphatase activity

3.19. Effect of tillage on α-galactosidase activity

3.21. Season variation in soil extractable nitrate .............................................................. 101

3.22. Seasonal variation in soil extractable ammonium ................................................. 102

3.23. Season variation in soil phosphorus ........................................................................ 103

3.24. Seasonal variation in soil organic matter .............................................................. 104

INTRODUCTION TO SOIL MICROBIAL DYNAMICS, MICROBIAL

effects on the performance of crops are of great agricultural importance. Moreover