10.5061/DRYAD.KWH70RXZP
Xiao, Rui
0000-0002-0075-1774
Nanjing Agricultural University
Qiu, Yunpeng
North Carolina State University
Tao, Jinjin
Nanjing Agricultural University
Zhang, Xuelin
North Carolina State University
Chen, Huaihai
North Carolina State University
Reberg-Horton, S. Chris
North Carolina State University
Shi, Wei
North Carolina State University
Shew, H. David
North Carolina State University
Zhang, Yi
Nanjing Agricultural University
Hu, Shuijin
Nanjing Agricultural University
Data from: Biological controls over the abundances of terrestrial ammonia
oxidizers
Dryad
dataset
2020
ammonia-oxidizing archaea
ammonia-oxidizing bacteria
Nitrification
soil carbon to nitrogen ratios
National Natural Science Foundation of China
https://ror.org/01h0zpd94
31600383
Ministry of Science and Technology of the People's Republic of China
https://ror.org/027s68j25
2017YFC0503902
a China Scholarship Council scholarship to YunpengQiu
CSC NO. 201306320137
NIFA, USDA
USDA-2012-02978-230561
2019-10-11T00:00:00Z
2019-10-11T00:00:00Z
en
41660 bytes
4
CC0 1.0 Universal (CC0 1.0) Public Domain Dedication
Aim: Ammonia-oxidizing archaea (AOA) and bacteria (AOB) are the primary
agents for nitrification, converting ammonia (NH4+) into nitrate (NO3-)
and modulating plant nitrogen (N) utilization and terrestrial N retention.
However, there is still lack of a unifying framework describing the
patterns of global AOA and AOB distribution. In particular, biotic
interactions are rarely integrated into any of the conceptual models.
Location: World-wide. Time period: 2005-2016. Major taxa studied:
Ammonia-oxidizing archaea and ammonia-oxidizing bacteria. Methods: A
meta-analysis and synthesis was conducted to obtain a general picture of
global AOA and AOB distribution and identify the primary driving factors.
A microcosm experiment was then conducted to assess effects of relative
carbon to nitrogen availability for heterotrophic microbes on AOA and AOB
in two distinct soils. A mesocosm experiment was further carried out to
characterize the effects of plant roots and their arbuscular mycorrhizal
fungi (AMF) on AOA and AOB abundances using hyphae- or root-ingrowth
techniques. Results: Our meta-analysis showed that soil carbon to
nitrogen (C/N) ratios explained the most variance in AOA and AOB
abundances, although soil pH had a significant effect. Experimental
results demonstrated that high cellulose and mineral N inputs increased
total microbial biomass and microbial activities, but inhibited AOA and
AOB, suggesting microbial inhibition of AOA and AOB. Also, AMF and roots
suppressed AOA and AOB, respectively. Main conclusions: Our study
provided convincing evidence illustrating that relative carbon to nitrogen
availability can dominate the abundances of AOA and AOB. Our experimental
results further validated that biotic competitions among plants,
heterotrophic microbes and ammonia oxidizers for substrate N predominantly
control AOA and AOB abundances. Together, these findings provide new
insights into the role of abiotic and biotic factors in modulating
terrestrial AOA and AOB abundances and their potential applications for
management of nitrification in an increasing reactive N world.
Experiment 1. A microcosm study examining the effects of different ratios
of carbon and nitrogen inputs on AOA and AOB. This experiment was designed
to determine the effect of resource C/N ratios on AOA and AOB through
quantifying the responses of AOA and AOB to differences in the relative
availability of organic C to mineral N. Soil samples were collected from
two distinct soils: a pine plantation soil (PINE) in Nanjing 32°03'
N, 118°46'E, Jiangsu Province, and a vegetable field soil (VF) in
Nantong (32° 01'N, 120°51'E), Jiangsu Province. The PINE and VF
soils contained 0.9 and 1.1 g.kg-1 of total N, 37.8 and 13.9 g.kg-1 of
total organic C, and had pH values of 4.08 and 7.50, respectively. Field
soil samples were sieved through a 2 mm sieve before being used for the
incubation experiment. Each soil was amended with organic C (cellulose)
and inorganic N [(NH4)2SO4] at four C/N ratios as follows: 10, 25, 50, and
100. For each soil, two levels of N inputs were designed (Low N at 75 and
high N at 200 mg N kg-1 soil) and four levels of cellulose were added at
each N level (C/N ratios at 10, 25, 50 and 100, respectively). These
resulted in 16 treatment combinations with 18 replicates per treatment (2
soils × 2 N levels × 4 C/N ratios ×18 replicates = 288 microcosm jars),
plus two controls (that is, soils with no C and no N amendments: 2 soils
×18 replicates = 36 microcosms). The soils were pre-incubated for one week
and the treatments were then applied. Cellulose (powder) were weighed and
well mixed into 50.0 g soil (dry equivalent) and placed into a 250 mL jar
with a surface area of 22 cm2. (NH4)2SO4 was applied in a liquid form. The
soil moisture was adjusted to 65% of the water holding capacity for each
soil. All the microcosm jars were randomly placed inside the room with a
room temperature (air-conditioned) at 22-25°C. The soil moisture was
maintained by periodically weighing the microcosm jars and adding
(spraying) distilled water with a syringe to compensate for any weight
loss (every 3 days). The soil samples were incubated for about three
months. To ensure sufficient O2, the cover for each jar was opened for 10
min each day for the 1st two months and every 4 days in the 3rd month. The
jar opening scheme with different frequencies at different times was
designed to ensure sufficient O2 for AOA and AOB, because high soil
respiration rate at the early stages consumed more O2 (as shown in
Supporting Information Figure S7). A randomly selected subset of
treatments (three replicates) was used for monitoring microbial
respiration. Sub-samples of soils (each jar representing one sub-sample)
were destructively taken at 3, 7, 14, 21, 35, 47, 60, 75, and 90 days
after the application of the treatments to determine the amoA gene copies
of the AOA and AOB populations, soil microbial biomass C (MBC), and
extractable N (NH4+ and NO3-). Soil samples were taken at 7, 35, 60 and 90
days after the application of the treatments and soil DNA were extracted
to determine the abundances of bacteria and fungi (more details below).
Soil MBC was measured using the chloroform-fumigation extraction method
(Vance, Brookes, & Jenkinson, 1987). NH4+ and NO3- concentrations
were extracted with 2 M KCl and detected on a flow injection analyzer
(Tecator Inc., Sweden). Soil respiration was measured by an
incubation-alkaline absorption method (Hu & van Bruggen, 1997).
Fresh soil (0.30 g) was extracted for DNA using MoBio Power soil TMDNA
isolation kits (San Diego, CA) according to the manufacturer’s
instructions. Real-time quantitative PCR was performed to determine copy
numbers of amoA gene of AOA and AOB, and to quantify bacterial 16S
ribosomal DNA and fungal 18S ribosomal DNA in the total DNA of the soil
sample using iCycler iQ 5 thermocycler (BioRad Laboratories, Hercules, CA,
USA). Primer sets amoA-1F/amoA-2R (Rotthauwe, Witzel, & Liesack,
1997), Arch-amoAF/Arch-amoAR (Francis, Roberts, Beman, Santoro, &
Oakley, 2005), Eub338/Eub518 (Fierer, Jackson, Vilgalys, &
Jackson, 2005) and ITS1f /5.8s (Fierer et al., 2005) were used for the
amplification of bacterial amoA gene, archaeal amoA gene, bacterial 16S
ribosomal DNA, and fungal 18S ribosomal DNA fragments, respectively. Real
time PCR was performed using the temperature profiles describes in
Supporting Information Table S2. Standard curves for real time PCR assays
was made as described in Di et al. (2009) for AOA and AOB and Fierer et
al. (2005) for bacteria and fungi, respectively. 2. A mesocosm experiment
assessing the impact of plant roots and arbuscular mycorrhizal fungi (AMF)
on AOA and AOB This experiment was conducted in the greenhouse at North
Carolina State University (NCSU), Raleigh, North Carolina, USA. Two
sources of soils [organically managed soil (OM) and conventionally-managed
soil (conventional)] were employed to examine how plant roots and their
associated AMF affected the abundances of AOA and AOB. These soils were
collected from two farming systems at the Center for Environmental Farming
Systems at NC State University (35°22'N, 78°02'W) that was
established in 1999 in Goldsboro, North Carolina, USA. While the
conventional fields had been applied with mineral N, the
organically-managed fields had received organic manures and cover crops
only since 1999 (Mueller et al., 2006; Tu, Louws, et al., 2006). Both
fields were planted with corn (Zea mays L.) prior to soil collection in
2014. Soil samples were partially air-dried and sieved through a 4 mm
sieve. The OM and conventional soils contained 24 and 21 μg.g-1 of
inorganic N, 178 and 42 μg.g-1 of labile C, and had pH values of 6.50 and
5.50, respectively. We employed hyphae- or root-ingrowth techniques to
examine how the presence of AMF only or roots with their associated AMF
affects AOA and AOB abundances in the two soils described above.
Plexi-glass mesocosms were used to manipulate roots and/or mycorrhizae,
and each microcosm was divided into six compartments with each
compartment measuring 13×14×15cm (width × depth × height) (Tu, Booker, et
al., 2006). Three compartments in a row were designated as HOST
compartments (containing host plants inoculated with AM fungi) and the
three adjacent compartments were designated TEST compartments to assess
the effects of AMF and/or roots on AOA and AOB. The HOST and TEST
compartments were separated by a replaceable mesh fabric panel
(Tetko/Sefar mesh, Sefar America, NY) that prevented plant roots or both
roots and AMF hyphae from growing into the TEST compartments,
respectively. Consequently, this leads to three treatments of AMF and/or
roots based on whether the TEST soil is accessible by roots and/AMF from
plants in the HOST compartment: (1) the control with no penetration of
plant roots and AMF to the TEST soil (CK), (2) penetration of AMF hyphae
to the TEST soil (AMF), and (3) penetration of both AMF hyphae and plant
roots to the TEST soil (Root). Three different mesh screens, that is, 0.45
µm, 20 µm, and 1.6mm, were used for the Control, the AMF treatment, and
the Root treatment, respectively. The AMF inoculum was a mixture of
multiple AM fungal species that were trap-cultured from an agricultural
soil, collected from the Center for Environmental Farming Systems at NC
State University, and was then pot-cultured to increase fungal biomass.
Twelve AM fungal species were identified and characterized according to
the International Culture Collection of (Vesicular) Arbuscular Mycorrhizal
Fungi (INVAM) (Tu, Booker, et al., 2006). The AMF inoculum consisted of
culture media containing spores, hyphae, and colonized root pieces. Each
compartment was filled with 3.5 kg soil mixed with 100g AM fungal
inoculum. Organically managed soil was applied with chicken manure at a
rate of 12.5 t ha-1, while conventional soil was supplied with urea at a
rate of 180 kg N ha-1. The chicken manure (Microstart60, Perdue
AgriRecycle LLC) was a pellet with 3-2-3 for N, P2O5 and K2O, finely
grounded and mixed with soil before the corn seeds were sown. For the
conventional soil, half of urea fertilizer (dissolved in deionized water)
was applied to the soil prior to seed sowing and the rest half was applied
4 weeks later. Each time, urea was and then applied to the soil. All sides
of all mesocosm units were covered in aluminum foil to block the lights
throughout the experimental period. Four corn seeds (Zea mays L., variety
F1 Incredible) were sown to each of the HOST compartments on the 4th March
2014 (days after sowing 0 day: DAS0). The mesocosm units were placed in a
temperature-controlled glasshouse by a randomized block design. All of
these treatments were replicated three times. The mean daily temperature
was 28 ± 2 oC. To ensure 16-h daylight for corns, overhead lights (400 W,
Son-T Agro) were used during the early evening hours. The
photosynthetically active radiation flux was 505 m mol m-2 s-1 with a day
length of 16 h. The plants were watered with deionized water as needed.
The corn plants were allowed to grow for 66 days. All plants were then
harvested, dried and weighed, and subsamples were analyzed for C and N
contents. Soil samples from the TEST compartments were collected for
analyses of AOA and AOB. Fresh soil (0.50 g) was extracted for nucleic
acid using FastDNA SPIN kit (MP Bio, Solon, OH, USA) according to the
manufacturer’s instructions. Quantitative real-time PCR was performed on
each soil sample (CFX96 Real-Time PCR Detection System, Bio-Rad, Hercules,
CA, USA) to determine the amoA gene copy numbers of AOA and AOB with the
primer sets CrenamoA23f and CrenamoA616r (Tourna, Freitag, Nicol,
& Prosser, 2008) and amoA-1F/amoA-2R (Rotthauwe et al., 1997),
respectively. Standard curves for real-time PCR assays were made following
serial dilutions of the plasmid DNA method (Qiu et al., 2018).