Nematology in the provisionof soil ecosystem services:nutrients and energy in the
soil food web
Teagasc,Environment Research Centre,Johnstown Castle,Wexford.
Bryan Griffiths
Acknowledgements
Jane Davidson Fiona Brennan
Jen Kennedy Vincent O’Flaherty
Susan Mitchell Xiaoyun Chen
Dave Roberts Tim Daniell
Jane Wishart Roy Neilson
Michael Bonkowski Nanjing Agricultural Uni.
SCRI sequencing service
Christine Hackett, BIOSS
Date for your diary
2nd International workshop on nematodes as
environmental indicators
5-6th July 2012
Ghent, Belgium
Scope of presentation
Ecosystem services is the new catchy title. Encompasses:
nutrient cycling; carbon sequestration; provision of clean
water; disease suppression.
This talk will concentrate on interactions in the food web
And large-scale applications of nematode community structure
Firstly, a reminder of the soil food web……
Below-ground biomass = 600 sheep ha-1; 1-2 tonnes ha-1How many ‘sheep’ below-ground?
Fauna 10% biomass = 60 sheep ha-1; 1-200 kg ha-1
The soil food web(de Ruiter et al. 1993, J Appl Ecol 30, 95-106)
Roots
PhytophagousNematodes
PredaceousNematodes
PredaceousMites
PredaceousCollembolans
NematodeFeeding Mites
Detritus
Bacteria
Bacteriophagous
NematodesBacteriophagous
Enchytraeids
Mites
Flagellates
Amoebae
SaprophyticFungi
Collembolans
Noncrypto-stigmatic Mites
CryptostigmaticMites
FungivorousNematodes
The soil food web - energy flows
Roots
PhytophagousNematodes
PredaceousNematodes
PredaceousMites
PredaceousCollembolans
NematodeFeeding Mites
Detritus
Bacteria
Bacteriophagous
NematodesBacteriophagous
Enchytraeids
Mites
Flagellates
Amoebae
SaprophyticFungi
Collembolans
Noncrypto-stigmatic Mites
CryptostigmaticMites
FungivorousNematodes
Carbon and nutrients essentially similar in termsof flow through the food web
Ecological efficiency drives nutrient recycling
Nematodes have lower efficiencies (ca. 10%) than protozoa (ca. 40%)
Nematodes Protozoa
1 gincrease in
biomass
Ingest 13 g bacteria Ingest 3 g bacteria
Excrete 0.9 µg N Excrete 0.09 µg N
But not the whole story…
If plants are grown with excess nutrients (i.e. no
nutrient limitation), fauna still increase plant growth
Recent research has related presence of protozoa to
increased number and length of roots
Due to grazing-stimulated production of plant
hormones by rhizosphere bacteria
Conceptual model of protozoan effects on rootgrowth
Bonkowski & Brandt 2003; Bonkowski, 2004 New Phytol. Tansley review
Generation of nematode-enriched soil
5µm mesh 1mm mesh
Mao et al., Soil Biol Biochem 2006; 2007
Generation of nematode-enriched soil
5µm mesh 1mm mesh
27.8 ± 2.8 nematodes g-1 soil 135.1 ± 13.1 nematodes g-1 soil
Generation of nematode-enriched soil
27.8 ± 2.8 nematodes g-1 soil 135.1 ± 13.1 nematodes g-1 soil
5µm mesh 1mm mesh
Plant growth experiment
Tomatoseed
27.8 ± 2.8 nematodes g-1 soil 135.1 ± 13.1 nematodes g-1 soil
5µm 1mm
Nematode treatments
Treatment
1mm 1mm mesh, 135 nematodes g-1
5m 5 mm mesh, 27 nematodes g-1
CE 5 m mesh + 135 C.elegans g-1
Mixed 5 m mesh + 135 mixed nematodes g-1
Auxin (IAA) content of nematode-enriched soil
0
25
50
1mm 5um C.elegans Mixed nematodes
0 5 10μg g-1
dry soil
DAYS
Plant root growth in nematode-enriched soil
30
40
50
1mm 5um "+Ce" "+Nem"
20
30
40
50
1mm 5um "+Ce" "+Nem"
a b a a
a b a a
Root length (cm) Root tips (number)
1mm 5m CE Mixed 1mm 5m CE Mixed
Molecular signals detected in plant roots
Valentine et al., in prep
Conclusions
Bacterial-feeding nematodes contribute to root
development.
Future work will try to identify the ‘genetic control
points’.
Long-term aim to manipulate nematode populations
and root growth in the field.
Bacteria fight back – 2o metabolite repression
Neidig et al 2011 Microb. Ecol 61:853
The soil INTERACTION web
Roots
PhytophagousNematodes
PredaceousNematodes
PredaceousMites
PredaceousCollembolans
NematodeFeeding Mites
Detritus
Bacteria
Bacteriophagous
NematodesBacteriophagous
Enchytraeids
Mites
Flagellates
Amoebae
SaprophyticFungi
Collembolans
Noncrypto-stigmatic Mites
CryptostigmaticMites
FungivorousNematodes
Molecular Biology for ecological studies
Moving ahead rapidly, due to economic
implications of plant-parasitic forms and the
Caenorhabditis elegans genome project.
Micro-arrays available for plant pathogens.
Many labs working on non-pathogenic forms
and information readily available on the web.
Aims
Develop a molecular method of profiling soil
nematode communities (because traditional
methods too time-consuming and skilled)
Validated against current techniques
Apply the methods in an agricultural context,
relating to effects of fertilisation.
Nematode morphology
• A skilled job
• Takes a long time
• Would analysis ofDNA be faster
Molecular approaches
Amplify SSU
Extract DNA
Sieve from 200g soil
48 hr Baermannfunnel extraction
Bead beat &kit purify
PCRamplificationof SSU
Molecular approaches
Amplify SSU
Extract DNA
Clone & sequence
phylogenetics
T-RFLP
Directed T-RFLP
T-RFLP advantages
Directed T-RFLP – peaks related to trophicgroup
Trace and pictures
Long-term effects of P fertilisation
The study area was in South East Ireland, on
which a long-term trial to study P for beef
production has been carried out since 1968,
with changes since 1999.
P applications
Year
Treatment (kg P ha-1 yr-1)
1 2 3
P0 P15 P30
1968-1998 0 15 30
Year
Treatment (kg P ha-1 yr-1)
1 2 3 4 5 6
P0 P0-30 P15 P15-5 P30 P30-0
19681968--19981998
1999-2009
0 00 0 1515 1515 3030 3030
0 30 15 5 30 0
Nematode abundance marginally affected
bbab
b
a
b
abcab
bc
aabc
c
bb
a
b
b
b
0.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
P0-0 P0-30 P30-30 P30-0 P15-15 P15-5
Ne
ma
tod
eN
um
be
rs(/
gD
ryso
ils)
Large(>125µm) Small(125-53µm) total
Morphological – channel ratio increases with P
c
b
aa
aa
60
65
70
75
80
85
90
P0-0 P0-30 P30-30 P30-0 P15-15 P15-5
NC
R(%
)
Molecular – bacterial-feeders increase with P
40
50
60
70
80
90
0-0 0-30 30-30 30-0 15-15 15-5
%b
ac
teri
al-
fee
de
rs
a
a
b a,b
b
b
Morphological vs Molecular
c
b
aa
aa
60
65
70
75
80
85
90
P0-0 P0-30 P30-30 P30-0 P15-15 P15-5
NC
R(%
)
40
50
60
70
80
90
0-0 0-30 30-30 30-0 15-15 15-5
%b
ac
teri
al-
fee
de
rs
a
a
b a,b
bb
Time required:
9 days vs. 2days
Summary and future work
• Primers allow us to amplify SSU of main soil
nematode types. Validated with traditional
techniques.
• Directed t-rflp designed and tested in silico and on
environmental samples
• Need to understand the differences in results
between morphological and molecular approaches
• Still some work to do but could be used to target
samples for morphological analysis
New projects include novel techniques
European Commission
funded project:
Soil ecological function
and biodiversity across
europe
www.ecofinders.eu
To link:biodiversity – function – ecosystem services
Not just nematodes:
Archaea, bacteria,
fungi, protozoa,
nematodes, micro-
arthropods, worms
+ functions: C cycling,
N cycling, water
retention.
A range of soils, climate and land use
The benefits of large-scale studies:An example from The Netherlands
Mulder et al. 2011 Advances in Ecological Research Vol. 44:
Nematode response to livestock intensity
Mulder et al., 2005 Naturwissenschaften
Thank youThank you…… Future directions:
More of the same!
Research on a broad front – applications of new
techniques; above-ground below-ground interactions;theory; interactions within the food web
National monitoring schemes increasinglyimportant (especially in Europe)
Development of trait based approaches
Trait differencesHeemsbergen et al 2004 Science
Significanttrend
-20
-10
0
10
20
30
3 6 7 9 11 12 13 16 18
Functional dissimilarity
Ne
td
ive
rsit
ye
ffe
ct
No trend
-10
-5
0
5
10
15
2 4 6 8
number of species
Ne
td
ive
rsit
ye
ffe
ct
Indices based on c-p and other traits havefacilitated ecological studies
Ferris, Bongers, de Goede (2001) A framework for soilfood web diagnostics: extension of the nematode faunalanalysis concept. Applied Soil Ecology 18: 13-29
Bongers, T., 1990. The maturity index: An ecologicalmeasure of environmental disturbance based onnematode species composition. Oecologia 83, 14-19.
Form and function: Metabolic footprints ofnematodes in the soil food web
Howard FerrisEuropean Journal of Soil Biology 46 (2010) 97-104
Form and function: Metabolic footprints