Nematology in the provisionof soil ecosystem services:nutrients and energy in the

soil food web

Teagasc,Environment Research Centre,Johnstown Castle,Wexford.

bryan.griffiths@teagasc.ie

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