Day, 20 June 2010 Agenda Point (13:30-14:30) Vigour ... … · Malavika Dadlani IN Ibrahim Demir TK...

Vigour Committee report 2009-2010

Day, 20 June 2010 Agenda Point (13:30-14:30)

Vigour Committee Members

Chair: Alison A Powell GB

Vice-Chair: Michael Kruse DE

Members: Patricia Brownfield US

Emanuela Casarini IT

Malavika Dadlani IN

Ibrahim Demir TK

Marie-Hélène Wagner FR

Hulya Ilbi TK

Mohammad Khajeh Hosseini IR

Seok Hyeon Kim K

Tim Loeffler NE

Gillian McLaren GB

Stan Matthews GB

Rasha El-Khadem (resigned 09/2009) AT

Yan Rong Wang CN


Structure:

Summary of work in 2009-2010; questions

Nature of work in VC; questions

Proposals for 2010-2013; questions

Vigour Committee report 2009-2010


Summary of work completed

2009-2010

Introduction of new tests into the Rules

Controlled Deterioration test for Brassica spp. In Rules

2010

Introduction of new species

Phaseolus vulgaris – Conductivity test. In Rules 2010

Glycine max – Conductivity test – Validated 2009; Rules Proposal 2010


Workshop

No ISTA workshops

British Association of Seed Analysts – SASA, Edinburgh

Publications

14 publications:

•3 method validation reports

•2 general articles on seed testing topics

•9 research publications

Provide background to test development


Highlights of research publications

Rate of germination test:

Maize:

Mean germination time in the laboratory estimates the relative

vigour and field performance of commercial seed lots of maize

(Zea mays L.).

Khajeh-Hosseini, M., Lomholt, A. and Matthews, S. (2009), Seed Sci. &

Technol., 37, 446-456.

Rate of physiological germination compared with the cold test and

accelerated ageing as a repeatable vigour test for maize.

Matthews, S., El-Khadem, R., Casarini, E., Urso, G., Khajeh Hosseini, M,

Nasehzadeh, M. and Wagner, M-H. Seed Sci. & Technol., In press

Cucurbits

Mean germination time estimates the relative emergence of seed lots of

three cucurbit crops under stressful conditions.

Mavi, K., Demir, I. and Matthews, S. (2009) Seed Sci. & Technol, 37,.


Cool germination test

Maize

Cool germination test as an alternative vigour test for maize.

Ilbi, H., Kavak, S. and Eser, B (2009) Seed Sci. and Technol, 37, 516 -

519.

Conductivity test

Cabbage

Vigour tests for cabbage seeds using electrical conductivity

and controlled deterioration to estimate relative emergence in

transplant modules.

Matthews, S., Demir, I., Celikkol, T., Kenanoglu, B.B. and Mavi, K.

(2009), Seed Sci. & Technol., 37, 736-746



Special projects

Controlled deterioration test:

approaches to reducing test durationM-H Wagner, I Demir, A Powell

Rate of germination test

Maize, cotton, cucurbits, pepper, oilseed rapeS Matthews, M-H Wagner, E Casarini, R El-Khadem, M

Khajeh Hosseini, I Demir, G McLaren

Conductivity test:

extension to Brassica spp.M-H Wagner, A Powell, S Matthews, I Demir

Machine vision:

application to assess rate of germinationM-H Wagner

Any questions?



Nature of work completed by the

Vigour Committee

What is our overall aim?

1. To develop new vigour tests

2. To add new species to validated vigour tests


Nature of work completed by

the Vigour Committee

1. To develop new vigour tests

• Background research

Is there a problem?

What is physiological basis for the problem?

Could this be used in a test?

Sometimes different tests for different species

Commercial seed lots

• Publication

• Comparative tests

• Validation

2. To add new species to validated vigour tests

•Background research

Is there a problem of vigour in the species?

Does the biological basis of the test suggest

it will apply to the new species?

Experiments on commercial seed lots

differing in vigour

•Publication?

•Comparative tests

•Validation


Examples of test development

Controlled deterioration test

Background research

Comparisons of field emergence:

2 years of trials; 2 field sites; 2-6 sowings per species per year.

10 vegetable species (6 Brassica spp.); 10-30 commercial seed lots

Comparisons of storage potential

2.5 year commercial storage – 2 ambient stores, 1 dehumidified store

4 species (1 Brassica sp). Regular samples.

Physiological basis of test = seed ageing

Empirical selection of MC and temperature to achieve separation of

lots in 24 hours.

Test against emergence and storage in each year.

Correlate test results with emergence and storage


Publications

At least 15 ; 7 on Brassica sp

Comparative tests

1995-1998: 5 seed lots x 3 test runs (4 labs)



Validation

Data from all test runs in method validation report

Accepted June 2009; Rules January 2010.


Controlled deterioration test: continued




Background research

Maize

Rate of germination (radicle emergence) shown to relate to vigour

1. Mean germination time (MGT) indicative of soil emergence in lab:

11 commercial lots (SST, 2006)

MGT

Final emergence -0.633*

Mean emergence time (MET) 0.749**

Mean seedling length -0.780**

CV seedling length 0.700*

2. Length of MGT (lag period) relates to uniformity of emergence

and seedling size: 10 commercial lots (SST 2007)

0

20

40

60

80

100

0 24 48 72 96 120 144

Germ

inatio

n (%

)

Time f rom set to germinate (h)

F

H

I

MGT=Σ(nt)/ Σn

Germination

at 20oC

Lag period

Longer

lag period

MGT relates to seedling size and uniformity in lab

MGT = 4.80 days

Faster germination

MGT = 5.84 days

Slower germination

3. MGT relates to single early count of radicle emergence at both

13oC and 20oC (SST, 2009):

7 commercial lots, 3 laboratories

y = -37.653x + 294.64

R2 = 0.95**

20

40

60

80

100

5 6 7 8

MGT at 13 o C (d)

6d

ge

rmin

atio

n a

t 1

3 o C

(%

)

(a) y = -43.829x + 200.69

R2 = 0.98***

20

40

60

80

100

2 2.5 3 3.5

MGT at 20 o C (d)

3d

ge

rmin

atio

n a

t 2

0 o C

(%

)

(b)

Low MGT = high early count High MGT = low early count

MGT also related to seedling size and uniformity in lab (SST, 2009)


Research has led to conclusion that:

•Rate of germination based on radicle emergence

relates to vigour in maize

•Rate based on : MGT (based on many counts)

and

single count of radicle emergence

Comparative tests

4 laboratories x 6 commercial lots x 2 test runs (SST 2010)

LotLab A Lab B Lab C Lab D Overall

mean

Cold test

G (%)Run 1 Run 2 Run 1 Run 2 Run 1 Run 2 Run 1 Run 2

498a

2

100a

1=

95a

1

100a

1

100a

1

100a

1

95a

1

100a

1

99a

1

99

1

5100a

1

100a

1=

93a

2

95ab

2

95ab

2

100a

2

85ab

2

95a

2

95a

2

98

2

288b

3

75b

3

80ab

3

88ab

3

83ab

3

80b

3

80bc

3

75b

3

81b

3

76

3

368bc

4

65bc

4

68bc

4

83b

4

95bc

4

75b

4

63cd

4

45c

4

70b

4

67

4

145cd

5

55bc

5

43cd

5

35c

5

70c

5=

73b

5

48d

5

15d

5=

48c

5

54

5

628d

6

40c

6

25d

6

50c

6

70c

5=

73b

6

25e

6

15d

5=

41c

6

50

6

r with

cold

test G

0.95 0.99 0.95 0.89 0.86 0.96 0.93 0.98 0.98

Validation

2nd comparative test: 5 laboratories x 6 commercial lots x 2 test runs

Data used for preparation of Method Validation Report (in progress).



Further development of assessment of rate of

germination : Machine vision

• Jacobsen tables

• T C 10 to 30 C

• Continuous watering

• 4 cameras by monitor

• Handmade sowing by zone

• 1, 4 or 16 zones per camera

• Image shot automation

• Image analysis

• Data treatment


Image n 60 (120 h at 15 C)

Image n 60 after

segmentation

2 samples A and E,

4 x 50 seeds

09_01_12_Mais_1_0.avi


Lot A Lot E

Lot ALot E


0

20

40

60

80

100

0 24 48 72 96 120 144 168

Ge

rmin

ati

on

(%

)

Time after sowing (hours)

Lot A 20°C

Lot E à 20°C

Lot A à 15°C

Lot E à 15°C

•Mean germination time based on automated counts


21-days emergence (%)

after an early sowing

Lot A Lot E

56 34

daily temperature

5

10

15

20

07-

avr

09-

avr

11-

avr

13-

avr

15-

avr

17-

avr

19-

avr

21-

avr

23-

avr

25-

avr

27-

avr

me

an

te

mp

era

ture

(°C

)

Lab test

(5 samples)

Emergence

AA 0.543

CM 0.457

CT 0.619

MGT 20 C

(France)

0.664 *

MJGT 13 C

(Scotland)

0.690 *

•Mean germination time related to emergence (2006 Workshop)

Introduction of a species to a validated test

Conductivity test: Phaseolus vulgaris, Glycine max

Background research

Problems of vigour documented

Evidence exists that conductivity relates to emergence

Comparative tests

Phaseolus vulgaris: 6 lots x 5 labs x 3 test runs

Glycine max: 5 lots x 7 labs

Validation

Phaseolus: Validated 2008; ISTA Rules 2009

Glycine max: Validated 2009; ISTA Rules Proposal 2010


Current work on test development: 1

Controlled Deterioration (CD) test

– Extension to new species –pepper, aubergine,

cucurbits

– Shortening the test



CD predicts vigour of watermelon

Year Correlations with emergence under

stress:

Correlation

with

germination

after storageLow

temperature

Mechanical Salinity

2005 LG 0.81* 0.83** 0.73* 0.83**

CD 0.91*** 0.92*** 0.85** 0.95***

2006 LG 0.69* 0.61* 0.72* -

CD 0.93*** 0.89*** 0.95*** -

Mavi and Demir, 2007


Aubergine:

CD germination and emergence in modules

y = 0.77x + 31.704

R2 = 0.9322

50

60

70

80

90

100

40 50 60 70 80 90 100Germination (%)

Fin

al em

erg

en

ce (

%)

y = 1.6667x - 74.917

R2 = 0.1778

Demir et al

50

75

100

Germ

ination a

fter

4 m

onth

s

50

75

100

20 40 60 80 100

Germ

ination a

fter

8 m

onth

s

Germination percentage (%)

Pepper: CD and storage

CD at 22% MC, 45oC

Basak, Demir et al, SST, 2006

CD:72h

CD: 24 h LG

LG

CD: 24 h

CD:72h

4 Months storage: 5oC, 8% m.c

8 Months storage: 5oC, 8% m.c


• Shortening the CD test

– Shorten equilibration time

– Replace germination after deterioration

• by EC measurement

• by early count of germination



Conductivity test for Brassica

– Initial EC predicts vigour?

– EC after CD



EC correlated to SG during storage of oilseed rape

18 commercial lots stored one year in two extreme conditions,

9 just harvested and 9 already stored from one to 4 years in seed companies

0

50

100

150

200

250

300

350

0

20

40

60

80

100

0 2 4 6 8 10 12

Co

nd

ucti

vit

y(µ

S.c

m-1

.g-1

)

No

rmal g

erm

inati

on

(%

)

Storage (months)

SG during storage at 10°C SG during storage at 30°C

EC during storage at 10°C EC during storage at 30°C

0

20

40

60

80

100

0 2 4 6 8 10 12

No

rmal g

erm

inati

on

(%

)

Storage at 30 C and 75% RH (months)

H1

H2

H3

L1

L2

L3

3847 48 51

6065

0.0

20.0

40.0

60.0

80.0

100.0

H1 H2 H3 L1 L2 L3

Initial conductivity and germination of 6 samples (3 hybrids and 3 lines)

EC SG

18 seed lots mean behaviour during storage


EC correlated to SG after CD test of oilseed rape

15 seed lots before ( )and after CD test ( ) at 16% or 20% MC

MC adjusted by imbibition or adding water method

y = -0.4813x + 119.7R² = 0.6908**

0

20

40

60

80

100

40 90 140 190 240

No

rma

l g

erm

ina

tio

n (

%)

Conductivity (µS.cm-1.g-1DW)


Rate of germination

– Cucurbits, cotton, pepper, oilseed rape


R2 = 0,80***

0

20

40

60

80

100

LT

E (

%)

R2 = 0,80***

0

20

40

60

80

100

HT

E (

%)

R2 = 0,83***0

20

40

60

80

100

MS

E (

%)

R2 = 0,81***

0

20

40

60

80

100

40 60 80 100

MGT (Hours)

SS

E (

%)

Watermelon

R2 = 0,81***

R2 = 0,76**

R2 = 0,71**

R2 = 0,53*

30 40 50

MGT (Hours)

Melon

R2 = 0,62*

R2 = 0,64**

R2 = 0,69**

R2 = 0,60*

20 30 40 50 60

MGT (Hours)

Cucumber

Rate of germination

(MGT; hours)

Low temperature

High temperature

Mechanical stress

Salinity stress

Emergence in conditions of:

Mavi, Demir and Matthews, SST, 2010

Fast Slow

y = -20.255x + 133.84

R2 = 0.83 ***

0

20

40

60

80

100

2 3 4 5

MGT (18 o C) d

% F

ield

em

erg

en

ce

y = 0.3442x + 44.743

R2 = 0.76 ***

0

20

40

60

80

100

0 20 40 60 80 100

% Germination 3d (18 o C)

% F

ield

em

erg

en

ce

S Matthews, B Ross, P Steel

Courtesy of Cotton Seed Distributors

Wee Waa, NSW

Australia

Cotton (Australia 2007)

Fast FastSlow Slow

Correlation of Physiological germination (2mm radicle ) after 3d at 10oC with:

Field emergence at 7 days 0.913***

Field emergence at 10 days 0.781**

Oil seed rape: Early counts at 3 days correlate with field emergence

McLaren, Cockerell and Matthews

y = 0.54x + 34.39

R² = 0.61

p < 0.01

50

60

70

80

90

100

40 60 80 100

Germination after 3 days (%)

Fin

al e

me

rge

nc

e (

%)

Any questions?


Proposals for 2010-2013

Test development

Research

Basis of vigour differences

Biotinylated proteins as quality markers in oilseed (rape,

sunflower)

Cell cycle and DNA degradation (RG test, maize)


Automation of germination time course• Identification of best time for single count

at different temperatures

Application to cotton, onion, oilseed rape, Viola spp



Research continued

Controlled Deterioration test

Modifications to reduce test time: •Adjustment of seed moisture content

•EC after deterioration – Brassica spp

•Single early germination count:

Leek, onion, pepper, tomato, watermelon

Conductivity test

Short test for chickpea

Extension to new species- leek, onion, radish, cauliflower

Flower seed

Use of Saturated Salt AA / CD

Grasses

Elymus nutans, E. sibiricus – alpine species, China

Vigour test development


Test development

Comparative tests

Accelerated Ageing

Cereals

Rate of germination / Controlled deterioration

Peppers, cucurbits

Conductivity

Brassica spp

Validation

RG test

Maize

Proficiency test

2nd PT round 2013 Conductivity test (Pisum sativum)

Workshops

Japan: with Moisture and Tetrazolium Committees

Turkey

Canada



Any questions?


Thank you!

Day, 20 June 2010 Agenda Point (13:30-14:30) Vigour ... … · Malavika Dadlani IN Ibrahim Demir TK...

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