Identification of Cowpea (Vignaunguiculata

Feed the Future Innovation Lab for Collaborative

Research on Grain Legumes

Identification of Cowpea (Vigna unguiculata

L. (Walp.)) Lines and Polymorphic Gene-

based microsatellite Markers for Resistance

to Aphids (Aphis Craccivora Koch)Ouedraogo P. Adelaide, Tignegre Jean-Baptiste, Batieno Benoit Joseph, Poda S. Leandre,

Traore Fousseni, Ouedraogo Jeremy, Bao-Lam Huynh, Philip Roberts, and Close Timothy

Feed the Future Innovation Lab for Collaborative Research on Grain Legumes

INTRODUCTION

MATERIAL and METHODS

RESULTATS

CONCLUSION and RECOMMANDATIONS

Presentation outline



Figure 1:(left) cowpea plant; (right) Seed characteristics of farmer preferred varieties, Kamboinse 2008

Figure 2: Cowpea based-meals

Figure 3: (TOP) Vigna unguiculata subsp. Unguiculata cv.-gr. Textilis(BOT) California Blackeye 27Photo by: Mitchell Lucas

Hay for feeding livestock.

Important source of income.

Cowpea is grown as a green manure

and as a cover crop (United States).


Figure 4: Cowpea production by region( average 1994-2014 ), FAOSTAT 2014

Figure 5: Cowpea production ( top 10 producers), FAOSTAT 2014


Aphis craccivora is one of the insects responsible for yieldlosses in cowpea.

It causes damage by altering the metabolism, takingnutrients from the plant, and by transmitting plant viruses(Blackman and Eastop 2000)

Production constraints: biotic, abiotic and socio-economic(Tignegré, 2010).

Yields losses related to insect can be up to 100% in sub-Saharan Africa (Singh and Jackai, 1985).


Relatively tinyAdults can be winged or winglessTwo reproductive pathwaysGreat adaptability

Figure 6: wingless and winged aphids


Figure 7:Resurgence of A. craccivora in farmer field 3 weeks after insecticide application


MAIN OBJECTIVE:

The aim of this study was to contribute to an increase in the yield of cowpea by the identification of aphid resistant lines and polymorphic gene-based markers.

SPECIFIC OBJECTIVES:

To screen cowpea genotypes for resistance to aphids.

To characterize populations of aphids that prevail in cowpea

growing areas.

To identify polymorphic markers for selection purposes.



MATERIALS

Kvx396-4-5-2D

Tiligré and Kvx295-2-124-99 for the polymorphism test.Figure 8&9:pictures of the 10 varieties of cowpea used for the aphid resistance test


METHODSSource and maintenance of experimental insects

Figure 10: Aphids sampling areas located in the three main agro-climatic zones in Burkina.

Figure 11: Cages used for aphides maintenance

Experimental design: Randomized complete block design with three (3) blocks, five (5) repetitions each.

Plants in each block were infested with only one of the three (3) aphid strains.


Observations were made every three days and consisted of

measuring some parameters:

• The number of leaves;

• Plant survival;

• The dynamics of the aphid populations.

Figure 12: Infestation technique


No aphid 1 to 5 aphids 5 to 20 aphids 20 to 100 aphids 100 to 500 aphids More than 500 aphids

Figure 13: Field layout


To look for polymorphic markers, DNA of each parent was extracted,

and amplified with each ninety-six (96) pairs of SSR primers.

The raw data of 20 DAS, 35DAS, 50 DAS, 65 DAS, one month after

infestation and data from the AUDPC were used for several analyzes.

XLSTAT 7.1 and SAS 9.4 software were used to analyze the data. The

formula used to calculate the AUDPC is:

nAUDPC = Σ [(Xi+1 + Xi) / 2] [ti+1 - ti]

i=1


RESULTS


Varieties 20 DAS 35 DAS 50 DAS 65 DAS AUDPCB301 3 4.6 5 4.8 202.5CB 27 2.8 1 2.4 2 87

IT97K-556-6 1.4 2.2 3 3 111KN-1 3.2 2.2 4 3.8 145.5KVX295-2-124-99 3.8 4 4.6 4.6 192N°2300 4.2 5 3.8 3.8 192NS-1 4.2 5 4 4 196.5

NS-Farako Bâ 3.2 5 4.8 4.6 205.5

SARC-1-57-2 3.2 3 4 5 166.5

SARC-1-91-1 2.8 2.8 2.8 2.8 126

Varieties 20 DAS 35 DAS 50 DAS 65 DAS AUDPCB301 1.6 3 2.8 2.8 120CB 27 1.6 2.8 2 2 99IT97K-556-6 0.2 0 1 2 31.5KN-1 1 1 1 0.6 42KVX295-2-124-99 1.6 3 3.8 3.8 142.5N°2300 3.8 3.6 4.4 4.4 181.5NS-1 3 4.4 4.8 4.4 193.5

NS-Farako Bâ 2.8 3.8 3.8 3.8 163.5SARC-1-57-2 1.6 1.6 2.4 2.4 90SARC-1-91-1 1.6 2.6 3 3 118.5

Table 1& 2: Means data of degrees of infestation and the AUDPC (Bobo and Kamboinsé)

IDENTIFICATION OF RESISTANCE SOURCES

THE EVOLUTION OF THE INFESTATION


Table 4,5 &6 : correlation test between degrees of infestation at 20 DAS ; 35 DAS ;50 DAS ; 65 DAS and the AUDPC (respectively aphids strain of Kamboinsé, Bobo and Pobe))Varia

bles 20 DAS 35 DAS 50 DAS 65 DAS AUDPC

20 DAS 1

35 DAS 0.712 ⃰ ⃰⃰⃰ 1

50 DAS 0.611 ⃰⃰⃰⃰ ⃰⃰⃰ 0.776 ⃰⃰⃰⃰ ⃰⃰⃰ 1

65 DAS 0.563 ⃰⃰⃰⃰ ⃰⃰⃰ 0.733 ⃰⃰⃰⃰ ⃰⃰⃰ 0.938 ⃰⃰⃰⃰ ⃰⃰⃰ 1

AUDPC 0.745 ⃰⃰ ⃰⃰⃰ 0.923 ⃰⃰⃰⃰ ⃰⃰⃰ 0.950 ⃰⃰⃰⃰ ⃰⃰⃰ 0.915 ⃰⃰⃰⃰ ⃰⃰⃰ 1

Variables 20 DAS 35 DAS 50 DAS 65 DAS AUDPC

20 DAS 1

35 DAS 0.446 ⃰⃰⃰⃰ ⃰⃰⃰ 1

50 DAS 0.545 ⃰⃰⃰⃰ ⃰⃰⃰ 0.613 ⃰⃰⃰⃰ ⃰⃰⃰ 1

65 DAS 0.582 ⃰⃰⃰⃰ ⃰⃰⃰ 0.557 ⃰⃰⃰⃰ ⃰⃰⃰ 0.918 ⃰⃰⃰⃰ ⃰⃰⃰ 1

AUDPC 0.666 ⃰⃰⃰⃰ ⃰⃰⃰ 0.850 ⃰⃰⃰⃰ ⃰⃰⃰ 0.919 ⃰⃰⃰⃰ ⃰⃰⃰ 0.883 ⃰⃰⃰⃰ ⃰⃰⃰ 1

Variables 20 DAS 35 DAS 50 DAS 65 DAS AUDPC

20 DAS 1

35 DAS 0.548 ⃰⃰⃰⃰ ⃰⃰⃰ 1

50 DAS 0.452 ⃰⃰⃰⃰ ⃰⃰⃰ 0.778 ⃰⃰⃰⃰ ⃰⃰⃰ 1

65 DAS 0.414 ⃰⃰⃰⃰ ⃰⃰⃰ 0.719 ⃰⃰⃰⃰ ⃰⃰⃰ 0.797 ⃰⃰⃰⃰ ⃰⃰⃰ 1

AUDPC 0.620 ⃰⃰⃰⃰ ⃰⃰⃰ 0.927 ⃰⃰⃰⃰ ⃰⃰⃰ 0.933 ⃰⃰⃰⃰ ⃰⃰⃰ 0.862 ⃰⃰⃰⃰ ⃰⃰⃰ 1

Varieties 20 JAS 35 DAS 50 DAS 65 DAS AUDPCB301 1.6 2.4 3.6 3.6 129CB 27 3.4 4.2 4 4.4 181.5IT97K-556-6 2.4 1.4 0.6 0 48KN-1 0.8 1 1 2 51KVX295-2-124-99 2.6 2.6 3.6 2.6 132N°2300 2.6 2.6 3.6 3.6 139.5NS-1 3.2 4 3.8 3.8 169.5NS-Farako Bâ 3.2 4 4 4 174SARC-1-57-2 1.2 1 1 2 54SARC-1-91-1 1.6 4 3 4 147

Table3: Means data of degrees of infestation and the AUDPC (Kamboinsé)


Fixedterm

Wald statistic

n.d.f. F statistic

d.d.f. F pr

A.Strains 10.22 2 5.11 120 0.007

Varieties 43.83 9 4.87 120 0.001

Strain*

Varieties18.27 18 1.01 120 0.448

N.Lea. D.INF. Surv.R.

N. Lea. 1

D.INF. -0.652 1

SURV.R. 0.728 -0.628 1

Table 7:Combined analysis of the AUDPC for the 3 strains of aphids.

Table 8: Correlations between the 3 variables (number of leaves, survival rate and degree of infestation)

SIMILARITY AND CORRELATION TEST


VARIETIES CLASSIFICATIONIT

97K

-556

-6

KN

-1

B30

1

KV

X29

5-2-

124-

99

SAR

C1-

91-1

CB

27

SAR

C-1

-57-

2

NS-

1

N°2

300

NS-

Fara

ko B

â

0

2

4

6

8

10

12

14

16

18

Dis

sim

ilari

ty

KV

X29

5-2-

124-

99

N°2

300

SAR

C1-

91-1

CB

27

IT97

K-5

56-6

NS-

1

SAR

C-1

-57-

2

NS-

Fara

ko B

â

B30

1

KN

-1

0

2

4

6

8

10

12

14

16

Dis

sim

ilari

tyFigure 14&15 :Dendrogram representing the three classes of varieties (left:strain of kamboinse; right:strain of Bobo)


KN

-1K

VX

295-

2-12

4-99

IT97

K-5

56-6

SAR

C-1

-57-

2

B30

1

SAR

C1-

91-1

N°2

300

NS-

1

CB

27

NS-

Fara

ko B

â

0

5

10

15

20

25D

issi

mila

rity

Resistant ModeratelyResistant

Susceptible

Figure 16 :Dendrogram representing the three classes of varieties (Strain of Pobé)

RESISTANTS MODERELY RESISTANTS SUSCEPTIBLESIT97K-556-6 B301 N°2300

VARIETIES SARC1-91-1 CB 27 NS-1KVX295-2-124- 99 SARC-1-57-2 NS-Farako BâKN-1

Tableau 9: Characteristics of the 10 varieties studied in the three aphid strains.


Bobo-Dioulasso Kamboinsé Pobé Mengao

Bobo-Dioulasso 0.00Kamboinsé 3.46 0.00

Pobé Mengao 4.20 0.25 0.00

CHARACTERIZATION OF APHID STRAINS

Tableau 9:Fisher's F associated with the square of the Mahalanobis


POLYMORPHISM TEST

24 polymorphic markers out 96.

Figure 18:Electrophoretic profile


CONCLUSION & RECOMMANDATIONS


Among the 10 varieties:

Four(4) resistant varieties, three (3) moderely resistant.

The Pobé strain is similar to that of Kamboinsé and these

two strains are different from the Bobo one.

Twenty four (24) polymorphic markers found .


Validation of polymorphic markers specially codominant

ones for selection purposes.

Molecular characterization of Bobo, Pobé and Kamboinsé

aphid strains in order to give more precise information

about the genetics of the different strains of aphids.


Date post:	02-Nov-2021
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Identification of Cowpea (Vignaunguiculata

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