banana plantain

1. A member of CGIAR consortium www.iita.org
In vitro culture for production of quality banana and
plantain planting material
GUEYE Badara
RTB in vitro propagation and conservation specialist (In vitro Genebank)
International Institute of Tropical Agriculture (IITA)
Headquarters: PMB 5320, Oyo Road, Ibadan, Nigeria
Web: http://www.iita.org/genetic-resources-center
2nd
International Workshop of the Learning Alliance on Banana Bunchy Top Disease Control in Africa.
9 to 14 March 2015. IITA Ibadan, Oyo Road, Ibadan, Nigeria

2. A member of CGIAR consortium www.iita.org
PTC gives opportunity to grow plant material in artificial conditions with
advantages such as:
-Possibility for sanitation (culture in sterile conditions, pathogen cleaning)
-Rescue and conservation (continuous availability, space saving, cost-effectiveness, cryo)
-Micro-grafting and micro-propagation (mass multiplication: Bioreactor, cell suspension,
polyshoot…)
-Germplasm exchange
-Door opened for more research (genetic transformation, somatic embryogenesis, haploid
production, embryo rescue, molecular and pathological/virological investigations…)
INTRODUCTION

3. A member of CGIAR consortium www.iita.org
Musa spp. GERMPLASM SANITATION
Vegetatively propagated crop: accumulation of pathogens over the
reproductive cycles
3 major viruses: BBTV, BSV and CMV
Disease control: Production of clean planting material through virus
eradication
Cleaning techniques through in vitro culture: Meristem regeneration,
Thermo-treatment and Chemo-treatment
Factors:
(a) the virus characteristics,
(b) the type of tissues treated
(c) the plant species
(d) the genotype

4. A member of CGIAR consortium www.iita.org
In vitro VIRUS ELIMINATION TECHNIQUES
FOR Musa spp.
In vitro Virus
elimination
techniques
Meristem regeneration Thermo-treatment Chemo-treatment
(+)
- Most efficient on
phloem-associated
viruses ( BBTV)
- Association with other
techniques
- « the smaller, the
better »
- Mainly heat-treatment,
cryo-treatment less
popular
- Widely used in vivo/in vitro
and associated with
meristem culture
- A wide range of potentially
antiviral molecules tested
- More efficient on CMV
(-)
- Less efficient for virus in
meristematic dome (CMV)
- Depends on the virus type
- mode of action not very
known
- Time consuming and
harmful
- Phytotoxicity and negative
effect on regeneration
- Mode of action (inhibition,
elimination, « silencing ») ???

5. Schematic representation of virus eradication processes for banana. The green , blue,
and red arrows represent the process of chemotherapy, meristem culture, and
thermotherapy, respectively (Lassois et al., 2012. In Maurizio Lambardi et al. (eds.), In press)

6. A member of CGIAR consortium www.iita.org
MICROPROPAGATION
Musa spp. are very responsive to in vitro PTC
Meristem regrowth in 4 to 6 weeks, followed by shoot development
Proliferation: subculture of fully developed plantlet

7. A member of CGIAR consortium www.iita.org
MICROPROPAGATION
Musa spp. multiplication using semi-solid culture medium
Plantain/banana proliferation medium
Average multiplication rate onto semi-solid culture medium
4:1
Components Qty / L
MS basal medium 4.43 g
Myo-inositol 100 mg
Sugar 30 g
IAA (Indol-Acetic Acid) 0.18 mg
BAP (Benzyl Amino Purine) 4.5 mg
Ascorbic acid 10 mg
Agar 7 g

8. A member of CGIAR consortium www.iita.org
Large scale multiplication
MICROPROPAGATION
DeRoi, SA
National
genebank, SA
Vitropic,
France

9. A member of CGIAR consortium www.iita.org
MICROPROPAGATION
Musa spp. multiplication using Temporary Immersion System (TIS) Bioreactor
Rocket kit shaker & shouthern sun bioreactor
Air-lift Method

10. A member of CGIAR consortium www.iita.org
MICROPROPAGATION
Musa spp. multiplication using Temporary Immersion System (TIS) Bioreactor
NACGRAB, Nigeria
Setis Method

11. A member of CGIAR consortium www.iita.org
MICROPROPAGATION
Musa spp. multiplication using Temporary Immersion System (TIS) Bioreactor
Average multiplication rate using TIS
16:1

12. A member of CGIAR consortium www.iita.org
HARDERNING
Acclimatization process
More than 95 % success

13. A member of CGIAR consortium www.iita.org
HARDERNING
Nursery, a vital step (DeRoi, SA)

14. A member of CGIAR consortium www.iita.org
Virus cleaning by meristem culture
+ Most popular and more efficient (on phloem-associated viruses like BBTV)
+ « the smaller, the better »
+ Association with other techniques
e.g.: On Musa acuminata (AAA, cv Williams)
- BBTV eliminated at 99 % elimination
- BSV at 76 % and 41 % for in vivo and in vitro respectively
-Balance with regeneration capacity: « the bigger the better »
- Less efficient on CMV, virus in meristematic dome (1 à 7 % clean)

15. A member of CGIAR consortium www.iita.org
Virus cleaning by Thermo-treatment
+ Mainly heat-treatment, cryo-treatment less popular
+ Widely used in vivo/vitro and associated with meristem culture
- Depends a lot on the virus type
- mode of action not very known
-Time consuming and harmful

16. A member of CGIAR consortium www.iita.org
Virus cleaning by Chemo-treatment
+ A wide range of potentially antiviral molecules tested
+ More efficient on CMV
+ Example: Virazole ® (Ribavirin, 1-B-D-ribofuranosyl-1-2-4-triazole-3 carboxamide),
synthetic analogue of guanosine, replication inhibitor)
-Phytotoxicity and negative effect on regeneration
- Mode of action (inhibition, elimination, « silencing »)

17. A member of CGIAR consortium www.iita.org
Recommandation: combination of techniques
- Meristem culture + thermo-treatment:
e.g.: On Musa acuminata (AAA, cv Williams)
- CMV eliminated at 38-70 % elimination
- BSV at 79 % and 67 % for in vivo and in vitro respectively
Banana and plantain virus cleaning

18. A member of CGIAR consortium www.iita.org
Banana and plantain virus cleaning
CHALLENGES
- Clean or not expressed (« opportunism » of the virus)???
-What about B genomes???
- Integration of the virus in the genome (BSV)
- Inhibition of the expression (BSV)
- Policy issues (germplasm exchanges)…

20. Recommandation: combination of techniques
- Meristem culture + thermo-treatment:
e.g.: On Musa acuminata (AAA, cv Williams)
- CMV eliminated at 38-70 % elimination
- BSV at 79 % and 67 % for in vivo and in vitro respectively
Banana and plantain virus cleaning

21. Banana and plantain virus cleaning
CHALLENGES
- Clean or not expressed (« opportunism » of the virus)???
-What about B genomes???
- Integration of the virus in the genome (BSV)
- Inhibition of the expression (BSV)
- Policy issues (germplasm exchanges)…