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ACP Project 1.3 International quarantine facility for the exchange of sugarcane germplasm among ACP...
Transcript of ACP Project 1.3 International quarantine facility for the exchange of sugarcane germplasm among ACP...
ACP Project 1.3
International quarantine facility for the exchange of sugarcane
germplasm among ACP countries
Mid-Term Review1 October 2012
MSIRIRéduit
Mauritius
International quarantine facility for the exchange of sugarcane germplasm among ACP countries
Implementing Institution: Mauritius Sugarcane Industry Research InstituteCountries targeted: ACP sugar producing countriesDuration of project: 4 yearsCost: Euros 918 510
INTRODUCTIONObjective:
To enhance the productivity and efficiency of the
sugarcane industry in ACP countries by promoting
germplasm exchange, making their industries more
responsive to the challenges in the global economy
Purpose of the International quarantine
To provide ACP countries with sugarcane plants free
from detectable pathogens in order to safeguard
their industry from potentially damaging diseases
Sugarcane diseases
Some 120 diseases recorded on sugarcane Most important ones are those known to be
transmitted by cuttings, referred as systemic diseases
Their most common mode of spread is through exchange of varieties between countries
28 systemic diseases are recorded The risks represented by them, if introduced,
can vary from minor to severe
How to minimize risk of accidental introduction of sugarcane diseases?
• Safe movement of sugarcane germplasm• Quarantine of germplasm• Effective testing (indexing) of material - use of
molecular tools• Movement of tissue culture plants
Sugarcane Quarantine in MauritiusMauritius has been regularly importing germplasm since 1914
A National closed quarantine is in existence since1946
The responsibility for the national quarantine has been vested on the MSIRI since 1953
Setting-up of an internationalquarantine station in Mauritius, which any member of the ACP countries can use to facilitate exchange of disease-free germplasm
International Quarantine Benefits:
Minimizing the risks of introducing potentially devastating diseases into ACP countries
Avoiding duplication of resources by establishing an international facility in one state
Using up-to-date disease detection technologies and tissue culture to speed availability of germplasm
Main Project Components
• Renovation of an existing quarantine to meet biosecurity level 3
• Setting up and equipping of a plant pathology Lab
• Renovation of a tissue culture facility and equipping
• Training of staff• Training of ACP members quarantine officials
project 1.3
Activity 1.1- Existing glass houses renovation
International quarantine facility designed to meet biosecurity level 3
Near completion
GH for imported germplasm & tissue culture plants
Activity 1.2 – Setting up of a new Plant Pathology Lab
1 2 3 4 5 6 7 8 9 10 11 12 13
352 bp
Near completion
Activity 1.3 – Upgrading of an existing Tissue culture laboratory
Completed
Tissue culture Laboratory designed & equipped
project 1.3
Activity 2.1
Training in molecular diagnostic of diseases
Major diseases not present in Mauritius: SCMV, SCSMV, Fiji disease, Downy mildewCIRAD provided training to staff in molecular detection of SCMV,SCSMV, and red leaf mottle virus
Activity 2.2 Diagnostic tools development
DEVELOPMENT OF DISEASE TESTING AND ELIMINATION PROCEDURE FOR POTENTIAL DISEASES
Based on: • Country of origin of germplasm• Requirements of importing country• Risk assessment analysis
Sugarcane Systemic DiseasesUNKNOWN ETIOLOGY
FUNGI VIRUSES BACTERIA
Ramu stunt Downy mildew Fiji leaf gall Gumming
Chlorotic streak Dry top rot Leaf fleck Leaf scald
Fusarium sett Mild mosaic Mottles stripe
Pineapple disease Mosaic Ratoon stunt
Red rot Red leaf mottle Red stripe
Smut Streak PHYTOPLASMA
Sclerophthora Streak mosaic Grassy shoot
Wilt Striate mosaic Green grassy shoot
Yellow leaf White leaf
Leaf yellows
Detection of Xanthomonas albineans
Primers and Probe designed from the
albicidin gene cluster sequence of
X albilineans
Primers/Probes checked for specificity
Conventional PCR & Real-time PCR tests
optimized Leaf scald disease
Xa specific fragment amplified by PCR
No amplification from Xcv
Highly specific as compared to ITS based primers
Xcv
isol
ates
Xcv
isol
ates
Wat
er c
ontr
ol
XaXa
Taqman ® Real-Time PCR for X albilineans detection
Inclusion of probe - added specificity guarding against non-specific annealing of PCR primer pairs
Amplification
curve observed
from all isolates
of Xa tested
Flat line for Xcv
isolates & water
control
Sugarcane Mosaic • Application of RT-PCR test for detection of
SCMV, SrMV, JgMV using Primer pair Oligo1n/2n
327 bp fragment amplified from poaceae potyviruses
Detection of Sugarcane yellow leaf virus
Multiplex RT-PCR optimized
Real-Time RT-PCR optimized
The virus displays high
genetic diversity
Development of real-time
RT-PCR tests for screening
genotypes in progress Leaf yellows disease
Multiplex RT-PCR for CUB, BRA-PER & REU genotypes of SCYLV
BRA-
PER CU
B REU
M
ultip
lex
Wat
er R
T-St
ep
Wat
er P
CR S
tep
Primers: REU-F/CPR, CUB-F/R & BRA-PER F/R
589 bp
450 bp
360 bp
Real-time RT-PCR specific for REU genotypes
REU genotypes amplified
No amplification from water controls, disease free plantlet, and plant infected with genotype BRA-PER
Activity 2.3
Tissue culture for elimination of diseases
Tissue culture/elimination of SCYLV- optimized
Month 0Month 1
Callus formation
3 subculturesof callus
Infected
Months 2-4
Months 5-6 RegenerationMultiplication & Rooting
Months 7-8
Diagnosis
Success rate of SCYLV elimination in vitro
Period Infected varieties
Successful regeneration
Virus elimination
% success for virus cleaning
1991-2001 18 16 16 1002001-2003 38 29 25 862003-2005 12 11 11 1002006-2008 13 10 10 1002008-2010 13 13 13 1002011-2012 8 6 6 100
Activity 2011 2012 2013 2014
1.1. Setting up of a glasshouse for imported germplasm
1.2 Setting up of a diagnostic lab
1.3 Upgrading of a tissue culture laboratory for disease elimination & multiplication of plantlets1.4 International advice on setting up the quarantine
1.5 Certification of the facility
2.1 Training of staff
2.2 Development of disease testing
3.0 Reception of germplasm in quarantine
4.0 Training of Plant health officials of ACP countries
International quarantine facility for the exchange of sugarcane germplasm among ACP countries
Expected Project Outputs• Quarantine facility available for ACP
countries to allow safe movement of germplasm
• Create awareness on diseases of quarantine importance
• Diagnostic protocols available to ACP countries
• Capacity building of plant health officials in ACP countries
Collaborators
• Dr Salem Saumtally• Mr Sonalall Dhayan• Mr Guy Triton• Dr Asha Dookun-Saumtally• Mr Nawshad Joomun• Mr Miguel Antoine
Acknowledgements