Effect of Climate Change on Emerging Natural
Toxins in Plants and Seafood Production
Επιδπάζειρ ηων Κλιμαηικών Αλλαγών ζηη
Δημιοςπγία Αναδςόμενων Φςζικών Τοξινών και
ζηην Παπαγωγή Φςηών και Θαλαζζινών
SAFEFOODERA-EMTOX project 7 FP
Dr. E. Ioannou– Kakouri,
Chief Chemist, EFSA focal point
Ministry of Health, State General Laboratory
44 Κimonos str, 1451 Nicosia, Cyprus
Επγαζηήπι για ηην Πποζαπμογή ζηην Αλλαγή ηος Κλίμαηορ
2 - 3 Νοεμβπίος 2011
Holiday Inn, Λεςκωζία
State General Laboratory
Ministry of Health
44 Κimonos str, 1451 Nicosia, Cyprus
Contents
Effect of Climate Change on Emerging Natural
Toxins in Plants and Seafood Production
SAFEFOODERA-EMTOX project 7 FP 2009-2011
www.emtox.gov.cy
1. Introduction
2. Role of SGL-WP3
3. Methodology
4. Results
5. Discussion
6. Finalization of project- Conclusions
Introduction
•Climate change has become one of the most critical
issues for the future of our planet. It involves significant
changes in the variability or average state of the
atmosphere, such as related to temperature, precipitation
and/or wind patterns, over durations ranging from
decades to millions of years.
•The effects of climate change will have implications
for food production, food security and food safety.
•In particular, the safety of feed and food products arising
from plant and marine production systems is expected to
be affected by climate change, specifically by increased
occurrence of natural toxins, including mycotoxins and
phycotoxins (marine biotoxins).
•The overall objective of the current research is to project
the impact of climate change on the occurrence of
(possible) feed and food safety hazards in both terrestrial
plant production systems and marine seafood systems in
Europe.
•The project particularly focuses on natural toxins in
cereal and shellfish production that are harmful to animal
and human health, including several key mycotoxins and
phycotoxins. Some of these are re-emerging.
•As a second objective, the project aims to quantify (as
much as possible) the relationship between climate change,
shifts in primary plant and marine production systems, and
the occurrence of (re-)emerging natural toxins. Hereby,
currently available predictive models will be used as well as
reliable databases.
SAFEFOODERA- EMTOX project 7 FP, 2009-2011
The project was coordinated by the RIKILT- Institute of Food
Research, Netherlands, other participating countries: Cyprus
(SGL) and Scandinavian countries, Sweden, Norway, Finland,
Denmark. The SGL was involved in WP3.
The project has 7 WP :
• Work Package 0 – Kick off meeting - coordination
• Work Package 1 – Climate change scenarios, DMI, DK
e.g. ENSEBLES database (Global and regional climate data)
• Work Package 2 – Cereal production Aarhus Univ, DK
• Work Package 3 – Mycotoxins in cereals, RIKLT, NL
• Work Package 4 – Phytoplankton production and blooms-No
• Work Package 5 – Harmful algae and marine toxins- Del-NL
• Work Package 6 – Integration of models RIKLT, NL
Role of State General Laboratory
Role of SGL
Participation in SAFEFOODERA- EMTOX project 7 FP, WP 0 & 3
1.
Organization of Kick-off meeting 28-29/4/09 and EMTOX webpage
(www.emtox.gov.cy) (WP 0)
2.
Mycotoxins in cereals (WP 3). Analysis of local cereal samples especially
wheat and barley and their products for re/emerging mycotoxins Τ-2, ΗΤ2, DON, ZON (fusarium toxins) and OTA &AFs.
3.
Availability, enrichment of the national database for the results of Τ-2,
ΗΤ-2, DON, ZON, OTA and AFs. analysis in cereal samples.
4. Collection of agronomic data. Completion of questionnaire by farmers.
Partners for providing samples etc:
1.
2.
3.
4.
5.
6.
7.
S.E.DI.S LTD ( Coop. Assoc. of Cereal Farmers, samples+ quest/res )
Cyprus Grain Commission (samples )
Department of Agriculture ( data for mycotoxins, samples+ quest/re )
Agriculture Research Institute (Samples+ Quest/re )
Health Services (Samples)
Meteorological Service (meteorological data)
Farmers (Samples+ Quest/res )
SGL and WP 3- Mycotoxins in cereals
•Mycotoxins are secondary metabolites which are
produced by a wide variety of fungal species in
a range of crops, cereals, nuts and fruits,
particularly during cultivation and/or storage
stages of production.
•Mycotoxins are chemically stable and – to a large extent
- resistant against most of the feed and food processing
steps , resulting in contamination of the final feed and
food products.
• Consumption of contaminated cereal based feed and
food product can cause harmful effects to human and
animal health.
SGL- Mycotoxins control
 The
presence of various toxic/carcinogenic Mycotoxins:
Aflatoxins B1, B2, G1, G2, Ochratoxin A (OTA) and other
mycotoxins in local and imported foodstuffs (cereals, nuts, dried
fruits, rice etc) has been monitored and controlled systematically
and effectively since 1990 in Cyprus, by the State General Lab.
(SGL) and the Health Services of the Ministry of Health.
 The samples for official control are collected according to a
specific sampling plan based on the EU Regulations, at critical
control points ( HACCP approach) such as:
import, primary storage, processing and market.
1990
1991
1992
1993
1994
1995
1996
1997
1998
1999
2000
2001
2002
2003
2004
2005
2006
2007
2008
2009
2010
% SAMPLES ABOVE LIMIT
MONITORING PROGRAMME OF
AFLATOXIN (HACCP)
IN CYPRUS 1990-2010
MARKET
12
ALL CCPs
10
8
6
4
2
0
Figure 1
INCIDENCE OF AFLATOXIN M1 IN MILK IN CYPRUS
1993-2010
% POSITIVE SAMPLES
70
60
50
40
30
20
10
0
1993 1995 1996 1997 1998
1999 2002 2003 2004 2005
2006 2007 2008 2009 2010
Fig.4 Afl M1 levels< aver 2-8 ppt
INCIDENCE OF OCHRATOXIN A IN FOODSTUFFS IN CYPRUS
1997-2010
100
% POSITIVE SAMPLES
90
80
RAISINS
70
COFFEE
WINE
60
CEREALS
50
40
30
OTA levels
within ML
20
10
0
1997 1998
1999
2000
Fig.
4 2006
2005
2007 2008
2009 2010
DON levels
within ML
Methods of Analysis of Mycotoxins
Extraction
With Methanol:Water or Acetonitrile:Water
+
NaCl or NaHCO3
 Filtration
(fluted paper)
 Dilution with water or PBS
 Filtration (GF/A)
Immunoaffinity column clean-up
Methodology of Analysis of Mycotoxins
In immunochemical methods specific and selective immunoaffinity
columns are used that contain specific antibodies for each mycotoxin.
Subsequently, the process of purification based on the antibody-antigen
effect takes place.
Methods of Analysis of Mycotoxins
• Aflatoxins B1, B2, G1, G2
• Ochratoxin A
• Zearalenone (ZON)
HPLC/FLD
• Deoxynivalenol (DON)
HPLC/PDA
• Trichothecenes
(T-2, HT-2 toxins)
GC/MS
Multi-Mycotoxin Analysis
LC-MS/MS
Results – Occurrence of Mycotoxins in cereals &
Products
Table 1. Levels
of mycotoxins in cereals and their products for the years 2008-2010
Sample
Parameter
Wheat
DON
OTA
Aflatoxins
Τ-2
ΗΤ-2
ΖΟΝ
6
7
25
1
1
3
5
7
7
1
1
3
1
2
0
0
0
0
Barley
DON
OTA
Aflatoxins
Τ-2
ΗΤ-2
ΖΟΝ
2
13
20
1
1
7
2
11
20
0
0
7
0
1
0
0
0
0
DON
OTA
Aflatoxins
Τ-2
ΗΤ-2
ΖΟΝ
1
0
19
1
1
4
0
0
10
1
1
2
1
0
3
0
0
2
DON
OTA
Aflatoxins
Τ-2
ΗΤ-2
ΖΟΝ
11
4
7
2
2
1
8 (?)
3
1
1
1
0
DON
OTA
Aflatoxins
Τ-2
ΗΤ-2
18
5
0
0
0
6 (?)
3
0
0
0
Corn
Flour
Pasta
Number Number of Number of Minimum Maximum
of
local
positive value (ppb)
value
samples samples
samples
(ppb)
Mean
value
(ppb)
Median value
(ppb)
Number of samples
over the maximum
limit
414
0,14
414
0,33
414
0,24
0
0
0
0
0
0
0,26
0,26
0,26
186
186
186
0,36
21,67
11,02
4,7
5,4
5,1
10
2
0
0
0
1
80
0,28
709
1,57
283
0,93
0
0
0
14
14
14
0
12
3
0
0
0
64,7
0,06
692
0,54
234,6
0,25
0
0
0
0
0
0
0
0
0
0
0
0
0
1
0
0
0
Results – Occurrence of Mycotoxins in Cereals
Results in cereals and their products (2008-2010)
Number of Samples
80
60
No of Positive
Samples
No of Samples
40
20
0
DON
ZON
OTA
AFLs
T-2
HT-2
Results for DON (2008-2010)
Results for ZON (2008-2010)
7
15
No of
Samples
10
No of Positive
Samples
5
0
BARLEY WHEAT CORN FLOUR PASTA
Number of Samples
Number of Samples
20
6
5
No of
Samples
4
3
No of Positive
Samples
2
1
0
BARLEY
WHEAT
CORN
FLOUR
Results
Table 2. Results
number of
positive
samples
1
5
2
1
0
1
8
1
Minimum Maximum Mean value
value (ppb) value (ppb)
(ppb)
414
0,27
1,4
2,0
414
0,67
1,4
3,1
414
0,47
1,4
2,7
0,26
2,8
2,0
0,26
14
2,0
0,26
9,4
2,0
No of samples
No of local samples
No of positive samples
0
BARLEY
WHEAT
Number of Samples
10
5
Results for ΟΤΑ (1997-2010)
No of samples
No of local samples
No of positive samples
5
0
BARLEY
WHEAT
15
10
No of samples
No of local samples
No of positive samples
5
0
BARLEY
WHEAT
Results for AFLs (1997-2010)
15
10
Median
Number of
value (ppb) samples
over ML
414
0
0,47
0
1,4
0
3,0
0
0
0,26
0
8,8
0
2,0
0
Results for ηην ZON (1997-2010)
Results for DON (1997-2010)
Number of Samples
Barley
Number of Samples
Wheat
Parameter Number of Number of
samples
local
samples
DON
9
5
OTA
13
7
Aflatoxins
68
22
ZON
7
5
DON
5
5
OTA
12
1
Aflatoxins
26
11
ZON
11
10
Number of Samples
Sample
of the levels of mycotoxins in wheat and barley from 1997-2010
80
60
No of samples
No of local samples
No of positive samples
40
20
0
BARLEY
WHEAT
Results and Discussion
Results 2008-2010
61 LOCALLY produced samples of wheat, barley & products were
analyzed, 21 of which were positive to some mycotoxins: AFs, OTA .
 163 analyses for 116 samples of wheat, barley, corn. & products were
analyzed, 38 positive samples were found for some mycotoxins: DON,
ZON, AFs, OTA.
• For DON 24 positive samples found, levels ranging between 64,7
and 709 ppb, within the maximum limit (ML=1250 ppb) for cereals
and of 750 ppb for cereal products.
• For ΟΤΑ 8 positive samples were found with the levels ranging
between 0,06 and1,57 ppb, within the ML of 5 ppb for cereals and of
3 ppb for cereal products.
• For the four Aflatoxins AFs 3 positive imported corn samples were
found with the levels ranging between 0,36 and 21,67 ppb. One corn
sample exceeded the ML of 10 ppb.
• For ΖON 3 positive samples were found with the levels ranging
between 4,7 και 14 ppb within the ML of 100 ppb for cereals and of
75 ppb for cereal products.
•. For Τ-2 and ΗΤ-2 no positive samples were found.
Results and Discussion for local wheat + barley 2008-2010
• 85 analyses for DON, ZON, OTA, AFs in 38 samples of local wheat+ barley
• Only 2 positive samples to ΟΤΑ were found (1 Wheat and 1 Barley), within the
maximum limit of 5 ppb for cereals.NO FUSARIUM TOXINS WERE DETECTED! These
findings are in accordance with relevant info for DON, ZON, OTA, , Afs, Fum B1
+B2 from the Agricultural Services!
Results for ΖΟΝ in Wheat and Barley
8
20
6
16
No of samples
No of samples
Results for DON in Wheat and Barley
4
2
12
8
4
No of positive samples
0
2008
2009
2008
2010
Results for ΟΤΑ in Wheat and Barley
No of local samples
2009
2010
Results for AFLs in Wheat and Barley
20
15
16
No of samples
No of samples
No of positive samples
0
No of local samples
12
12
9
6
3
No of positive samples
0
2008
No of local samples
2009
2010
8
4
No of positive samples
0
2008
No of local samples
2009
2010
Finalization of the project
1. Completion of analysis of local wheat, barley etc and hay
for mycotoxins, as hay is stored in the fields.
2. Completion and evaluation
producers/farmers.
of
questionnaires
by
3. Processing and utilization of the results- enrichment of the
database.
4. Final meeting with the EMTOX partners of the project,
final results.
5. Arrangement of Meeting for informing all stakeholders in
Cyprus, may be in May 2011.
6. Continue of monitoring of local
mycotoxins for the next years
wheat, barley etc for
Results of the completed
questionnaires by farmers
 8 Cypriot cereal farmers, mainly wheat
 11 Questionnaires ( 1 farmer for wheat +barley,
1 farmer for wheat + barley + oat)
• 7 Wheat samples
• 3 Barley samples
• 1 Oat sample
11 questionnaires 9 wheat, 2 barley, 1 oat
1. What is your postal code?
All farmers in Nicosia
district area (11)
2. Optional: fill in your farm identification
code
5 of 8 farmers answered (8)
3. Do you grow wheat in 2011 in one or more
plots?
1 plot (1), > 1 plots (10)
If so, would you please fill in the questions below for one particular random
plot? The sample at harvest needs to be taken form the same plot.
4. Which name or code do you use for this plot? 7 of 8 farmers answered
(10)
5. What is the soil type of this plot?
Red soil (10), Different soils
(1)
6. What was the main cultivation of the soil of
this plot? E.g. deep tillage / surface tillage/ ….
Surface tillage (8), Deep
tillage (2), Tillage with
plow (1)
7. What crop did you grow on this plot in the
previous season?
Wheat (2), Potatoes (5),
Barley (2), Fallow (2)
8. What was the sowing date of the wheat in
this plot?
12/10/2009 – 30/12/2009
9. What wheat cultivar variety (or hybrid) did
you sow and – if known- what is the resistance
number for Fusarium spp. for this hybrid?
Cypriot wheat varieties :Ourania (6), Dimitra (1),
Athieni (1), Ekavi + various genetic lines (1)
10. What is your address and phone number?
All the plots are located in Nicosia district
11. What is the flowering date of the wheat in
this field? The flowering date is the date at
which 50% of the wheat field is flowering.
Beginning of February – End of April 2010
12.When do you expect to sow the wheat?
End of October 2010 – January 2011 (5), After the
rain (3), Fallow (1)
13. Which fungicide did you use during
cultivation of the wheat and during which
phenological phase did you apply this? Please
fill in the scheme below
Fungicide used
Monival (1), Pyriphos (6), Denocamines (2),
Douspan (1), Mancozeb (1), Vitavax (1), None
(2)
14. What was the expected harvest date of the
wheat?
Period of application (crop stage)
First stage of growth (10-15 cm) (4), Beginning of
March (15-20 cm) (2), 25-30 cm of growth (1), Before
seeding (1)
Beginning of May – June 2010
15. What was the actual harvest date of the
wheat?
1/6/2010 – 6/8/2010
Did you postpone harvesting due to bad
weather conditions?2 If so, by how many days?
7 of 8 farmers answered “No” (10)
Discussion of Questionnaires
• 11 Questionnaires, 8 farmers 12/10/2009 – 30/12/2009,
mainly wheat.
• Cypriot wheat varieties Ourania (6), Dimitra (1), Athieni
(1), Ekavi, Most resistant for Cypriot climate is Ourania.
• Red soil, surface tillage more of the farmers
• Fungicide used: Chloropyriphos (Pyriphos+Douspan),
Dithiocarbamates (Mancozeb, Vitavax).
• Trials are made by Agricultural Research Institute for new
hybrids of wheat (wheat + rye) to improve resistance to
climatic changes of Cyprus (less irrigation and more
salinity).
• Most of the wheat consumed in Cyprus is imported and a
small quantity of the locally produced is used for human
consumption while the rest + barley is used as feeding
stuff.
Conclusions
• No fusarium toxins (DON, ZON, HT-2, T-2) were
detected in Cypriot wheat and barley samples (38) for
2008-2010 in contrast with what is observed in other
European countries.
• Cypriot variety of wheat Ourania is the most resistant
to fusarium spp. and adapted to climate conditions
• Only 2 positive local samples for OTA were found.
• These findings are in accordance with relevant info
from the Agricultural Service for DON, ZON, OTA, Afs,
Fum B1 +B2!
• In general, all the analyzed samples (116) of wheat,
barley, corn, flour and pasta were within the MLs of EU
legislation.
– Only 38 positive samples were found for some mycotoxins:
DON, ZON, AFs, OTA
Acknowledgements
• To Mrs E. Christou, Head of Food Contamination &
Natural Toxins Lab of SGL
• To Mrs A. Anastasi, Head of IT Unit of SGL
• To Dr. M. Christophidou, Mrs M. Christodoulou. Mr D.
Stefani, Dr D, Kafouris, K.Roti, analysts in the above
Lab. of SGL
• To Dr H.J. (Ine) van der Fels-Klerx coordinator and other
EMTOX partners
• To Dr Rebecca Chrysafi of Research Promotion
Foundation of Cyprus