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Mycotoxins, climate change and food security: do we know enough?
Prof Naresh Magan DSc
Dr Angel Medina and Dra Alicia Rodriguez
Applied Mycology Group
AgriFood Institute, Cranfield, Bedford MK43 0AL, U.K.
Contents of my talk
Background to climate change issues –context for spoilage moulds/diseases and mycotoxins in staple food crops
Impact of water stress x temperature on mycotoxigenic fungi
Effect of water stress x temp x CO2 on growth and toxin production
Modelling climate change on a global scale to predict impacts on mycotoxins
Future perspectives
Food security is a global issue: prices of somestaple grains have increased in the last 5 years
Climate change: will put pressure on foodsupply, quality, and sustainability world-wide
What will changes in rainfall patterns, drought,temperature and CO2 have on staple foodproduction systems pre- and post-harvest?
Interaction between plant stress andfungal/pest infections will impact on fungalspoilage and mycotoxin contamination
Estimates: +25% losses due to fungal spoilage
Background
Predicted temperature change models in relation to industrial activity
0
100
200
300
400
500
600
700
1920 1940 1960 1980 2000 2020 2040 2060 2080
CO2 (ppmv)
CO2…
Predictions of CO2 based on the available models
Drought /rainfall patterns: total annual rainfall patterns may shift and interact with temp. & CO2 changes
Bebber et al. (2013; Nature Climate Change) have predicted: plant pathogens and pests are moving at about 3-5 km/year towards
the poles.
Expected additional no. of pests per country. Predicted from their model relative to
current levels (Bebber et al. 2014. New Phytologist)
−200 −150 −100 −50 0 50 100 150 200 250 300 350 400 450 500 550
Fischer & Knuttl (2015). Anthropogenic contributions to global
occurrence of heavy precipitation and temp. extremes. Nature
Climate Change: reduction in extremes from 1000 to 200 days
Climate change threatens food
security
Effects of climate change on crops?
Maize: Increasing need for irrigation; otherwise
>5-10% yield penalty; no maximum CO2 levels
determined
Rice: needs 12-38oC for canopy development;
Doubling CO2 promotes photosynthetic rate by
30-40%; also increases rice biomass by 20-25%
Soybeans: water stress would increase
vulnerability; higher temps adversely affect yield
by 40-50%
CO2 % change % change oC % change % change
> ambient biomass yield increase biomass yield
Yr
1 700 +12 +15 +4 -14 -18
2 700 +26 +26 +4 -16 -35
3 700 +17 +16 +4 -1 -6
Mean: +18 +19 -10 -20
Using “Climatron” environmental growth chambers (David Lawlor,
2000; Rothamsted Research)
Effects of increased CO2 (x 2) and temperature on biomass and grain production of winter wheat
[controls at 350 µl/l]
Different predictions by different studies
Climate change can affect crop diseases:
what impacts on mycotoxin production?
Crop(resistance)
Pathogen(changing population)
Environment(changing
climate)
Pre- and Post-
harvest spoilage &
Mycotoxin severity
Our focus has been in mycotoxins – why?
Mycotoxins - naturally occurring toxic secondary metabolites
heat stable/difficult to destroy even during processing
Produced by Aspergillus, Penicillium, Fusarium and Alternaria genera
Contaminated at any time: field, harvesting, drying, transport and storage, milling and in finished products
Low mould counts do not mean that food is free of mycotoxins, as moulds can die but the toxins will remain
Comparison of potency of carcinogens in test animals
Compound Dose * Relative potency
trichloroethylene 3 1
carbon tetrachloride 0.02 150
benzidine 0.005 600
dimethyl nitrosamine 0.0005 6000
Sterigmatocystin 0.00003 100,000
aflatoxin B1 0.000001 3,000,000
* = g/kg/day to produce tumours in 50% of test animals over a life time
ochratoxins
FumonisinsAflatoxinsTrichothecenes
Raw materials, moulds and mycotoxins
Commodities for which EU legislative limits existAflatoxin OTA Patulin Fusarium toxins
_______ ______ _______ ____
Groundnuts + +
Nuts + +
Dried fruit + +
Cereals + + +
Maize + + +
Spices + +
Baby foods + + + +
Coffee +
Cocoa +
Grape juice +
Fruit + + + (apple)
Milk, egg +
Wine +
Climate change impacts on mycotoxins: directly and indirectly? Increased plant diseases/pest reproduction and
interaction with plants? YES Effects on biodiversity of microbiota on plant
surfaces? YES Will this lead to more mycotoxin contamination
of staple crops??? Will different mycotoxins become important on
a regional basis??? Will this make existing legislation out of step
with potential problems??? EU borders: RASFF 30% rejections-mycotoxins
Maize silkingdates in (a) 2016 as reference and (b) + 5 °C scenario in 2050.
Significant impacts on fungal infection/pests
Battilani et al., 2012. EFSA Report
(a)
(b)
Maize harvest date in (a) 2016 as reference and (b) + 5 °C scenario in 2050.
Impacts on diversity, infection and toxin contaminstion
Battilani et al., 2012. EFSA Report
Maize: Map of predicted risk of aflatoxin B1 contamination in maize in the +2 and + 5°C climate change scenarios. Used predicted 2079 meteorological conditions vs existing ones (Battilani etal., 2012. EFSA Report). No account of CO2
+2oC +5oC
Ecology of mycotoxigenic fungi
Consider different interacting factors which will affect growth and mycotoxin production
Water stress (aw) x temperature changes
Climate change: Aw x temperature x CO2
1.00
0.90
0.80
0.70
0.60
0 10 403020 50
Temperature (oC)
Wat
er
acti
vity
(a w
)
Fusarium/Alternariaspecies
Penicilliumspp
Aspergillus spp.
Diagrammatic profiles of growth/no growth limits for the three key mycotoxigenic genera
Potential changes in mycotoxins due to Temp and water stress interactions
Aw Tmax range T+3 T+5
/temp
Alternariol 0.95 100-500/25 20-40 5-20(A.alternata) 0.90 5-20/25 0 0
Ochratoxin A 0.95 >50/20 >50 30-50(P.verrucosum) 0.90 30-50/20 30-50 30-50
Ochratoxin A 0.95 1500-2000/20 1000 0-500(A.carbonarius) 0.90 500-1000/20 0-500 0-500
(Pen/Asp: ng/g) Magan et al. (2011) Plant Pathol.
Integrating molecular, ecological stress and toxin data: A.flavus and aflatoxin B1
Water x temperature impacts
Systems approach: combining gene
expression, growth and aflatoxin B1 production
Expression of 10 genes in the biosynthetic
pathway: aflF, aflD, aflE, aflM, aflO, aflP,
aflQ, aflX, aflR, aflS
+
Radial Growth +Aflatoxin production
Used a mixed growth model: growth associated product
formation model (Shuler and Kargi, 2007)
Comparison of the observed vs. predicted aflatoxin B1 production.
0
0.5
1
1.5
2
2.5
3
3.5
4
0 1 2 3 4
Alat
oxin
pre
dict
ed (p
pm)
Aflatoxin observed (ppm)
Model validation outside of the experimental area
Factors Conditions
Temperature (oC) 37 37 40
Water activity (aw) 0.90 0.95 0.90
Growth, µ (mm day-1) 0.29 0.59 0.14
Observed AFB1 production
(µg g-1)
3.96±0.20 2.68±0.14 0.00
Predicted AFB1 production
(µg g-1)
4.90±0.00 3.75±0.18 0.00
Table 4. Model validation outside the regions in which the model was developed
Abdel-Hadi et al. (2012), & Medina et al. (2013) Journal of Royal Soc. INTERFACE.
Climate change: important factors to
consider
Water stress (water activity, aw)
Temperature fluctuations (+2 to +5oC)
CO2 (x2 and x3 present value)
Very limited data if any on
mycotoxigenic pathogens
Temperature, water and CO2 interactions on growth, gene expression and aflatoxin B1
production by A. flavus
0
2
4
6
8
10
12
14
350 350 350 650 650 650 1000 1000 1000
0.97 0.95 0.92 0.97 0.95 0.92 0.97 0.95 0.92
Gro
wth
rat
e (
mm
of
rad
ium
/day
)
CO2 concentrationWater activity
A. flavus NRRL3357-YES-34oC
0
2
4
6
8
10
12
14
350 350 350 650 650 650 1000 1000 1000
0.97 0.95 0.92 0.97 0.95 0.92 0.97 0.95 0.92
Gro
wth
rat
e (
mm
of
dia
mte
r/ d
ay)
CO2 concentrationWater activity
A. flavus NNRL 3357-YES-37oC
x 2 CO2 x 3 CO2Air
A.flavus: absolute aflD expression relative to the control (=0.99 aw/30oC)
0.6
0.8
1
1.2
1.4
1.6
1.8
300.91 0
300.91650
300.911000
300.99 0
300.99650
300.991000
370.91 0
370.91650
370.911000
370.99 0
370.99650
370.991000
Calibrator: 30ºC, 0.99 aw, 350 ppm CO2
Rel
ati
ve
exp
ress
ion
Regulatory gene: aflR gene relative expression: A. flavus cultures on Glucose-based medium Calibrator: 30ºC, 0.99 aw, 350 ppm CO2
0.6
1.6
2.6
3.6
4.6
5.6
6.6
30 0.91
350
30 0.91
650
30 0.91
1000
30 0.99
350
30 0.99
650
30 0.99
1000
37 0.91
350
37 0.91
650
37 0.91
1000
37 0.99
350
37 0.99
650
37 0.99
1000
Re
lativ
ea
flR
ge
ne
exp
ress
ion
Temperature, water activity and CO2 concentration
0
200
400
600
800
1000
1200
1400
1600
1800
350 650 1000 350 650 1000 350 650 1000
0.97 0.97 0.97 0.95 0.95 0.95 0.92 0.92 0.92
ng
of
AFB
1/g
CO2 concentrationWater activity
A. flavus NRRL 3357 - AFB1
production at 37oC on YES
Summary impacts: A. flavus NRRL3357-YES
Temp (oC) aw CO2 (ppm) aflD aflR AFB1
34
0.97650 = = =
1000 = = =
0.95650 = = =
1000 = (x3.6) =
0.92650 = (x24.4) (x2.6)
1000 = (x2.0) (x2.0)
37
0.97650 (x4.6) = (x30.7)
1000 (x6.5) = (x23.8)
0.95650 (x6.4) (x14.6) (x79.2)
1000 (x3.2) (x43.9) (x78.5)
0.92650 = (x40.4) (x15.1)
1000 (x22.5) (x1680) (x23.8)
Medina et al. (2014) Frontiers in MicrobiologyMedina, A., Rodríguez, A., Sultan, Y & Magan, N (2015). Climate change factors and A.flavus: effects on gene expression, growth andaflatoxin production- World Mycotoxin Journal.Medina, Rodriguez and magan (2015). Current Genetics. In Press.
Stimulation of relative expression of biosynthetic pathway genes
by these 3-way interacting factors and increasing phenotypic
AFB1
Studies on maize: A. flavus NRRL3357
Comparison of relative gene expression and
aflatoxin production on maize under normal and
elevated conditions of CO2 x temp x aw
Temp (oC) aw CO2 (ppm) aflR AFB1
30
0.99650 = =
1000 = =
0.91650 (x 2) (x2.7)
1000 = =
37
0.99650 (x2) (x3.6)
1000 (x3) (x5.0)
0.91650 (x2) (x1.8)
1000 = (x1.5)
RNA sequencing now completed and being analysed in collaboration with NCSU, USA and USDA,.
Coffee: contamination with ochratoxin A
Contaminated with:
Aspergillus westerdijkiae (A. section Circumdati)
A. carbonarius (A. section Nigri)
0
20
40
60
80
0.95 0.98 0.99
OT
A (
ng
g-1
)
Water activity
A. westerdijkiae (B 2) 30°C
Effect of aw x CO2 on OTA production by A.westerdijkiae grown on a coffee medium at 30
and 35°C (Akbar & Magan, unpublished data)Note: scale ranges are different for OTA production.
0
1000
2000
3000
4000
5000
6000
0.9 0.95 0.97
OT
A (
ng
g-1
)
Water activity
(a) A. westerdijkiae (B2)
30°C
Effect of aw x CO2 x temperature on OTA production by A. westerdijkiae (B2) on coffee beans stored at (a) 30 and (b) 35°C (Akbar & Magan, unpublished data)Note: scale ranges are different for OTA production.
Models which have been used to predict mycotoxins in relation to climatic conditions
Predictive models: for Fusarium head blight and DON uses regional weather parameters during ripening (DONCAST)
Geostatistics approaches: to identify spatial hotspots which are high risk areas for mycotoxins (Europe; USA)
Agricultural production system simulators: in AustraliaDeveloped an Aflatoxin Risk Index (ARI) for peanuts/maize
Maize: Related seasonal temp and soil moisture during critical silking period to determine the ARI. Hot/dry conditions. Peanuts: Fractional amounts of available soil water during pod filling determined the ARI. Real time model developed (Chauhan et al., 2008, 2010)
Variable N° of stations EMDa
Mean Temperature 20828 48.36
Mean Minimum
Temperature11550 64.94
Mean Maximum
Temperature11544 64.96
Rainfall 27375 41.71
World meteorological data available in FAOCLIM
Variable Africa Asia America Europe Asia Antarctica World
Mean T 605 1256 2765 765 515 90 5996
Rain 3395 2172 5611 1389 915 48 13530
Inventory of time series (number) of variables considered according to
continents.
Worldwide prediction
Risk map – A. flavus in maize
Relative risk maps for ochratoxin A in grapes
Predictive models which exist at present: are they good enough to predict impact of climate change?
Impact of drought stress x CO2 changes on crop physiology and interaction with mycotoxinproducing fungi – required urgently
Cycling of drought/temperature events - effects on plant /fungus interface and mycotoxin production
Metabolomics: changes in mycotoxins produced by specific species??? Ratios ?? Switch between mycotoxins?
Future perspectives I
New mycotoxins by existing drought tolerant fungal species pre- and post-harvest??
Relationship between climate change factors, especially temp x CO2 increases may stimulate pest reproduction in staple crops
This could have a significant impact on mycotoxincontamination by causing damage
Food security in both developing and developed countries could be profoundly compromised under such scenarios
Future perspectives II
Acknowledgements:
Dr Angel Medina, Dr David Aldred, Dra. Alicia Rodriguez, Esther Baxter (Cranfield); Pof. Rolf
Geisen, Dr. Markus Schmidt-Heydt (Max RubnerInstitute, Germany; Dr. Roberto Parra (Monterey
Tech, Mexico)
Dr Deepak Bhatnagar (USDA) and Prof. Gary Payne (NCSU, USA) help with RNA Seq of A.flavus
(analysis in progress)
Pre-harvest: Wheat fusarium ear blight
• Decreases yield
• Produces mycotoxins (damage human/animal health)
• Important across Europe, North and South America
Climate change – wheat flowering dates
predicted to be earlier: implication for
FHB and mycotoxins?2050s High CO21980s
Madgwick et al., 2011, Eur J Plant Path 130, 117-131
Climate change – predicted increase in %
plants with Fusarium ear blight: what does
this mean for mycotoxins - DON??1980s 2050s High CO2
0
400
800
1200
1600
0.95 0.98 0.99
OT
A (
ng
g-1
)
Water activity
(a) A. carbonarius (ITAL 204) 30°C
0
20
40
60
0.95 0.98 0.99
Water activity
(b) A. carbonarius (ITAL 204) 35°C Air (400ppm)
CO2 (1000ppm)
Growth of F. graminearum: elevated temperature and CO2 (1000 ppm; 3x ambient)
conditions at 2 aw levels (Medina & Magan, unpub. data)
0
5
10
15
20
25
25 30 35 40
Gro
wth
rat
e (
mm
dia
met
er/
day
)
Temperature oC
Fusarium graminearum
aw 0.995+ CO2
aw 0.98+CO2
aw 0.995
aw 0.98
0
0.5
1
1.5
2
2.5
3
GS50 GS60 GS65 GS75
350
700
350+4C
700+4C
Growth stage
Log
10
CF
Us
cm2
Biodiversity: fungal populations affected by climate change on flag
leaves/ears of wheat
Magan & Baxter, 2000
Different sub-arrays shown in different colours red = ochratoxin A, light brown = aflatoxin, dark blue =
trichothecenes (type A), light blue=trichothecenes; dark green =fumonisins
This corresponds to the colours of the frames surrounding thehybridized spots after microarray analysis
Schmidt-Heydt & Geisen (2007) Int. J. Fd Microbiol.
MYCOCHIP ARRAY
Maize: colonisation by A. flavus NRRL3357 at 30oC
-1
1
3
5
7
9
11
13
15
17
19
350 650 1000 350 650 1000
0.91 0.99Re
lati
ve
afl
R g
en
e e
xp
res
sio
n
Water activity x CO2 concentration
aflR gene (30°C)0
0.2
0.4
0.6
0.8
1
1.2
350 650 1000 350 650 1000
0.91 0.99Rela
tive a
flD
gen
e e
xp
ressio
n
Water activity x CO2 concentration
aflD gene (30°C)
Key: where P is the AFB1 production (ppm) and b1, b2, α, and β are parameter estimates from the model and µ was calculated based on a period of 9 days growth.
[g] – linear model for gene expression
Abdel-Hadi et al. (2012) Journal of Royal Soc. Interface (aflatoxins)Medina et al. (2013). Journal of Royal Soc. Interface (fumonisins)
11
0
tRT
ab
eeXgPw
Linked: Gene expression; growth, aflatoxin B1 production
Biosynthetic genes: aflatoxin production: Grouped according to expression profile at temperature x water activity combinations, to the gene cluster. Group 1 – light grey; Group 2 – dark grey (Schmidt-Heydt et al. 2010: Int J Fd Microbiol)
Ratio of aflS/aflR in relation to aflatoxin biosynthesis and
different parameter combinations
Parameter combination Ratio aflS/aflR Aflatoxin [ng/g]
25oC/0.90 0.5 3.7
35oC/0.90 0.3 4.7
25oC/0.95 7.4 830.2
30oC/0.95 7.1 3016.9
25oC/0.99 1.5 1957.3
30oC/0.99 2.7 2758.7
0
50
100
150
200
250
300
350
350 650 1000 350 650 1000
0.91 0.99
Re
lati
ve
afl
D g
en
e e
xp
res
sio
n
Water activity x CO2 concentration
aflD gene (37°C)
0
20
40
60
80
100
120
140
160
180
350 650 1000 350 650 1000
0.91 0.99
Re
lati
ve
afl
R g
en
e e
xp
res
sio
n
Water activity x CO2 concentration
aflR gene (37°C)
Maize: colonisation by A. flavus NRRL3357 at 37oC
Boundaries for growth and mycotoxinproduction
We now have significant knowledge of the impact of environmental stress factors on mycotoxigenic fungi: germination, growth, toxin production
F.culmorum DONF.graminearum DONF.langsethiae T-2/HT-2F.verticillioides FumonisinsP.verrucosum OTAA.flavus AflatoxinsA.carbonarius OTAA.westerdijkiae OTAAlternaria alternata Alternariol, AME, TZA, ALTXII
Magan & Sanchis, 2004; Magan et al., 2011; Magan et al., 2015
Effects of climate change on
maize?
Increasing need for irrigation; otherwise
>5-10% yield penalty; no maximum CO2
levels determined
Climate change Threat for agriculture
Maize is sensitive to water
stress and A.flavus is able to
grow in drought stress
conditions
Crop diseases, food security and
climate change
Need information to guide strategies for
adaptation to impacts of climate change on crop
disease losses
Need publicity to show how improved crop
disease control can contribute to climate change
mitigation
Need vigilance to maintain/improve crop disease
control despite changing pathogen/climate to
ensure global food security
New mycotoxins by existing drought tolerant
fungal species pre- and post-harvest??
Relationship between climate change factors,
especially temp x CO2 increases may stimulate pest
reproduction in staple crops
Could have a significant impact on mycotoxin
contamination by causing damage
Food security in both developing and developed
countries could be profoundly compromised under
such scenarios
Aflatoxin biosynthesis gene cluster
The genes involved in aflatoxin production are clustered together
Yu et al. (2004)
Re
gu
lato
ryg
en
es
Sst
ruc
tura
lg
en
es Schmidt-Heydt et al. (2010)
- Some genes of this cluster act
together in different groups
- Ratio aflR/aflS & aflatoxin
production (under different
environmental conditions)
Abdel-Hadi et al. (2012)
- A.flavus can grow over a wide
range temperature x aw, butaflatoxin production is over a
narrower range.
- Relationship between activity of
aflD gene with regulatory genes
(T/aw)Sst
ruc
tura
lg
en
es
F. verticillioides CO2
0
2
4
6
8
10
12
14
16
24 25 26 27 28 29 30 31 32 33 34 35 36 37 38
Gro
wth
rat
e (
mm
dia
met
er/
day
)
Temperature oC
Fusarium verticillioidesaw 0.995+ CO2
aw 0.98+CO2
aw 0.995
aw 0.98
Growth of F.verticillioides under increased temperature and CO2 (1000 ppm; 3x ambient) conditions at 2 aw levels (Medina & Magan, unpub. data)
