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di Aspergillus flavus e festione della filiera, un future per maiscoltura e “latte italiano di qualità” 9 febbraio, 2017, Universita Cattolicà del Sacro Cuore, Piacenza

di Aspergillus flavus e festione della filiera, un future per maiscoltura e “latte italiano di qualità” 9 febbraio, 2017, Universita Cattolicà del Sacro Cuore, Piacenza

Peter J. Cotty,Agricultural Research Service, U.S. Department of AgricultureSchool of Plant Sciences, University of Arizona, Tucson

Peter J. Cotty,Agricultural Research Service, U.S. Department of AgricultureSchool of Plant Sciences, University of Arizona, Tucson

1970s, Arizona

1980s, ArizonaFirst Tests

1988

Vegetative Compatibility Analyses

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CAP = Cluster Amplification Pattern

Simple Sequence Repeat Analyses

First Patent Application: 1989 First Conference with Environmental Protection Agency: 1992 Used on commercial crops in US since 1996. Three Products Currently Registered Over 1 Million Acres Treated Annually Registered Target Crops: Maize Grain, Maize Silage,

Pistachios, Cottonseed, Peanut (Almond & Fig in registration process).

Aflatoxin Biocontrol in the US

Biocontrol in Africa There is Severe Human Exposure to Aflatoxins in Several

African Nations. Products registered for use in Nigeria, Kenya,

Senegal/Gambia. Target Crops: Maize & Groundnut.

The Wall Street JournalTuesday March 16, 1993

The Wall Street JournalTuesday March 16, 1993

Biocontrol of aflatoxins exists as a practical tool because of farmers.

Genotypes of Aspergillus flavus, an important cause of aflatoxin contamination, vary widely in ability to

produce aflatoxins.

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Per

cent

of

A. f

lavu

s

0

10

20

30

40

50

60

1 2 3 4 5 6

Field 1 Field 2

Aflatoxin Produced in Culture (log ppb)

Aflatoxin-ProducingPotential

Aflatoxin-ProducingPotential

Field 1 = Low,3,400 ppb

Field 1 = Low,3,400 ppb

Field 2 = High,54,000 ppb

Field 2 = High,54,000 ppb

Aflatoxin Production by A. flavus from Two Fields

Fungi Vary Across Areas in Aflatoxin-Producing Ability

Farmers apply atoxigenic strains to reduce the average aflatoxin-producing potential of fungi on farms & this reduces vulnerability of crops to aflatoxin contamination

Fungi Vary Across Areas in Aflatoxin-Producing Ability

Farmers apply atoxigenic strains to reduce the average aflatoxin-producing potential of fungi on farms & this reduces vulnerability of crops to aflatoxin contamination

Mycological Research 101:698-704; 1997.

As Applied After Fungal Growth

The atoxigenic fungi are applied on non-viable grain (sorghum, wheat, and barley are used).

Biocontrol Products: AF36, Prevail™, Aflaguard™, AflaSafe™, AF-X1™, FourSure™

As AppliedAs Applied

J F M A M J J A S O N DMonth

0

100

200

300

400

500

=total crop biomass=total crop biomass=total A. flavus=total A. flavus=soil A. flavus=soil A. flavus

Aspergillus Populations Rapidly Increase with Crop BiomassFounder Effects Drive Shifts in Population Structure.

Aspergillus Populations Rapidly Increase with Crop BiomassFounder Effects Drive Shifts in Population Structure.

Properly timed applications of atoxigenics can direct population shifts

to improve the safety of Aspergillus populations associated with crops

Afl

ato

xin

B1

(ng

/g X

10,

000)

Isolates (%) in Applied Atoxigenic Strain

0

1

2

3

4

5

6

7

8

9

0 20 40 60 80 100

Dots Represent Values for Replicate Plots

Aflatoxin in Crop versus Atoxigenic Incidence

Cotty, 1994. Phytopathology:1270-1277.

Aflatoxin (ppb)

Area Samples (#) AF36 (%) Mean Range

GraysonNorth

17 96 a 12 a 0 to 48

GraysonSouth

16 98 a 15 a 0 to 38

GraysonControl

8 24 b 230 b 5 to 530

Commercial Maize: North Central Texas 2008

Means in the same column with different letters are significantly different by Tukey’s HSD test, P < 0.001.

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Fungi move between fields and across areas.

Take a bigger view: Not just one field, one farmer, one crop.

We can protect whole agricultural systems from aflatoxins including all affected industries and improve the environment.

57% AF36

92% AF3660% AF36

42% AF36

1% AF364% AF36

3% AF36

0% AF36

Blue dots represent the percent of the A. flavus

community composed of AF36

Soil Community Before Treatment Community on Crop After Ginning

Application Rate = 10 lb/acre

0.5 miles

Treated Field Treated Field

Application of Atoxigenic Strain AF36 in Commercial Cotton Influences the Composition of Fungi on Crops in both Treated and Nearby Fields

Data from 564 vegetative compatibility analyses.

Field Fields

AF36

(% A. flavus)

S strain

(% A. flavus)

A. flavus

(CFU/gram)

type (#) 1996 1997 1996 1997 1996 1997

Treated 3 4% ab 85% a 52% a 4% d 582 a 365 a

Adjacent 4 2% b 48% b 41% a 18% c 411 a 157 a

Diagonal 4 2% b 16% c 52% a 33% b 61 a 100 a

Other 4 9% a 9% c 43% a 50% a 109 a 98 a

Composition of Aspergillus flavus Communities in Soil of Treated and Nearby Fields in May 1996 Prior to Application of AF36 and

in May 1997 One Year After Application

TreatedAdjacentOther

Diagonal DiagonalOther Other Other

OtherOtherAdjacent

Adjacent

82%

53%

96%100%

0% 0% 0% 1% 17% 11%

77%74%

0%

20%

40%

60%

80%

100%

Pa

rha

mS

oil

Ru

the

rfo

rdS

oil

Pa

rha

mC

orn

Ru

the

rfo

rdC

orn

Pa

rha

mC

orn

Ru

the

rfo

rdC

orn

2008 2008 2008 2008 2009 2009

Per

cen

t o

f A

sper

gil

lus

flav

us

Grayson County, Texas: Carry Over to the Second Year Crop

= Atoxigenic isolate AF36

= S Strain, produces very high aflatoxins

Crop Treatment Year

Crop Year After Treatment

Soil Before Treatment

Percent AF36™

Percent Aflaguard™

Percent other A. flavus

Not Treated in 2015 Atoxigenic Biocontrol Agents on Harvested Corn

From Commercial Field Northcentral Texas

Area-Wide & Multi-Year Effects in Commercial AgriculturePie charts indicate percent A. flavus composed of the two EPA registered

active ingredients on maize harvested from the indicated spots. Values in each pie result from Vegetative Compatibility Analyses on 16 isolates (160 total).

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Equator

15° N

30° N

Equator

30° S

15° S

Wuhan(37 m)

Nanning(80 m)

Nampula(431 m)

Machakos(1,138 m)

Kaolack(26 m)

Phoenix(331 m)

Peanut

Corn

Dallas(131 m)

Pistachio

Corn

ACRPC, Phoenix

IITA, Ibadan

KALRO Katumani, Machakos

Italy

Cotton

During Crop DevelopmentIrrigation, Weed Control, Fertilize

Best cultivars, Insect Control

During transportRapid, Dry, No Damage

During ProcessingSort, Cull, Discard, Add binders.

During StorageDry, prevent moisture, cool

Prevent damage: insects, rodents

Prevent Formation of Dangerous Aflatoxin Levels

Protect Crops From Field…

…to Mouth

Since 2009 thousands of farmer’s fields have been treated in Nigeria, Kenya, Senegal, Burkina Faso, Zambia, Tanzania, and Ghana.

Reductions in aflatoxin contamination consistently average over 80% both at harvest and after storage.

Single applications are made at 10 kg/hectare

Nigeria

Senegal

As in the US, in Africa the biocontrol fungi survive between seasons allowing

for long-term and cumulative benefit.

“Emerging Risks in the Global Food System”

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74 ppb - Treated1,133 ppb - Control

93.5% Reduction

Mutomo, Kitui County, Kenya: 2012 TestsShort Rain Season Farmer Field Trials

DeadlyAverage 2,750 ppb!

Range 1,790 ppb to 3,710 ppb.

Safe Food

510 ppb

Aflatoxin Content(ppb)

Selection of atoxigenics for Kenya: Probst, et al. 2011. Plant Disease 95:212-218.

0

100

200

300

400

500

600

700

Not Treated Treated

20 ppb

µg/

kg (

ppb)

Tot

al A

flato

xins

Nigeria: Treated and control crops were harvested and stored under poor conditions for four months.

Atoxigenics stay with the crop from treatment, ………through harvest, storage, and consumption.

Atehnkeng, et al. 2014. Biological Control 72:62-70.

Area-wide Programs provide efficacy across cropping systems and additional health benefits. Area-wide Programs are less expensive (per hectare), provide for long-term commitment to aflatoxin elimination, protect all crops and remediate the area’s reputation.

Biocontrol fungi move from treated fields across areas

Multiple Crops Benefit From the Same Biocontrol Fungus

Biocontrol is Area-wide Management Area-wide Aspects can be Optimized

Atoxigenics persist in soils, on crop debris, on non-crop plants

Manufacture is easy. Biocontrol products like AF-X1 are made by a Farmer Organization in Arizona, and in a factory in Nigeria. Factories to make biocontrol products are under construction in Kenya and Senegal. manufacturing under construction at Katumani Station of KALRO in Machakos, Kenya. The facility will make Aflasafe KE01

December 2014

April, 2016

Producing the Biocontrol Product:

Manufacturing

Agricultural Research Service, USDASchool of Plant Sciences, University of Arizona

Work in Africa led by: Dr. Ranajit Bandyopadhyay Int. Institute Tropical AgricultureIbadan, Nigeria

Work on Tree Crops led by: Dr. Themis Michailides

University of California, DavisParlier, California

Work in Italy led by: Dr. Paola Battilani Universitá Cattolica del Sacro CuorePiacenza, Italy

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Agricultural Research Service, USDASchool of Plant Sciences, University of Arizona

There are many atoxigenic strainsSelect strains best adapted to rotations, ecosystems, & climates

Crops are infected by complex communities of diverse fungiWe can influence aflatoxin-producing ability of fungal communities resident in production areas

through crop rotations, agronomic practice, and by applying atoxigenic strains

Atoxigenics are Already Present on the CropJust increasing the frequency of endemic strains & natural interference with contamination

Treatments have Long-Term Influences & Cumulative Benefits

All Crops in an Area May Benefit From Applications

Atoxigenic Strains can be Applied Without Increasing Infectionand without increasing the overall quantity of A. flavus on the crop & throughout the environment