1. by Hendrix Chalwe Zambia Science Conference-October 2016
PREDICTING PRE-HARVEST AFLATOXIN INCIDENCE IN GROUNDNUTS USING SELECTED SOIL AND WEATHER PARAMETERS
by Hendrix Chalwe
Zambia Science Conference-October 2016
Nov-18

2. INTRODUCTION: DEFINITIONS
Aflatoxins are secondary metabolites of fungal species Aspergillus flavus and Aspergillus parasiticus commonly found in agricultural crops such as maize, groundnuts, cottonseed, and tree nuts.
Aflatoxin-formation is usually associated with drought stress before harvest while moist, humid storage conditions increase susceptibility after harvest.
Aflatoxins are among the most potent mycotoxins.
The commonest classes of aflatoxins include B1 , B2 , G1 and G2
(Bennet and Klich, 2003)

3. STRUCTURES OF CLASSES OF AFLATOXINS

4. EFFECTS OF AFLATOXINS ON HUMAN AND ANIMAL HEALTH
High levels of aflatoxins can cause acute toxicity, and potentially death, in animals as well as in humans.
Humans: stunting in children, immune suppression, cancers and genetic mutations
Animals: liver damage , reduced production of milk and eggs in cattle and poultry, respectively.
(Richard, 2007).
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5. EFFECTS OF AFLATOXINS ON TRADE
High levels of aflatoxins limits access to international markets (Murphy et al, 2006).
For instance: ZABS Standard on peanut butter is 15 µg/kg (ZS 723, 2008).
In September, 2016, ZABS withdrew thousands of 1 L containers of Lyons peanut butter from shops in Lusaka resulting in a huge economic loss.
Nov-18

6. PREVALENCE OF AFLATOXINS IN GROUNDNUTS IN ZAMBIA
Groundnut (Arachis hypogaea L.) is the second most important food and cash crop of Zambia (Ross and Klerk, 2012)
However, the crop has very limited access to international markets due to high concentrations of aflatoxins in kernels.
Several reports reveal high concentrations of aflatoxins in raw and processed groundnuts
(Njoroge, et al. 2016; Ismail et al., 2014; Mukuka and Shipekesa, 2013)
Nov-18

7. MANAGEMENT OF PRE-HARVEST AFLATOXIN INCIDENCE
Traditional measures mostly target preventing fungal infection at field level (Murphy, et al 2006).
There exists several good agricultural practices (GAPs) that are capable of reducing aflatoxin levels in groundnuts including: resistant varieties, healthy seeds, crop rotations, appropriate weeding among others
The study focus is on GAPs that contribute to the effective soil water management to avoid stress during pod-development such as: timely planting, mulching, appropriate tillage and manures.
(Waliyar et al., 2013).
Since aflatoxin- formation begins at field level, the ability to predict pre-harvest aflatoxin concentrations offers an opportunity for timely and appropriate interventions
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8. OBJECTIVES Specifically:
To develop a statistical model that predicts pre-harvest aflatoxin content in groundnuts using selected soil and weather parameters
Specifically:
To evaluate the effect of end-of-season drought stress (low/no rainfall, higher ambient and soil temperatures, declining soil moisture) on pre-harvest aflatoxin concentrations in groundnuts.
To evaluate the effect of soil organic matter on pre-harvest aflatoxin concentrations in groundnuts.
To evaluate the effect of exchangeable calcium on pre-harvest aflatoxin concentrations in groundnuts.
To develop an aflatoxin risk index (ARI) using selected soil and weather parameters.
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9. MATERIALS AND METHODS Field and greenhouse trials
Conducted research trials on GAPs with an influence soil moisture, temperature and chemical fertility.
Field trials-based on common farming practices
Greenhouse trials-controlled environment and follow ups
The study sites included: Kasisi, University of Zambia, Liempe Farm and Msekera
Nov-18

10. DATA COLLECTION/STATISTICAL ANALYSES
At each of the study sites, land was ploughed conventionally with a tractor- mounted disc plough
Groundnut variety: MGV 4 (average yields ranging from 1.5 to 2.5 ton/ha and takes between days to mature).
Fully matured pods were harvested and sampled for aflatoxin testing
The sampled pods were dried to about 8 % moisture content, shelled and then tested for aflatoxin content using the Reveal Q+ Aflatoxin Testing Kit.
The data collected were subjected to either the ANOVA or the alternative Kruskal Wallis Test at 95 % Confidence Interval.
Nov-18

11. PROTOCOL FOR AFLATOXIN TESTING
Weigh 10 g of ground sample
Mix sample with 65% ethanol by shaking for three minutes.
Dilute the extract and insert a single test strip for 6 minutes
Read the aflatoxin concentration by placing the strip into an aflatoxin reader
Nov-18

12. EFFECT OF MOISTURE STRESS ON PRE-HARVEST AFLATOXIN CONTENT
A pot experiment was set up at University of Zambia, located at ’S and ’E
4 moisture levels equivalent to 25, 50, 75 and 100% crop water requirements from flowering to maturity.
There was 93 % reduction in total aflatoxin concentrations with increased water supply (p value=0.007, R-sq = 0.77).
Higher soil moisture availability is likely to enhance the production of phytoalexin and thus minimize fungal infections (Wotton and Strange, 1987).
Zambian Standards on aflatoxins in peanut butter (ZS 723: 2008), set at 15 ppb total aflatoxin.
Only the 100 % treatment produced groundnuts that would meet the tolerable limit.
Mean aflatoxin content as a function of water supply. Error bars represent SEM
Nov-18

13. EFFECT OF MOISTURE STRESS ON PRE-HARVEST AFLATOXIN CONTENT
A pot experiment was set up at University of Zambia, located at ’S and ’E
4 moisture levels equivalent to 25, 50, 75 and 100% crop water requirements from flowering to maturity.
There was 93 % reduction in total aflatoxin concentrations with increased water supply (p value=0.007, R-sq = 0.77).
Higher soil moisture availability is likely to enhance the production of phytoalexin and thus minimize fungal infections (Wotton and Strange, 1987).
Zambian Standards on aflatoxins in peanut butter (ZS 723: 2008), set at 15 ppb total aflatoxin.
Only the 100 % treatment produced groundnuts that would meet the tolerable limit.
Mean aflatoxin content as a function of water supply. Error bars represent SEM
Nov-18

14. EFFECT OF TYPE OF TILLAGE ON PRE-HARVEST AFLATOXIN B1
An experiment was conducted at Msekera Research Station located at ° S and ° E in Chipata, Zambia.
Design: RCBD consisting of 3 tillage types
Untied ridges
Tied ridges, and
Untied ridges with a grass mulch
There was 75 % reduction aflatoxin concentrations due to type of tillage (p value=0.049, R-sq = 0.95).
Tied ridging was most effective while mulching and the untied ridges remained less effective.
Results may be attributed to better soil moisture retention with tied ridging
Mean aflatoxin content as a function of tillage system. Error bars represent SEM.
Nov-18

15. EFFECT OF TYPE OF TILLAGE ON PRE-HARVEST AFLATOXIN B1
An experiment was conducted at Msekera Research Station located at ° S and ° E in Chipata, Zambia.
Design: RCBD consisting of 3 tillage types
Untied ridges
Tied ridges, and
Untied ridges with a grass mulch
There was 75 % reduction aflatoxin concentrations due to type of tillage (p value=0.049, R-sq = 0.95).
Tied ridging was most effective while mulching and the untied ridges remained less effective.
Results may be attributed to better soil moisture retention with tied ridging
Mean aflatoxin content as a function of tillage system. Error bars represent SEM.
Nov-18

16. EFFECT OF TIME OF PLANTING ON PRE-HARVEST AFLATOXIN B1
An experiment was conducted at Msekera Research Station located at ° S and ° E in Chipata, Zambia.
Design: RCBD consisting 3 phase planting, with 3 replicates
1st planting was at the on set of 1st effective rains, while the 2nd and 3rd planting were about a week apart after the first planting.
There was 95.7 % increase in aflatoxin concentrations with delays in planting (p value=0.001, R-sq = 0.99).
The increased aflatoxin content can be attributed to increased exposure to late season drought
Mean aflatoxin content as a function of time of plating within a season
Nov-18

17. EFFECT OF TIME OF PLANTING ON PRE-HARVEST AFLATOXIN B1
An experiment was conducted at Msekera Research Station located at S and E in Chipata, Zambia.
Design: RCBD consisting 3 phase planting, with 3 replicates
1st planting was at the on set of 1st effective rains, while the 2nd and 3rd planting were about a week apart after the first planting.
There was 95.7 % increase in aflatoxin concentrations with delays in planting (p value=0.001, R-sq = 0.99).
The increased aflatoxin content can be attributed to increased exposure to late season drought
Mean aflatoxin content as a function of time of plating within a season
Nov-18

18. RAINFALL AMOUNTS IN THE FINAL 4 WEEKS
Table 3. Aflatoxin contamination as influenced by seeding date in relation to drought stress during the latter part of the growing season.a
Planting date
Rainfall during final 30 days of the season
Mean aflatoxin B1 concentration in kernels
(mm)
(µg/kg)
31st December, 2014
99.0
4 a
8th January, 2015
27.3
37 b
15th January, 2015
0.0
89 c
aMeans for aflatoxin contamination followed by the same letter are not significantly different according to Fisher’s Protected LSD at p < 0.05.
Nov-18

19. EFFECT OF MANURE APPLICATION ON PRE-HARVEST AFLATOXIN CONTENT
Nov-18
Figure 3: Total aflatoxin content as a function of manure application
There was 27 % reduction in total aflatoxin concentrations due to the application manure (p value=0.04, R-sq = 0.84).
Waliyar et al. (2013) reported a 42 % reduction in total aflatoxin content using farmyard manure at a rate of 2.5 tons/hectare before planting.

20. EFFECT OF MANURE APPLICATION ON PRE-HARVEST AFLATOXIN CONTENT
Nov-18
Figure 3: Total aflatoxin content as a function of manure application
There was 27 % reduction in total aflatoxin concentrations due to the application manure (p value=0.04, R-sq = 0.84).
Waliyar et al. (2013) reported a 42 % reduction in total aflatoxin content using farmyard manure at a rate of 2.5 tons/hectare before planting.

21. EFFECT OF MANURE ON SOIL MICROBIAL ACTIVITY
Figure 1: Soil microbial activity as a function of manure application
Soil microbial activity increased with an increase in manure application (p =0.00, R-sq=0.9153)
Increased microbial activity likely to supress the dominating effect of Aspergillus
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22. EFFECT OF MANURE ON SOIL MOISTURE RETENTION
Nov-18
Figure 2: Volumetric water content as a function of manure application
Soil moisture retention increased with increased manure application.
A higher soil moisture content enhance the production of phytoalexins and thus minimize fungal infections (Wotton and Strange, 1987).

23. EFFECT OF GYPSUM APPLICATION ON PREHARVEST AFLATOXIN
Research site
Range of mean total aflatoxin content
Conclusions
Kasisi
ppb (3.5)
Application of gypsum did not affect the distribution of total aflatoxin across treatments
University Farm
ppb (3.0)
UNZA (Pot experiment)
ppb (22.6)
Three trials each consisting 4 levels of gypsum were set up at 3 different locations.
Contrary to many authors (Bowen et al, 1996; Waliyar et al. 2013, Guchi, 2015), the application of gypsum (source of calcium) did not show any significant effect on aflatoxin content, p-values > 0.05.
Nov-18

24. CONCLUSIONS Good soil water management through:
Appropriate tillage types (p value=0.04, R-sq = 0.95),
Application of manures (p value=0.04, R-sq = 0.84), and
Early planting (p value=0.001, R-sq = 0.99).
correlated well with pre-harvest aflatoxin concentrations in groundnut kernels.
Therefore, type of tillage, application of manures and timely planting are potential factors in predicting pre-harvest aflatoxin risk in groundnuts.
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25. (Major multi-factor and single-factor experiments)
WAY FORWARD
Conduct more focused field research trials on:
(Major multi-factor and single-factor experiments)
Planting dates-rainfall, temperature and soil moisture contents
Type of tillage-soil moisture and temperature fluctuations
Soil organic matter-soil moisture and temperature fluctuations
Minor research trials
Variety-ease of infection due to pod characteristics such as seed size, pod strength
Planting spacing-natural growth response
Gypsum applications-time and rates
Validation: Farmers fields in the vicinity of the experimental sites
Nov-18

26. THANK YOU
Nov-18