March 25, 2013 | By Dr. Terry Mabbett

Getting the Measure of Mycotoxins

Testing controls grain quality along every step of the supply chain

Getting the measure of mycotoxins is no easy task especially in the wide range of cereal crop commodities providing feed grain and the main ingredients of finished feed. Many field fungal pathogens and storage molds synthesize mycotoxins. Each group of fungi, including the aflatoxin-producing Aspergillus molds or the large number of Fusarium fungal pathogens delivering a broadside of Deoxynivalenol (DON), Zearalenone (ZEA), T-2 and HT-2 mycotoxins, has its own environmental requirements.

Each mycotoxin is the signature chemical and calling card left by a specific fungus or group of related fungi. Many of these highly versatile microbes "start life" as disease causing pathogens on cereal crops and subsequently extend into grain spoilage and mycotoxin contamination at any stage along the supply chain from on-farm storage to bags of finished feed.

Mycotoxins in the field

Field factors including fungicide treatment (by seed dressings or foliar sprays), fertilizer treatments and irrigation regimes, and the inherent disease resistance of the corn or wheat variety grown will play a part in the nature and magnitude of mycotoxin contamination. And watching over all this cereal agronomy is the weather as a wild card. The majority of cereal pathogens, and certainly Fusarium spp such as Fusarium graminearum responsible for wheat head blight and stalk and ear rot in corn, respond favorably to cool, cloudy, moist and humid conditions

Other fungal molds prefer it hot and dry. Classic case is the aflatoxin producing Aspergillus fungi, mainly A. flavus and A. parasiticus. They responded to the unprecedented high temperatures and accompanying drought experienced across the U.S. Corn Belt in 2012 with rapid growth rates and aflatoxin contamination in maize grain. Aspergillus spp are predominantly storage fungi and do not generally contaminate cereal grain prior to harvest. However, drought stress and insect damage often high in dry weather and drought stressed crops may allow infection by Aspergillus fungi and therefore aflatoxins production prior to harvest.

Mycotoxins moving into store

Mycotoxin first appears on the panicles of standing cereal crops, but cleaned grain arriving at the farm silo or loaded onto trucks for off-farm shipment is the first opportunity to test for what and how much mycotoxin is there. Getting the measure of mycotoxins from now on is a matter of what to test for, at what stage and how often.

Farmers and traders generally know the range of mycotoxins they need to test for in relation to the type of cereal and where grown. However, the appearance of one mycotoxin can often act as a "marker" for others because both are produced by closely related molds enjoying similar field conditions for infection and mycotoxin production. For instance, DON and ZEA will often occur together. These two mycotoxins are produced by Fusarium graminearum and also by a number of other closely related Fusarium fungi which infect a range of cereal crops.

Proper grain cleaning to remove all crop debris and especially the glumes (integuments surrounding small grain cereals like wheat) go a long way in preventing mycotoxin and fungal molds responsible from entering the post-harvest and processing stage.  The extent to which mycotoxin making molds become active in store to contaminate grain, and subsequently animal feed, will depend on grain moisture content and the conditions of storage. 

Maintaining the balance

Twelve percent grain moisture is generally given as the figure below which fungal mold activity ceases, but the situation is more complex than that. Moisture level within the grain and in the surrounding air is dependent on temperature because warmer air has a greater water holding capacity

Moisture inside the kernels of stored grain establishes an equilibrium level (balance) with the air outside, and the resulting relative humidity (R.H.) may be sufficiently high to encourage growth of deteriorative organisms including mycotoxin making fungi. Bacteria, fungal molds (including mycotoxin producers) and mites require a minimum R.H. of 90%, 70% and 60%, respectively. Insects depending on species need an R.H. level between 30% to 50%.

Grain storage specialists utilize this information to relate equilibrium moisture content of grain and R.H. of the surrounding air for a range of stored cereals. Relationships calculated for a range of cereals stored at a temperature of 25C are shown in Table 1. Grain moisture content in equilibrium with an R.H. of 70% (shown in bold type) is the figure beyond which the stored cereal grain becomes at risk of microbial damage and therefore mycotoxin contamination. In practical terms this means grain scheduled for storage at 25C should be dried to and maintained at that maximum moisture level. Equilibrium grain moisture contents and corresponding R.H. levels are recalculated for higher or lower storage temperatures.

Getting the measure of mycotoxins

Exactly when and where to test along the supply chain, is now formalized into Hazard Analysis Critical Control Point (HACCP) testing. Experienced operators will already be aware of inherently high risk points where testing has always been advised. This may be grain loads coming in from different parts of the farm and experiencing different growing conditions including soil moisture and irrigation levels. Mycotoxin testing will alert the farmer to any real-time mycotoxin problems and provide important farm and crop data for use in later years.

Routine testing of grain loads from different sources for mixing in silos or during feed manufacture is clearly a priority for managers of central grain depots and feed mills. Any point along the supply chain where grain and other debris can accumulate, whether in conveyors at the grain store or feed mixers and bins in the feed mill, are high risk points for fungal growth and mycotoxin accumulation.

Mycotoxins are not randomly or uniformly distributed throughout static grain or feed loads but tend to occur in so called "hot spots" corresponding to damp spots and pockets of mechanically damaged or insect infested grain that encourage mould growth. Sampling a moving stream of grain, feed ingredient or finished product reduces any selection bias associated with sampling and testing a static load. Static grain loads should be probed many times and all over, with sub samples bulked to produce a more representative gross sample for testing.

Speed and sensitivity of testing are the twin main thrusts of mycotoxin testing over the last two decades, including actual time taken to obtain an accurate and actionable reading and result. If a test result is achieved on-site within minutes, then "rogue loads" can be dealt with promptly and isolated without contaminating the main bulk of grain, feed ingredient or finished feed. When samples were taken back to laboratories for testing, no prompt action was possible for suspicious loads with visible mold or a musty smell, without holding up operations.  Traditional methods to analyze mycotoxins include fluorometer and LC methods using immunoaffinity (IA) columns. Chosen for their ability to isolate mycotoxins in simple or complex sample types, IA columns ensure precise measurements whether in the field or laboratory setting and provide the most comprehensive range for detection of single or multiple mycotoxins. 

Maximum protection against the threat of mycotoxin contamination is achieved when inbound raw materials such as rice, corn, wheat, cottonseed meal or barley are screened prior to storage or use in feed production. This need to know on-site and on time led to the development of portable on-site testing equipment like the VICAM Vertu® Lateral Flow reader — used to identify and quantify key mycotoxin hazards at critical control points right along the feed and grain supply chain. VICAM has gone one stage further with introduction of five-minute on-site quantitative strip tests to detect DON (DON-V™) and alflatoxins (Alfa-V™). 

VICAM’s quantitative strip tests are designed for use with the Vertu® Lateral Flow Reader and can be performed with a minimum of on-site training and technical expertise. These novel tests employ the highly sensitive and selective monoclonal antibodies required to accurately measure DON and alfatoxins in grain or feed material.

Current passion is to drive down testing times to several minutes. More important is the overall on-site picture to allow prompt testing by nonscientists, on the spot and anywhere along the supply chain, to secure prompt accurate and repeatable results over the sensitivity range required. Whether the actual testing time is three or five minutes is secondary. 

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