**2. Advantages of using infrared spectroscopy to manage fungal and mycotoxic risk for wheat, barley, and corn**

Fungus can be detected by microbiological methods involving visual, microscopy, and microbial-cultural methods.

Conventional methods of mold detection are based on direct observation by eye or by microscope of thalli, contaminated foodstuffs, or microbial cultures. These methods are time consuming and require viable samples and a good deal of expertise. Counting methods are difficult to apply to fungi because, during their reproduction, a spore generates a mycelium that can in turn divide itself into tens of individuals. Furthermore, a fungal contamination may not be visible at the surface of grains [Hirano, et al., 1998, Pearson, et al., 2001].

Other methods are based on molecular biology or on the detection of antigens specific to given molds. Organisms, either dead or alive, can be detected by the polymerase chain reaction (PCR) by copying a large number of DNA sequences that are originally present in small quantities (with a multiplicative factor on the order of 109). By amplifying certain genes of toxigenic strains, PCR serves as a tool to determine the risk. Various researchers have tested PCR to detect *Fusarium* contamination in corn [Jurado et al., 2006; Jurado et al., 2005; Nicolaisen et al., 2009]. These methods are rapid, sensitive, and can be automated. They are good qualitative methods (e.g., good selectivity) but offer only average precision in quantitative terms (they are called "semiquantitative"). These techniques are thus very reliable, provided the fungal strain to be detected is known beforehand, and so are used as referential methods. With such methods, a grain is deemed of suitable microbiological quality if less than 10 000 germs of the storage flora per gram of grain are detected.

New approaches are based on detecting constituents and fungal metabolites. Such approaches exploit the fact that molds have specific characteristics that distinguish them from other eukaryotes. These characteristics include the regulation of certain enzymes, the synthesis of lysine amino acid by a particular metabolic route, extremely structural characteristics (e.g., the Golgi apparatus), and genetic characteristics (e.g., haploid). From among these attributes, two types of compounds can be used as indicators of a fungal contamination.

The secreted compounds are synthesized compounds such as soluble carbohydrates (e.g., disaccharide trehalose and polyhydric alcohols such as mannitol or arabitol) or products of the metabolization of complex carbons such as volatile aldehydes, alcohols, ketones, spores, primary metabolites, secondary metabolites (i.e., volatile compounds). The last item gives rise to the characteristic fungal odor and is often detected by an electronic nose. For nonvolatile compounds, other tools such as infrared spectroscopy seem better suited.

The structural compounds of mold can also be used for their detection. The main polysaccharides of the cell wall of mold are the α et β (1-3) glucans, as well as chitin. Ergosterol is a component of fungal cell membranes.

Chitin may absorb infrared light, making it useful for infrared spectroscopy [Nilsson, et al., 1994; Roberts et al., 1991]. The main inconvenience in using this component as an indicator of fungal contamination is that chitin is not limited to fungi; it is found in insects, diatoms, arachnids, nematodes, crustaceans, and several other living organisms [Muzzarelli, 1977]. In addition, it may take different forms, each of which requires a specific detection technique. Roberts et al. [Roberts, et al., 1991] estimates the quantity of mold on barley by detecting this molecule but also detects glucans by near-infrared spectroscopy.

must be less than a given threshold; the limit for corn is 8 µg/g.

Ergosterol, however, is more specific to mold. This molecule, which may still be called provitamin D2, is a C24-methylated sterol (and is part of the subgroup of organic compounds that are soluble in lipids) and is found in the cell membranes of yeasts and filamentous fungi. This molecule is not found in animal cells [Verscheure et al., 2002] and is in the minority among the sterols found in higher plants [Pitt et al., 1997] and insects [Weete, 1980]. Griffiths et al. [Griffiths et al., 2003] demonstrated that ergosterol is the primary sterol found in molds: it represents 95% of the total sterols, with the remaining 5% being ergosterol precursors from *Leptosphaeria maculans*. This specificity makes this molecule a potential tracer of fungal activity. It is generally agreed that the ergosterol content of grains
