1Queensland Department of Primary Industries & Fisheries, Agency for Food and Fibre Sciences, Farming Systems, PO Box 2282, Toowoomba, Qld 4350. www.dpi.qld.gov.au Email email@example.com firstname.lastname@example.org email@example.com
2GxE Crop Research, Esperance, WA 6450 Email firstname.lastname@example.org
Grain colour defects, including staining and black point, have been a problem for Australian barley growers for a number of years. This has resulted in thousands of tonnes of malting barley being downgraded each year. Over the last decade, research has been conducted into many aspects of these disorders, including objective assessment, biochemical evaluation, crop management, storage effects and resistance breeding. A number of key findings in the area of barley staining, including sources of improved resistance and crop management, will be presented.
Future Australia barley varieties will have increased levels of resistance to grain colour defects, as recent screening techniques have identified many resistant breeding lines.
Barley, Black Point, Staining
Thousands of years ago barley “production” was only in dry Mediterranean climates. Human migration has since seen barley production expanded into more semi-tropical and tropical regions. With these changes in growing environments, barley has been required to deal with a number of new environmental stresses, most of which are disease threats. One significant change has been the move to climates when rain is more prevalent during the crop harvest period than in the growing season. Wet, humid conditions during harvest induce a number of problems. Rain during grain harvest can cause pre-harvest sprouting which results in a loss of quality. However, where high levels of humidity or even low levels of pre-harvest rain occur, grain can take on a stained appearance. Consequently, the environmental conditions prior to harvest has a direct impact of the appearance of barley (Edney et al. 1998).
Barley grain colour defects comprise of three distinct components. The first is stained grain caused by fungal proliferation after preharvest rain. In Australia, both saprophytic and pathogenic fungal species have been identified on barley grain. Recently Young and Loughman (2001) typed the fungal strains on Western Australian barley after it was subjected to pre-harvest rain. The second is a yellow pigmentation usually resulting from light rain or high humidity just prior to harvest. The third is black point, brought about through humid conditions between anthesis and harvest and appears as a brown/black discolouration on the germ.
The first two components can be measured using a colourimeter or spectrophotometer where the staining has an impact on the brightness of the grain. In comparison, the pigmentation is yellow appearance. Black point is measured visually and expressed as percentage.
The Australia malting barley classification and handling sector has been disjointed in its approach to a colour standard. Differences existed between grain handling companies in the classification of grain brightness. More, noteworthy is that brightness has now been removed as a specification from the national grain receival standards for malting barley. However, an objective standard remains in that hectolitre weight (HLW) has been considered as an indirect measure of possible pre-harvest sprouting. Hence, a decrease in HLW implies an increase in grain volume due to the grain imbibing water prior to harvest. With any pre-harvest sprouting condition, doubt exists in the buyers mind, over the quality of that grain. More work is required in this area to provide a more robust relationship between changes in HLW, pre-harvest sprouting and impacts on quality.
While the effect of staining can be related to fungal infection of the seed coat, the biochemistry behind the yellowing remains uncertain. Reuss (2001) suggested that an increase in the yellow appearance could be as a result of increases in non-enzymic browning reactions. In contrast, Fox et al. (2001b) suggested that increases in the yellow appearance could be associated with an increasing level of phenolic pigments in the outer layers of the grain. These contrasting results may be related to the fraction of grain analysed. Fox et al. (2001a) pearled the grain and analysed only the outer layers, whereas Reuss (2001) used a ground grain fraction. This area requires further research to resolve these conflicting results.
As described above, black point and a brown/black produces discolouration over the germ. Limited biochemical studies have been conducted to unravel the biochemistry of this defect. Williamson (1997) first proposed that peroxidase enzymes may be involved in black point formation in wheat. Sulman et al (2001) suggested that while this may also be the case for barley, there was no clear relationship between the level of black point and peroxidase. Further studies by Hadaway et al (2003) indicated a possible correlation between black point susceptibility and peroxidase banding on electrophoretic gels. In focusing attention away from the possible enzymes involved in catalysing black point formation, Michalowitz et al (2002) reported that after extraction of the “:black point”, an increase in the amounts of phenolic acids, namely feurlic and coumeric acids, was found in both barley and wheat, compared to extracts from non-black point grain. This suggests that the black pigment could be a large polyphenolic compound. Additional information regarding the role of phenolics was provided by Sulman et al (2003) who reported that black point barley germinates at a slower rate than non-black point. This suggests a possible role of phenolic acids in this temporary “dormancy-like” condition. Further research is needed to resolve this complex biochemistry.
Kernel staining or discolouration present a number of problems for grain growers, grain handlers and processors. Abramson et al. (1983; 1998) showed the negative effects of higher moisture contents and fungal contamination on stored barley. Hence the importance of an objective method to evaluate kernel discolouration to eliminate the potential problems of storing grain with high levels of fungal contamination. Near Infrared spectroscopy (NIR) has been shown to be a reliable objective method (Roberts et al. 1991; Fox et al. 2001b). It has an added benefit as other grain constituents such as protein and moisture could also be predicted simultaneously. Objective detection of staining has also included image analysis. Luo et al. (1999) used image analysis to determine discolouration in barley and a number of other grain types. Another aspect of objective assessment has been considered by others in this area. Young and Loughman (2001) and Lamper et al. (2000) have compared a level of discolouration and ergosterol in barley and wheat, respectively. The results showed a positive relation between the levels of discolouration and ergosterol. This suggested that measuring ergosterol may be a more quantitative method of detecting the level of fungal staining.
While objective measurement of discolouration has been important in terms of grain handling and marketing, another important aspect would be knowing what fungi was present on the grain. An alternative approach would be to measure a specific biochemical component that could indicate the strains of fungus present. Pekkarinen et al. (2000) demonstrated that by measuring proteases, it was possible to distinguish between Fusarium strains. Danks et al. (2001) reported a new assay for the rapid detection of fungal presence by the level of mycotoxins.
The increasing demand on high quality grain for human and animal consumption has seen changes in farming practices where winter and summer crops are growing in succession. Additional changes in farming practice have seen crop stubble not being completely removed or incorporated into the soil. Hence, disease threats exist with the potential for carryover on stubble. In terms of kernel discolouration in regions where maize (Zein) is grown as a summer crop, a potential exists for pathogenic fungi to infect subsequent winter crops. One specific fungus, Fusarium graminearum, has caused significant damage to barley and wheat crops in North America (de la Pena et al. 1999). In particular, a toxin produced by the fungus, deoxynivalenol (DON), has been reported to cause illness in animals fed on infected grain and also gushing in beer produced from infected barley.
While objective methods for detecting discolouration or the fungus responsible for causing discolouration could assist in managing infected grain, other options include on farm management. Agronomic studies (Young 1997; 1999) indicated that a number of factors including sowing date, heading date and harvest technique impacted on the level of kernel discolouration. Environmental conditions have a strong impact on discolouration with results suggesting that quicker maturing varieties left in the field for too long had higher levels of discolouration than slower maturing varieties. While it may be possible to reduce the level of kernel discolouration through agronomic practices, the best option is development of resistant varieties. While strategies for evaluating and selecting resistance to kernel discolouration have varied, a number of studies have shown that it was possible to breed some ‘resistance’ to kernel discolouration. Brinkman and Luk (1979) had shown that the angle of the head, with heads at < 90o nodding angle having lower levels of discolouration. Miles et al. (1987) demonstrated that while the environment had a significant effect on kernel discolouration, there were genetic differences in incidence between breeding lines and commercial varieties.
In terms of opportunities for improved varieties, Gebhardt et al. (1992) showed that breeding for resistance was possible with the six-rowed variety, Chevron, as the resistant parent. Goblirsh et al. (1996) presented data suggesting an association in the inheritance between low diastatic power and kernel discolouration resistance in a breeding population. Young (1997) also reported a number of breeding lines that exhibited a high level of resistance to staining. All of these reports would suggest that resistance to kernel discolouration was a heritable trait and therefore it should be possible to breed for resistance. De la Pena et al. (1999) first reported genetic markers for resistance to kernel discolouration. A number of markers on all but one chromosome were identified. Importantly, two markers on chromosomes 5H and 1H were associated with resistance to discolouration and maturity.
In addition, more recent studies have identified markers for black point. Markers have been identified on chromosomes 2H (Amanda Able pers comm.) and 5H (Cheng Dao Li pers comm). Sulman et al. (2003) also reported levels of heritability from 39 to 70+% in a number of populations suggesting that development of resistance to this grain defect was readily achievable through selection.
While it would be considered ideal to develop varieties resistant to colour defects, it is important to note that fungal infection and/or pigmentation is an indication of environmental events just prior to harvest. These may impact either in the short- or long term on the quality of that crop. Industry must remain pro-active in developing technologies that will provide objective assessment of potential performance at delivery to avoid long-term negative attitudes to a variety based on defects not specifically related to a variety.
A number of issues remain to be fully addressed in barley grain colour and these will require a whole of industry approach. Some of those issues include:
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