A genetic approach to improving post-harvest quality

Lead Research Organisation: Harper Adams University
Department Name: Crops and Environment Sciences


Minimal processing adds significant value to fresh produce, however, it also increases its perishability reducing shelf life and leading to waste of the produce and the resources used to grow it. This project is aimed at post harvest discolouration, a significant cause of quality loss in a wide range of fresh produce such as sliced apple, cut cabbage and lettuce. The main issue we are addressing is postharvest discolouration of lettuce in salad packs. UK lettuce production/imports are worth £240m farm gate but the retail value of UK processed salads is £800m. However, Tesco have recently reported that 68% of their salads are thrown away; the situation is similar for other retailers. There is therefore a need to improve postharvest quality to reduce waste and deliver consistently good quality products to consumers. Modified atmosphere packaging can provide control but once the pack is opened oxygen enters resulting in discolouration. Growing conditions also influence postharvest discolouration but are difficult to control in field crops. We are proposing breeding lettuce varieties with reduced propensity to discolour as a way to address the problem. To do this we need to understand the genetics and biochemistry of discolouration.
We are building on previous research we have done which identified genetic factors controlling the amount of pinking and/or browning that developed on lettuce leaves in salad packs 3 days after processing. However, we do not know what compounds or which genes are involved and we now intend to find this out by a multidisciplinary project involving three universities; Harper Adams University, Reading and Warwick, a lettuce breeding company, a lettuce grower, a salads processor and the Horticulture Development Company.
We have produced a set of experimental lettuce lines which we know show differences in the amount of pink or brown discolouration they produce. We will grow and process these lettuces in a way that mimics commercial production. We will then assess the salad packs for the amount of discolouration developing over 3 days, which is the current best before date for supermarket salads. We can then link this information to the plant's DNA profile to identify genetic factors for discolouration and DNA markers which can be used by plant breeders.
The same lettuces will also be analysed for compounds produced by a biochemical pathway called the phenylpropanoid pathway. This is thought to produce the pigments that cause discolouration. We know from other studies in a plant called Arabidopsis the genes which control the phenylpropanoid pathway and we have found the same genes in lettuce. We will see how these genes behave in lettuce plants that produce a lot of discolouration and ones that don't discolour. We will also see how the genes behave under different growing conditions. We can link these gene expression patterns to the amount of pinking and browning to see which genes are the key ones. Once we have done this we can look for naturally occurring versions of the genes which give a reduced discolouration.
The compounds produced by the phenylpropanoid pathway influence other things such as pest and disease resistance, taste etc. We do not want to reduce the amount of discolouration by breeding but end up with lettuce susceptible to pests or with poor taste, so we will assess lines which show high discolouration or no discolouration for their resistance to aphids and mildew and for taste to see if there are any differences. There are some compounds produced by the pathway which are colourless but still provide some resistance so by knowing the genetics and biochemistry breeders will be able to carry out smart breeding.
We will see if the results for lettuce hold true for other crops by seeing how the key genes behave in apple and cabbage and whether this is related to the amount of browning that develops when they are processed and look for genetic differences in these crops

Technical Summary

The research aims to provide tools for breeding crop varieties with reduced propensity for postharvest discolouration. The crop system for study is postharvest discolouration in lettuce which is a major problem for the UK salad supply chain. A reported 97,000 tonnes of whole head lettuce and bagged salad are wasted annually in the UK at a cost of £234m. A major cause of this is post harvest loss of quality due to discolouration (pinking and browning). We will use a multidisciplinary approach involving phenotyping, QTL analysis, bioinformatics, genomics, transcrptomics and metabolomics to achieve the following objectives.
1. Increase understanding of the genetics of pinking and browning in lettuce
2. Determine the role of phenylpropanoid (PP) pathway in lettuce discolouration.
3. Determine whether non PP pathway genes have a role in lettuce discolouration
4. Test the robustness of a genetic approach to reducing discolouration
5. Identify potential sources of beneficial alleles for key genes.
6. Assess the potential impact on pest and disease resistance and taste.
7. Test the applicability of the findings from lettuce to cabbage and apple.

Industry partners from the supply chain will provide expertise in breeding, crop agronomy and production and processing and consumer acceptance.
We will use an improved lettuce linkage map and an F7 RIL population to carryout QTL and eQTL analysis in order to understand the role of PP pathway genes in determining phenotypic variation. We will use RNA seq combined with bulk segregant analysis and also exploit lettuce sequence data to attempt to identify new genes associated with discolouration. A lettuce diversity set will be mined to identify beneficial alleles for use in smart breeding aimed at improving the discolouration phenotype while minimising the impact on other agronomically important traits. We will carry out an initial assessment of the applicability of our results to other crops (apple and cabbage)

Planned Impact

The ultimate beneficiaries of this research will be consumers who rely on visual appearance to judge quality of fresh produce. Discolouration is one of the major factors limiting current designated shelf life of fresh produce and one of the most common reasons for product failing to meet shelf life targets making it a major cause of food waste post-purchase. The minimum requirement for shelf life of pre-packed lettuce to the consumer is currently five to six days however, salad packs generally only survive three days from time of purchase until 'end of life' and it is estimated that 97,000 tonnes of whole head lettuce and bagged salad are wasted in the UK at a cost of £234m. The main cause of this wastage was loss of quality in a relatively short time (i.e. short shelf life). There is therefore a high demand from consumers for products with longer shelf life.
The research will have wider societal benefits by reducing food wastage and resultant reduction in wastage of inputs such as fertilizer, water and pesticides as well as leading to improved land use through improved marketable yield.
Other stakeholders in the lettuce supply chain will also benefit from the research as it will lead to improved postharvest shelf life of fresh produce which will reduce waste, reduce costs and delivery of consistently good quality products.
Lettuce breeders will benefit directly from the research as it will provide underpinning knowledge and molecular breeding tools to facilitate breeding of lettuce varieties with reduced propensity to develop post harvest discolouration. Rijk Zwaan as a partner in this project will exploit the results directly in their lettuce breeding programme and expect to have developed lettuce varieties with improved phenotype for post harvest discolouration 5-10 years from completion of the project.
Processors and retailers of fresh produce will benefit from the research because it will lead to development of improved raw materials for their products. Currently modified atmospheric packaging (MAP) can be used to control discolouration and prolong shelf life. However, this technique has a number of disadvantages and limitations. These include: added production costs; the need for specialised equipment and significant imprecision. The huge natural variation in raw materials makes it almost impossible to optimise gas formulations for each product and complex mixed leaf packs inevitably create compromises. More importantly, once a pack is opened, oxygen re-enters resulting in development of discolouration after purchase, the consumer therefore gains little from MAP. More aggressive MAP approaches for fresh produce (e.g. packaging in targeted anaerobic conditions) are not widely adopted because of the potential impact on food safety - a result of an increased risk of growth of anaerobic human pathogens. In addition, MAP is not used with whole heads either for retail or raw material for a processing factory (where they may be held in an intake store for 7-10 days). Breeding of new varieties with improved shelf life will remove or reduce the need for MAP reducing production costs and improving product quality throughout the supply chain. The value of the project to the supply chain is illustrated by the involvement of Bakkavor (the largest processor of fresh produce in the UK ) in the project
Lettuce growers will benefit from the research. Growing conditions have been shown to influence postharvest discolouration particularly the timing of water availability; however, this is difficult to control in UK field crops. Breeding crop varieties with an inherent reduced propensity to discolour regardless of the growing environment is a sustainable and cost effective way to address the problem as growing such varieties has no added production costs and will reduce costs due to wastage. The value of the research to lettuce growers is illustrated by the involvement of Gs Fresh (the largest UK salad growers) in the project.


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Description This research has provided molecular breeding tools to facilitate breeding of lettuce varieties, with reduced propensity to develop post-harvest discolouration, and underpinning knowledge of biochemistry enabling more efficient and effective use of both existing and new markers.
• We have defined QTL for two distinct discolouration events (pinking and browning) and shown that they are under distinct genetic control, although the biochemical processes involved may be related.
• We have improved the linkage map of lettuce cross Saladin x Iceberg with the addition of over 450 new markers which allows it to be anchored to the UC Davis lettuce genome. This provides the ability to identify gene candidates underlying any regions of interest identified in QTL analyses.
• Correlation analyses have identified a number of potential biochemical components of the pathways, some unique to each symptom, some common to both. QTL for differentially expressed genes associated with the development of the two different symptoms and QTL for biochemical components of the PPO pathway have identified at least one novel association between differentially expressed genes and the browning phenotype. Using these resources (improved marker density linkage map, flanking markers for QTL and an anchored underlying genomic sequence), breeders can identify additional targets genes and biochemical components in breeding programs. In addition, selection of lines showing high and low levels of pinking and / or browning could be incorporated directly into back-cross breeding programs.
• Collection of environmental data during the field trials has allowed further investigation into the impact of the scale and timing of environmental variability on the biochemical and genetic processes involved in subsequent discolouration development.
• The work has also led to the exploration of new techniques for examining the spatial distribution of particular compounds in tissues using Desorption Electrospray Ionisation (DESI) mass spectrometry.
Exploitation Route The bulk of the findings from the work are of importance to crop breeders. The new understanding that pinking and browning have been presented and discussed with leafy crop producers, processors and breeders, the key conduits for taking the work forward. It is anticipated that the knowledge that pinking and browning are related but distinct processes will enable breeders to phenotype for these two processes. The identification of a number of genes that show differential expression in high and low discolouring extremes from the population studied also identifies an alternative approach for breeders to pursue. Breeders will need to incorporate this information into their breeding programmes before the growers and processors are able to utilise the genetics components of the work.The work studying the effect sof environment have been incorporated into growing algorithms at the collaborating growing business. This demonstrates a clear impact and early from the work.
Sectors Agriculture, Food and Drink

URL https://www.harper-adams.ac.uk/research/fprc/
Description G's Fresh have used data on environmental factors from the project and incorporated this into their IceCAM predictive model when looking at internal data sources. They have identified increased pinking on lettuce ribs in relation to minimum temperature, total rainfall and maximum radiation during crop growth. This has allowed them to schedule harvest dates to minimise crop waste.
First Year Of Impact 2018
Sector Agriculture, Food and Drink
Impact Types Economic