Crocus, saffron-omics and the highest value crop

Saffron, Crocus sativus and origin label

Saffron, Crocus sativus and a protected origin label

Saffron, the stigma of Crocus sativus, is the highest priced agricultural product (often €/$25 or £15 per gram) and a good example of a profitable crop with sustainability, cultural and social values, and high labour demand. I have been discussing –omics studies of the crop – the DNA, RNA, metabolites and secondary products – at the annual meeting of a European Science Foundation COST programme Saffronomics.

www.Saffronomics.org logo

www.Saffronomics.org logo

The ‘Action’ aims to coordinate research on Saffron-omics for crop improvement, traceability of the product, determination of authenticity, adulteration and origin to provide new insights that will lead a sound Saffron Bio-Economy. Despite the high price, the spice costs only a few pence/cents per portion, and adds enormously to the flavour and colour of many dishes. Biologically, saffron is the species Crocus sativus, as recognized by Linnaeus, and it is a sterile triploid with 2n=3x=24 chromosomes.

Audience for annual meeting

Audience for annual meeting

The programme of our Annual Meeting opened with the genomics sessions – the DNA, RNA, genetics and epigenetics. I don’t usually start reviews with, nor indeed include, my own talk, but here its content sets the scene for other work discussed at the meeting. I talked about the work of Nauf Alsayid, who shows the lack of any clear DNA differences between any accessions of saffron – whether from Kashmir, Greece, Italy, Spain, Holland or Iran. I cited a paper from 1900, itself reporting work back to 1844, where the French botanist Monsieur Paul Chappellier reported “for the Saffron, there is only known a single and unique species; for ages it has not produced a single variety”, writing that he was importing bulbs Naples, Athens, Austria, Spain, Cashmere and China (Chappellier P 1900. Creation of an improved variety of Crocus sativus. J. Royal Horticultural Society XXIV Hybrid Conference Report 275-277 – brilliant download, even available free for Kindle!). Plus ça change, plus c’est la même chose!

Highest quality Saffron from Thiercelin 1809

Highest quality Saffron from Thiercelin 1809

After my talk, Jean Marie Thiercelin, the seventh generation of the major saffron and spice company http://www.thiercelin1809.com told me that his grandfather knew Paul Chappellier, and he commented in the history of saffron production in France: Chappellier knew how to produce 10 to 15kg per ha before the First World War. After the war, saffron production stopped altogether in France, but it has restarted this century, with now some 137 growers on 37 ha but production of only some 5kg per ha.

Continuing with the talks, a DNA-sequence level study of saffron by Gerhardt Menzel with Thomas Schmidt (Dresden) analysed of several Gigabases of genomic survey sequence data, revealing about ten distinct tandemly repeated satellite DNA sequences that could be used to identify chromosomes in saffron by in situ hybridization. The species has a 78% repeat content in the DNA, with about 6% being the rDNA, and many different classes of transposons.

Giovanni Giuliano - High trhougput sequencing of saffron RNA and gene discovery

Giovanni Giuliano – High throughput sequencing of saffron RNA and gene discovery

Giovanni Giliano (with Sarah Frusciante, Italy) demonstrated the carotenoid cleavage dioxygenase from saffron stigmas catlayses the first step in saffron crocin biosynthesis, a clear example of the pathway to the critical secondary product giving saffron its value (http://www.pnas.org/content/111/33/12246.short).


Slivia Fluch - Saffronomics Genomics Working Group Leader

Slivia Fluch – Saffronomics Genomics Working Group Leader

Both Matteo Busconi and Silvia Fluch (Austria) discussed epigenetic differences detected from different saffron collections: important for both understanding the controls on gene expression and for determining the origin of samples. Each producing area seems to have distinct profiles. Caterina Villa (Porto) reported results from use of the plant ‘barcoding’ primers ITS and matK with high resolution DNA melting analysis for saffron authentication, and more detail about the chloroplast genomes was presented from Bahattin Tanyolac and his Turkish colleagues. Although wild species of crocus are of interest from several points of view, only one paper, from Joze Bavcon (Slovenia) discussed these in detail, with a report of the natural hybrid Crocus reticulatus x C. vernus.

Joze Bavcon Crocus of Slovenia Book Cover

Joze Bavcon Crocus of Slovenia Book Cover

The next group of talks discussed the saffron metabolome, the analysis of different constituents of Crocus. Crocus is one of the few species to have its own international standard (ISO3632: http://j.mp/isosaffron ), and both quality and purity are measured (including contamination with stamens and pollen, along with detection of adulteration. Several participants were involved in the formulation of the standard, and Gianluca Paredi reported improvements that need less than the ISO methods needing no less than 23g of stigmas! Natural colours from plants such as Buddliea, Calendula, Curcum, Gardenia, safflower (Carthamus Asteraceae), cochineal (from the insect) and turmeric are widely mixed with saffron.

Chair of the Saffronomics Action Professor Maria Tsimidou

Chair of the Saffronomics Action Professor Maria Tsimidou

The Saffronomics project leader, Maria Tsimidou (Greece), used the three ISO3632 peaks for saffron – colouring strength from crocins absorbing at a peak wavelength of 440nm, aroma from safranal at 330nm, and taste (flavour) from picrocrocin at 257 nm – for examination of quality and authenticity of commercial saffron samples. Of 16 samples, 3 were adulterated, and half of the pure samples were graded in ‘category I’. Another amazing figure quoted was the price of saffron in quantity: of 75 tonnes imported to one county, only 35% is priced at more than $500 per kg. Authentic saffron could not be produced for anywhere approaching $1000/kg (typically $10-$15000/kg), so all this bulk product is fraudulent. Technology sessions in the meeting covered alternative quantification approaches to spectroscopy: Laura Ruth Cagliani in Milan tested  different solvents for extraction for NMR-based metabolomic characterization of authentic saffron distributed within the COST partners as well as the NMR evidence of absence of plant adulteration in those saffron samples.

Moschos Polissiou Saffronomics

Moschos Polissiou Saffronomics

A leading group from Thessaloniki was able to detect adulteration with as little as 15% cochineal. EA Petrakis and Moschos Polissiou demonstrated how FT-IR spectroscopy is promising to quantify small amounts of adulterants in saffron – safflower, Gardenia and tumeric – where diffuse reflectance mode provides rapidity, ease of use and minimal sample preparation. Other important reports discussed aging effects on profile of secondary metabolites (Paraskevi Karastamati Greece) and detection of herbicide residues (Christina Mitsi).

H stable Isotope Map from http://www.earthmagazine.org/article/cold-case-files-forging-forensic-isoscapes

H stable Isotope Map from http://www.earthmagazine.org/article/cold-case-files-forging-forensic-isoscapes

Micha Horacek (Austria) presented new results looking at the ratios of stable isotopes in saffron, a technique increasingly used to determine the origin of all agricultural produce. He showed the impressive map of with the gradient of water (hydrogen and oxygen) isotope ratio from North to South and from East to West in Europe. He also showed the differences in nitrogen stable isotope ratios depending of fertilizer use, and sulphur which depends on the underlying geology. Current work with saffron shows considerable year-to-year variation in the position of accessions from different regions of Europe, but the data is still being collected. Soon Micha will be getting a sample of our own, Leicester-lab-produced, saffron to add to his map!

Our hosts at RIKILT, the Food Safety and Quality Institute, Wageningen University, have much advanced applied science on food quality. An eye-opening talk by John van Duynhoven told us about rehydration of freeze dried blanched carrot with dynamic assessment of water movement in samples with and without blanching, freeze drying at -28 and -150C. Another series of images showed water transport and the impact of pre-cooking of rice, using magnetic resonance imaging MRI as a functional measurement of rice cooking. The final section discussed why crackers don’t crack: vapour transport during shelf life of crackers! Modelling of the nature of water transport links processing & formulation to the structure and on to functional and storage implications.


Fran Azafran - a school book about saffron

Fran Azafran – a school book about saffron

For ESF – COST projects, dissemination and public understanding are important, and participants were treated to a preview of a series of six school books about Fran Azafran and Franny Azafran by Manuel Delgado from Cuenca, Spain. I look forward to seeing these in full, and hopefully to their availability in other languages too.


At the podium

At the podium

Like the best of the projects, I feel that saffron science has moved in the last decade, (including research in the consortia www.crocusbank.org and www.saffronomics.org) with notable fundamental, technical and applied outcomes of our research. We know about its relatives and genome structure, key genes, metabolic processes and the key secondary products, and even understand epigenetic control, corm growth and dormancy. After 4000 years of being sold fake saffron, the fraudsters know now that we can test for saffron purity and quality!

Marta Rodlan (Vice Chair of the Action), Jose Antonio Fernández Perez and Jean Marie Thiercelin: key people in saffronomics

Marta Rodlan (Vice Chair of the Action), Jose Antonio Fernández Perez and Jean Marie Thiercelin: key people in saffronomics

Saffronomics Meeting Book Cover

Saffronomics Meeting Book Cover

Selection on a CONSTANS-like gene between two red oaks

Selection on a CONSTANS-like gene between two red oaks

Selection on a CONSTANS-like gene between two red oaks

Oaks can provide a model to study hybridization as they often maintain species integrity and distinct environmental adaptations despite recurrent gene flow. Lind-Riehl et al. identify candidate genes potentially involved in local adaptation in the interfertile species Quercus rubra and Q. ellipsoidalis, which are characterized by contrasting adaptations to drought. They find that a CONSTANS-like (COL) gene is nearly fixed on alternative alleles in both species, as reflected by high FST values (55–80 %) between neighbouring Q. rubra and Q. ellipsoidalis populations. All other markers show low interspecific differentiation (approx. 5 %). They conclude that this COL gene may play a role in adaptive divergence and reproductive isolation (via flowering time), making it a promising candidate speciation gene.

New plant journal

Image: Wikimedia Commons.

Image: Wikimedia Commons.

It just had to happen, but we didn’t know it would take nearly 150 years to come to fruition. And fruition is an apt word because the creation of a new botanical journal has recently been announced by the publishers behind Nature, the world’s premier general science journal. Imaginatively entitled Nature Plants, this new organ is due to be officially published in January 2015 but already has interweb presence with a blog and can be ‘followed’ on such social media as Facebook and Twitter. Its aim is to provide a fully rounded picture of the most accomplished and significant advances in the plant sciences, and will cover ‘all aspects of plants be it their evolution, development or metabolism, their interactions with the environment, or their societal significance’. Furthermore, along with original research, Nature Plants will also deliver ‘Commentaries, Reviews, News and Views’ from across the full range of disciplines concerned with the plant sciences (i.e. a bit like the Annals of Botany…). However, with topics covered in the journal including (deep breath) ‘agronomy, genomics, biochemistry, metabolism, biofuels, metabolomics, biophysics, molecular biology, cell biology, photosynthesis, defence physiology, development, plant–microbe interactions, disease resistance, proteomics ecology, secondary metabolism, economics, sociology, evolution, symbiosis, food security, systems biology, forestry and water use’, I do hope they leave something for other – more established – botanical journals, such as the Annals of Botany!

[Have others heard that the original Nature – in keeping with its soon-to-be somewhat impoverished science coverage – is being retitled Nature Cosmology, Palaentology and Non-botany? Whilst we wish this new venture well, it will be interesting to see if anybody publishes in the new journal because, and despite the undoubted cachet and kudos associated with the word Nature in the article’s citation, it won’t have an Impact Factor (IF) for a few years. Now, who wants to risk having publications on their CV in journals with no IF with potential damage to promotion prospects and career advancement (not that IFs should be used for such purposes – see e.g. EASE statement on inappropriate use of Impact Factors? Just saying. – Ed.]

Ongoing gene flow among Australian alpine Poa

Ongoing gene flow among Australian alpine Poa

Ongoing gene flow among Australian alpine Poa

Recently radiated groups pose a taxonomic challenge even with extensive molecular data, as they may be genetically differentiated only at small and/or patchy regions of the genome. Griffin and Hoffmann investigate species’ structure among the Poa that dominate the Australian alpine zone, which have radiated in the last 0.5–1.2 million years. Using a Bayesian approach to co-estimate nuclear and chloroplast gene trees with an overall dated tree, they find that most species are not genetically distinct, despite distinguishable phenotypes, which suggests recent adaptive divergence with ongoing inter-taxon gene flow.

Genetic and morphological signature of ancient Agave cultivation

Genetic and morphological signature of ancient Agave cultivation

Genetic and morphological signature of ancient Agave cultivation

Several species of Agave were cultivated prehistorically for food and fibre in Arizona, USA, and relict populations that remain in the landscape today can provide an insight into ancient cultivation practices. Parker et al. test whether formerly cultivated Agave parryi populations still bear a predictable genetic and morphological signature by comparing populations of uncertain origin with known wild and cultivated populations. They find less genetic and morphological variation and stronger population differentiation in populations actively cultivated in the past than in wild counterparts, and conclude that where archaeological information is lacking, the genetic signature of Agave populations can thus be used to infer their evolutionary history and to identify fruitful sites for archaeological investigation of pre-Columbian cultivation practices. The same approach can clearly be adopted for other species in similar situations.

Adaptation for rodent pollination in Leucospermum arenarium

Adaptation for rodent pollination in Leucospermum arenarium

Adaptation for rodent pollination in Leucospermum arenarium

Numerous studies have documented flower feeding in angiosperms and adaptions for non-flying mammal pollination, but the underlying interactions and structures that facilitate this pollination mode are less well known. Johnson and Pauw investigate the pincushion Leucospermum arenarium (Proteaceae) and find that the nectar is unusually viscous and, although produced deep inside the perianth, is translocated via capillary ducts to the petal tips where rodents can access it without destroying the flowers. Although flowers are presented at ground level, when raised experimentally to higher positions seed production is not reduced, indicating that selection through female function does not drive the evolution of geoflory. This reliance on rodent pollination has apparently evolved despite the fact that they are very adept at removing pollen through grooming.

The Martian by Andy Weir

TheMartianA while back I asked for recommendations for science fiction reading involving botany. On our Facebook page Claire Soulsby suggested The Martian by Andy Weir. I’ve had a stroke of luck* and have been stuck in bed feeling sorry for myself. This has given me plenty of time to read it.

The story begins after a freak dust storm on Mars causes the Ares 3 team to abandon their mission, leaving behind their dead crew member, Mark Watney. Watney awakes to find himself alone with the habitat and no hope of rescue for years, and enough food for months. The challenge is to produce food, water and oxygen to keep him alive and then to establish contact with NASA to coördinate a rescue.

The key to survival is first botany. It’s Watney’s ability to grow food on Mars that keeps him alive long enough to get a fighting chance. As everybody knows Botanists are pretty much the closest thing we have to superhero geniuses, so Watney is able to engineer all the fixes he needs to make in the hab to make farming, and everything else he needs, happen.

The book reads like hard-SF for the most part. The science is plausible and by relying on near-future science it means that Weir puts his character in a believable danger. The start of the writing process was planning a hypothetical Mars mission, including contingency plans for what might go wrong. Then he realised the contingency plans would make the basis of a story.

Most of the story is told through log entries. This works to explain the problems and the solutions. It also gives a plausible reason for why the character comes across the way he does. I vaguely remember someone saying there are no characters in Shakespeare plays, just plot devices. In a similar way, I’m not sure there are many characters in this book. In a couple of other reviews, people think the characterisation is weak. Watney does things, but he’s not changed much by them. When the next problem comes along in the book, he simply settles down to solve it so while there are many problems, I don’t know if there are many setbacks or catastrophes. Watney’s job at times seems to be to set up the next problem.

Fortunately, the problems are interesting enough to pull the story along. It’s also a change to read something where not every scientific problem can be solved by basic physics.

There’s a video of a talk he gave at, including a reading of the first chapter.

*Not good luck. I tend to avoid that.

Other reviews

Temperature and reproductive success in arabidopsis

Temperature and reproductive success in arabidopsis

Temperature and reproductive success in arabidopsis

Seed yield and dormancy status are key components of species fitness that are influenced by the maternal environment. Huang et al. grow the Arabidopsis thaliana ecotype Burren (Bur), which is adapted to a cool, damp climate, under conditions normally experienced by the Cape Verdi Isle (Cvi) ecotype, which is adapted to a hot, dry climate. They find that viability of pollen is unaffected, but limited filament extension relative to that of the pistils results in failure to pollinate. Both seed yield and dormancy are reduced, suggesting that higher temperatures predicted in climate change scenarios will impact on the seed performance of the cool-adapted ecotype.

Something fishy in the veggie patch…

Image: Nada Meeks, www.fineartamerica.com.

Image: Nada Meeks, www.fineartamerica.com.

The ‘alpha’ category is widely regarded as the best of its kind; think of alpha (males) in the context of animal behaviour and Aldous Huxley’s Brave New World, an α+ grade on your exams or the sports cars from Alfa Romeo. But omega – right at the other end of the Greek alphabet – is also merit-worthy, especially when it’s omega fatty acids (FAs), which are polyunsaturated FAs needed for human metabolism. However, since they cannot be made de novo by the human body – and are therefore considered ‘essential’ – it is necessary to acquire them in the diet.

Two of the three essential omega FAs needed for human metabolism – Eicosapentaenoic acid (EPA) and Docosahexaenoic acid (DHA) – are derived from marine sources, such as fish. The third – alpha-Linolenic acid (ALA – which, despite its name, is still an omega fatty acid!) comes from plant products and is used in the body to produce EPA, which in turn is used to generate DHA. One way of getting your essential omegas is to consume milk produced by cows that have grazed on fresh grass/red clover, whose milk has been shown to increase in ALA as a result. But if you are milk-averse or lactose intolerant this won’t work for you. Another dietary strategy is to eat fish. However, with concerns about dwindling fish stocks, and recognising that fish themselves actually get their omegas from the algae that they have ingested, a more imaginative – and plant-based – avenue is being promoted.

Using a GMO (genetically modified organism) Noemi Ruiz-Lopez et al. have successfully demonstrated high-level accumulation of fish-oil omega-3 long-chain polyunsaturated fatty acids in a transgenic (which includes at least one gene from an alga…) oilseed crop plant. Using heterologous genes (i.e. genes from organisms different to the host crop species) the Rothamsted Research (Harpenden, UK) -based team have developed Camelina sativa (like arabidopsis, a member of the Brassicaceae) whose seeds accumulate up to 12 % EPA and 14 % DHA (which levels are equivalent to those in fish oils). On the back of expectations that this could represent a sustainable, terrestrial source of these fatty acids, Rothamsted Research has applied to Defra (the UK government’s Department for Environment, Food and Rural Affairs) ‘to conduct a field trial of Camelina plants that have been genetically modified to produce omega-3 oils that may provide health, environmental and societal benefits’. Interestingly, one of the enzymes in the 5-gene cassette used to genetically manipulate EPA levels in the plant is derived from Phytophthora infestans – the potato blight-causing oomycete (definitely NOT a fungus) which infamously caused so much devastation to the potato crop of Europe in the 19th century.  Maybe this is an opportunity for that notorious plant pest to do some good for a change! And something to ponder as you fry your naturally omega FA-enriched fish in GM-enhanced camelina oil…? Regardless, let us hope that false flax (an alternative common name for the plant) does not give false hope but, rather, provides ‘gold-of-pleasure’ (another of its common names). And that this 21st century fish oil project has more to offer than the 19th century’s over-promising, under-delivering pedlars of ‘snake oil’! Here’s a video showcasing the work at the 2014 UKPSF meeting.

[For more on the proposed GM trials, there is a dedicated Questions and Answers Section on the Rothamsted Research website. But what we really want to know is whether there is a hidden agenda to use the GM-crop to produce jet fuel for the F-22 raptor supersonic fighter aircraft, which apparently can fly very well using biofuel produced from Camelina… In which case, maybe GM stands for Go Mach – Ed.]

[Update - since this piece was originally penned, not only has the GM trial been approved but it has taken place and the crop harvested. It is anticipated that the results will be published in an open access journal later this year - Ed.]

Predicting mixed-species litter decomposition

Predicting mixed-species litter decomposition

Predicting mixed-species litter decomposition

The biomass-ratio hypothesis states that ecosystem properties are driven by the characteristics of dominant species in the community. Tardif et al.  measure decomposition rates of litter from four herb species at three sites along a correlated climatic gradient of temperature and precipitation in order to test the predictive value of the hypothesis. They find that community-weighted means of monoculture values provide good predictions of mixed-species’ litter decomposition, converging to the predicted values with increasing species richness and in climates less favourable to decomposition. The results support the idea that the biomass-ratio hypothesis, operationalized as community-weighted means, could offer the opportunity to predict ecosystems processes at larger spatial scales and in a changing environment.