This blog post is part poem, part scientific explanation. An unusual combination, but one I had fun creating. This topic grabbed my attention from the moment I walked beneath the Super Blue Moon on 30th August 2023 at 23:30 at night - a night for inspiration!
As I stood admiring the bright orb in the sky, I turned my attention to the white roses at my feet. Illuminated by the reflected light, I noticed the dark pink flecks on the petals. I idly thought about how they must have been caused by raindrops concentrating the UV rays, causing the flowers to 'burn' and develop freckles, like me. I found this idea quite enchanting and decided there and then I wanted to write a poem about it, exploring the effects of environmental stressors on both our lives. A fanciful thought, as I've never written half a poem before in my life! A challenge is a challenge though - how do you know if you're the next Edgar Allen Poe if you don't give it a go?
Off I set, with the seed of prose stuck in my head but to my surprise, in my initial research unveiled that freckles on the flowers were not actually caused by the sun, but something I found altogether more interesting. I hope you enjoy the poem, which should enlighten you and read on afterwards to explore the scientific explanation!
How these freckles came to be
- Claudiomics
Gardens overflowing, with glorious roses growing,
But many petals, have many freckles.
And by poems end you’ll end up knowing,
How these freckles came to be.
Spores hover in air and hold – the infamous Grey Mould,
Rose riled up with derision,
Shouting “How could they be so BOLD?!”
As the mould prepares to invade.
Botrytis cinerea – a fungus that rots,
Pathogen genocide declared,
But the rose screams “I think NOT!”,
As a silent War of Roses is wrought.
Botrytis lands, through air that is damp,
On roses ready to rage.
And petals sense these perilous PAMPs,
Preparing genes to repel.
ERF, MYB and NAC, kick in as rose is attacked.
Botrytis feels doom, as roses bloom,
Evade their digestive tract,
Forcing the foe to fall back.
The Spores are dismayed, for in this crusade,
Rose reigns victorious, feeling glorious!
But dark blood is spattered, for the price is paid,
In pockmarked petals with freckles.
A War of Roses is told, so hopefully now you see,
That internal ballet, prevents decay,
How these freckles came to be,
They're borne by battling Botrytis.
Well, there you have it! A dramatised poem about a microscopic battle between rose petal cells and the fungus Botrytis cinerea. Now I'll delve a little into the science, so you can revisit a few different aspects of the poem to clarify its meaning.T
The Science
Introduction: Why Botrytis is a Billion Dollar Bother
What is Botrytis cinerea?
As I started looking into these freckles (figure 1 a-c), I found out that they were actually lesions caused by the fungus Botrytis cinerea, which is also known as grey mould (figure 1d-f).
Figure 1. (a-c) Lesions developed at different stages on infection on Rosa hybrida petals due to the fungus Botrytis cinerea (a, none; b, light; c, heavy). Photos taken from my garden. (d-f) Botrytis cinerea (d, Photomicrograph of hyphae with terminally emerging microconidiophores, taken from (1); e, Bright field image of Botrytis cinerea conidia treated with nanopores for 10 minutes, with the dotted box showing germinated spore, taken from (2); f, Botrytis cinerea infection on a strawberry also affects the leaves, taken from (3)).
Roses are one of the worlds most sought-after cut flower, making up more than a third of the total cut flower industry, and an estimated €1.3 mil in revenue was lost in 2002 due to Botrytis cinerea (4,5). This necrotrophic pathogen kills host tissue to feed on the decaying matter and can infect over 200 different plant species causing widespread economic losses (4,5).
Why study this pathogen?
Due to the widespread use of chemical fungicides (which are harmful to the environment, as well as consumer and farmer health), multi-drug resistant strains of Botrytis cinerea are appearing rapidly (6). This fungus has mutated to express less proteins where drugs can bind, and more proteins that pump fungicides out - rendering them far less effective (6). Current management of this disease should be re-examined and research turned towards unravelling the complex biochemical interactions of host defence. This will provide alternative solutions to fungicides, such as natural molecules which induce host defence pathways, and offer some respite from this pathogenic rascal.
A study carried out on strawberries infected by Botrytis cinerea concluded that selective breeding to obtain a high resistance crop would be too tedious and never result in a completely resistant fruit (3). One day, they hope to create a genetically modified strawberry line which is disease tolerance, but as of now the public's scepticism of genetically modified organisms (GMOs) prevents this from becoming a reality (3). Another study conducted on roses (7), provides more information on rose petal defence mechanisms, which I will now focus on as this was the paper I relied on to curate some of the lines in my poem!
Rose Petal Immunity against Botrytis cinerea
Recognition of the Pathogen
Immunity comes in two different stages: recognition of the pathogen (harmful organism), and retaliation (fighting back). The plant must recognise and identify if a foreign object is harmful and if so, how to respond.
Plants are able to do this by having immune receptors called pattern recognition receptors (PRRs) on their cell surface (figure 2)(7). These PRRs bind and recognise pathogen-associated molecular patterns (PAMPs) which are on the surface the invading pathogen and were mentioned on line 15 of the poem... perilous they are! (7). PRRs can also recognise damage-associated molecular patterns (DAMPs) which are its own cellular components and only exposed when the cell is damaged (figure 2 a,b)(7). Necrotrophic pathogens such as Botrytis cinerea secrete polygalacturonases to degrade cell walls, resulting in the release of oligogalacturonides which are the plant's DAMPs and will activate downstream cellular signalling (8).
"And petals sense these perilous PAMPs"
Figure 2. Showing the internal biochemical signalling within a rose petal cell after Botrytis cinerea invasion a. A pattern recognition receptor (PRR) on a plant cell membrane recognising pathogen-associated molecular patterns (PAMPs) from a fungal spore. b. A PRR recognising a damage-associated molecular patterns (DAMPs) from its own cell makeup. c. Phytohormones induced by downstream signalling after PAMPs and DAMPs are recognised. d. Transcription factors (TFs) induced by downstream signalling, leading to induction of genetic manipulation in the nucleus (more proteins made!). Diagram made using BioRender software.
Response to Invasion
After the pathogen is recognised, the receptors activate signalling cascades that kick-start the immune response. Phytohormones (plant hormones) are responsible for activating other plant hormones and signalling molecules such as transcription factors (TFs) which trigger the expression of genes related to defence (7).
Liu et al., were able to determine 7658 genes that were significantly changed 48 hours after rose petals were inoculated with B. cinerea (7)! That's a solid war effort!
Phytohormones... or fight-o-hormones...
Within the defence-induced phytohormones (figure 2c.), 325 genes were activated (7). The Brassinosteroid class made up over half of these upregulated genes, and other notable hormones include gibberellins, ethylene, abscisic acid and cytokinins, among others (7). Many of these hormones activate transcription factors, which can travel into the nucleus of the cell and affect transcription (a process required to make new proteins from DNA) (figure 2d) (7). Brassinosteroids are steroid hormones which ordinarily stimulate seedling development and the reproductive process, but under invasion by Botrytis cinerea activates genes to remodel the cell wall architecture to decrease digestability (8).
Transcription Factors
Liu et al., found 188 defence-responsive transcription factors that were significantly activated after applying fungal spores to the petals, mainly within the TF families ERF, MYB, NAC (mentioned within the poem line 17), as well as WRKY and bHLH, among others (7).
"ERF, MYB and NAC, kick in as rose is attacked,"
Further research has been conducted by Liu et al., which advanced their knowledge of the immune response of roses (Rosa sp.) and showed that the simultaneous activation of cytokinin and inhibition of abscisic acid phytohormones increased rose immunity to Botrytis cinerea (9). This process was induced by the same transcription factor - the transcriptional repressor Rosa hybrida (Rh)WRKY13 (9). This repressor binds to both the promoter regions and prevented expression of the cytokinin degradation gene (RhCKX3) as well as the abscisic acid response gene (ABA insensitive4) (9). Blocking cytokinin degradation and preventing abscisic acid transcription increased Botrytis cinerea immunity (9).
Conclusion
I’ve never given much consideration to the defence mechanisms within plants, but I now have a newfound respect for what plants have to deal with on a day to day basis (apart from being chopped apart by us).
To summarise the rose immune response - Botrytis cinerea is recognised by PRRs on the surface of rose cells, triggering an onslaught of phytohormones and TFs which work together to fight back against this fungus by altering the cell wall to decrease digestibility (7,8). However, this immune response is not always successful and infection can end up degrading blooms, causing great economic losses in the cut flower industry.
Liu et al., have suggested one means to improve resistance without the use of fungicides by increasing the cytokinin phytohormones while simultaneously decreasing abscisic acid activity (using the same transcription factor), they reported a significant increase in resistance (8). Applying molecules such as these to naturally induce resistance to Botrytis cinerea makes the use of ineffective and harmful fungicides redundant, and with more research more of these alternatives will become mainstream. Some researchers are in favour of creating genetically modified organisms which would be resistant to this fungus - what are your thoughts? Would you buy GMO roses? Eat a GMO strawberry? Let me know in the comments below!
"GMO? OMG NO!!!" - Somebody, probably.
When it comes to this fungus though, it isn't all doom and gloom. In fact, some grape farmers actually apply Botrytis cinerea to their grapes so the fungus can cause 'noble rot' (10,11). The fungus breaks down glucose and cell walls within grapes to produce a sweeter, tastier wine and even boosts the amount of polyphenolic antioxidants which are beneficial to consumers (10,11). Now, I'm not telling you to go and suck on a mouldy grape here. What you do with mouldy grapes is up to you. But I hope you enjoyed the read, and think back to this poem whenever you see petal freckles out and about. Or maybe even when you're enjoying a glass of botrytized wine!
- Claudiomics
References
1. Grindle M. Phenotypic Differences Between Natural and Induced Variants of Botrytis cinerea. Microbiology. 1979;111(1). https://doi.org/10.1099/00221287-111-1-109
2. De Angelis G, Simonetti G, Chronopoulou L, Orekhova A, Badiali C, Petruccelli V, Portoghesi F, D’Angeli S, Brasili E, Pasqua G, Palocci C. A novel approach to control Botrytis cinerea fungal infections: uptake and biological activity of antifungals encapsulated in nanoparticle based vectors. Sci Rep. 2002;12(7989). https://doi.org/10.1038/s41598-022-11533-w
3. Petrasch S, Knapp SJ, Van Kan JAL, Blanco-Ulate B. Grey mould of strawberry, a devastating disease caused by the ubiquitous necrotrophic fungal pathogen Botrytis cinerea. Molecular Plant Pathology. 2019;20:877-892. https://doi.org/10.1111/mpp.12794
4. Dean R, Van Kan JA, Pretorius ZA, Hammond-Kosack KE, Di Pietro A, Spanu PD, Rudd JJ, Dickman M, Kahmann R, Ellis J. The top 10 fungal pathogens in molecular plant pathology. Mol Plant Pathol. 2012;13(4):414–30.
5. Williamson B, Tudzynski B, Tudzynski P, Van Kan JAL. Botrytis cinerea: the cause of grey mould disease. Molecular Plant Pathology. 2007;8:561-580. https://doi.org/10.1111/j.1364-3703.2007.00417.x
6. Leroch M, Mosbach A, Walker A, Fillinger S, Mernke D, Schoonbeek H, Pradier J, Leroux P, Waard M. Fungicide-Driven Evolution and Molecular Basis of Multidrug Resistance in Field Populations of the Grey Mould Fungus Botrytis cinerea. PLOS Pathogens. 2009;5(12):e1000696. https://doi.org/10.1371/journal.ppat.1000696
7. Liu X, Cao X, Shi S, Zhao N, Li D, Fang P, Chen X, Qi W, Zhang Z. Comparative RNA-Seq analysis reveals a critical role for brassinosteroids in rose (Rosa hybrida) petal defense against Botrytis cinerea infection. BMC Genet. 2018;19(62). https://doi.org/10.1186/s12863-018-0668-x
8. Nafisi M, Fimognari L, Sakuragi Y. Interplays between the cell wall and phytohormones in interaction between plants and necrotrophic pathogens. Phytochemistry. 2015 Apr;112:63-71. doi: 10.1016/j.phytochem.2014.11.008. PMID: 25496656.
9. Liu X, Zhou X, Li D, Hong B, Gao J, Zhang Z. Rose WRKY13 promotes disease protection to Botrytis by enhancing cytokinin content and reducing abscisic acid signaling. Plant Physiology, Volume 191, Issue 1, January 2023, Pages 679–693, https://doi.org/10.1093/plphys/kiac495
10. Azzolini M, Tosi E, Faccio S, Lorenzini M, Torriani S, Zapparoli G. Selection of Botrytis cinerea and Saccharomyces cerevisiae strains for the improvement and valorization of Italian passito style wines. FEMS Yeast Research. 2013;13(6):540-552. https://doi.org/10.1111/1567-1364.12054
11. Nikfardjam MSP, László G, Dietrich H. Resveratrol-derivatives and antioxidative capacity in wines made from botrytized grapes. Food Chem. 2006;96:74-79.
Commentaires