The Polyploid Marsh-orchids

Polyploid is a feature of many plants. Much of our agricultural croppage is polyploid (4n, 6n, 8n or even 16n) because higher chromosome numbers equate to higher yield. Compare your carrots to the roots of the wild carrot to see what I mean.
At its final division during meiosis a pre-germ cell's pairs of chromosomes disassociate and half migrate to one new cell and the the other half to the other cell. Thus two haploid or 1n cells are formed. At fertilisation two haploid cells fuse to form a single 2n cell. This mixing of chromosomes, half from both parent organisms ensures genetic diversity and is the basis behind why there are two sexes in all higher organisms. However, occasionally in plants the final cell division goes wrong and all chromosomes migrate to one of the new cells (the other will die). This new pollen cell or seed cell can lead to a new fertilised cell with a chromosome count higher than 2n. Instant speciation.

Polyploidy gives a species several evolutionary advantages. Spontaneous mutations of the genetic code are random. Most will confer no advantage to a plant, some will be detrimental, but the odd one does; and this will persist. Having four copies of each gene means that any negative effects of a mutation of one of them is minimised. During meiosis a process of chromosome cross-over can mix gene from both parents onto one chromosome. New combinations like this may give the plant an advantage, and having four sets of chromosomes increases the chances of this. Autopolyploids can be larger than either parent, so robustness and being easier to find by pollinators helps them survive, but the converse may occur with allopolyploids like the Heath spotted, as there is now an element of possible inbreeding. The new genetic make-up may confer the ability to colonise a different habitat to either parent. We can see this in the British Marsh-orchids. This may be due to a wider range of enzymes being capable of being produced, or double-doses of enzymes. The evidence for the evolutionary advantages of polyploidy is said to be the richness of the British endemic flora when there has been but 8000 years separation from continental Europe.

I have been trying to find out what the mechanism is for a new tetraploid (4n) plant to arise. Some propose that it is two 2n zygotes at fertilisation creating a tetraploid descendant, others proposing a triploid intermediate. Suppose it is pollen that is 2n. That fertilises a haploid (1n) cell, creating a triploid (3n) plant. Unfortunately, it is known that triploid hybrids formed from the union of diploid and tetraploid parents is rarely fertile. That final cell division at meiosis is disturbed with odd numbers of each chromosome migrating to the two new cells. Even if one has a full complement of chromosomes, any incomplete duplication of some chromosomes throws everything off kilter.
The idea that two 2n germ cells come together however means that evolution plays its Joker. If 1 in 10,000 germ cells is 2n, then the chances of two of these coming together is 1 in 100 million. Obviously the numbers may be much less than that, but even so, any seed produced also has to survive to flowering just to continue, and then be self-fertilised until sufficient descendant plants have formed the first colony of a new tetraploid.

Because it is two 2n parents involved we know that it will be plants such as D. incarnata and D. fuchsii involved as the parents. Other diploid Dactylorhiza species have also been implicated for some mainland continental polyploid species, but those two are the only ones which have been the original parents of the British and Irish tetraploid Dactylorhiza species - though there may be other possibilities. I have deliberately used the word species here, though some may disagree. From plastid haplotype analysis it is shown that in each of the British species it is D. fuchsii that is the ovule or seed (maternal) parent and D. incarnata that is is the pollen (paternal) parent. Hybrids will show a morphological appearance closer to the maternal parent due to the extra non-nuclear DNA they carry into their offspring and stabilisation of the genome (whatever that means) can indicate how relatively long ago the polyploid event took place. Because in all but one of the five (or six) British polyploids involved the same parent species it is no wonder that some do regard them as all the same species. In fact when Denholm and Bateman did an extensive morphometric analysis of the polyploids in 1983 they concluded that they were too similar to be regarded as species and they were placed as British subspecies of the the continental D. majalis. Molecular techniques since then have refined this view and placed them as separate species in their own right. This is because the parents were likely different populations separated by both time measured in thousands of years, distance measured in hundreds of miles, and probably habitat too. It is largely ancestral populations of the current species involved with the polyploid events.

There is a tendency towards losing one of the inherited ITS markers over time in the polyploid plants. The degree of this is a marker of the relative age of the polyploid species. Retention of both ITS (to an extent) is more frequent in the plants growing in UK, Ireland and Sweden. They result from more recent polyploid events. In specimens from the Alps, Mediterranean and Turkey retain the ITS from only one ITS. This displays their more ancient origins.

The allopolyploids (those where the parent were two different species) of Britain comprise

Southern Marsh-orchid - Dactlyorhiza praetissima

Northern Marsh-orchid - D. purpurella

Narrow-leaved Marsh-orchid - D. traunsteineroides

Irish Marsh-orchid - D. occidentalis

Hebridean Marsh-orchid - D. ebudensis

Additionally there is one autopolyploid with one species (D. fuchsii) providing both 2n gametes

Heath Spotted-orchid - D. maculata

Going against current thinking I have included D. ebudensis as a full species and will explain why on its own page. Meanwhile the questions that must be asked are:

1. If these, and other allopolyploids across Europe. have similar `parentage', how come they look different? The answer lies in one of two theories.
The Post-differentiation theory holds that once polyploidy had occurred the populations undergo some genetic modification and selection for a particular habitat; one that is different to either parent species. This can happen through methylation or de-methylation of certain cytosine bases of the DNA, affecting the expression of certain genes. Whilst not changing the actual genetic code, this process is heritable from parent to offspring, and allows the offspring to colonise habitats that are off-limits to either parent. There is evidence that base methylation helps a species overcome climatic factors.
Additionally there will be secondary gene flow through hybridisation with either of the parent species. And it should not be forgotten that polyploidy permits increased chances for random mutation leading to adaptive changes. Some populations exhibit genetic markers not normally found in the polyploid parent species indicating localised hybridisation, introgression, and stabilisation. They may only vary modestly in terms of morphology from other less hybridised examples of the same species
The Pre-differentiation model states that it is the variation within the parent populations that defines what the polyploid looks like. Some polyploids are quite recent, while others are more ancient. The parent species will have changed over that time.
Of course, both may occur as they are not mutually exclusive, but the resulting polyploids can follow different evolutionary paths to either parent.

2. Did autopolyploidy occur just once for each species? Difficult to answer. D. ebudensis almost certainly has occurred just once based due to its extreme localisation, and that its habitat and locale is less than 3,000 years old. D. purpurella is older but is still thought to to have arisen once and spread from that point. D. occidentalis is restricted to Ireland so again a single event. For the other two species the situation is less clear. Others arose in southern Europe while the ice sheets covered Britain and their origin is further back. But it is likely that they arose from one event.
Does this imply that the creation of a new allotetraploid is an extremely rare event, or is it that most new polyploids do not find a habitat that favours them over their parent species and so die out as an entity? There has been a suggestion that the continental species D. majalis, the Broad-leaved Marsh Orchid, may have a multiple origin, with subsequent merging of the different lines. It is perhaps the oldest European allopolyploid, perhaps dating back to before the last glaciation when it would have been inhabiting the Mediterranean area. This geographical concentration and time available would allow this. D. praetermissa is another species which also may have a multiple origin, which will be discussed on its own page. 
 

Despite all the complaints by orchid observers concerning this genus and hybridisation, there seems to be only limited hybridisation of the allopolyploid species in the wild, with little gene flow btween the species. In fact, the F2 generations are only fertile if D. incarnata is involved. There have been found few maculata genetic markers in traunsteineri or majalis despite them growing close to each other. These does appear to be some barier in place that helps the allopolyploids maintain their species individuality, and integrity. This is further justification for them each to have full species status.

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Narrow-leaved Marsh Orchid   Hebridean Marsh Orchid   Heath Spotted Orchid