An update on a microscopic study of orchid leaves

Another little study done.   Just over a year ago I summarised some of the work I was doing looking at orchid leaves under the microscope to see if they helped identification.   I added a lot more data in 2018, especially on helleborines (Epipactis) and over the last few days I have revised my summary in a Powerpoint presentation.    If anyone is interested then it can be downloaded by following this link to my dropbox account  Orchid Leaf Edges TS

In all I examined 36 orchid species, and ignoring the Schedule 8 plants and those without leaves, there are another 5 on which I would like to collect data.   All were examined at x100 from small slivers of often damaged leaves using a home microscope.  In the field it is possible to get up to around x40.

Most have regular, toothed, leaf edges, with peaks and troughs, and I used several terms to describe those tooth patterns, to try to capture subtle differences.   I measured the width of the teeth, and the peak height over the trough.    Where teeth were angled I measured the average slant.   Some though, such as the butterfly orchids (Platanthera) and the bee orchids (Ophrys), had entire edges (i.e. not indented).  Some had spotted leaves, some had purple tinged edge cells.   Some had stomata on the upper leaf surface, others did not.   Two other measures were the shape and appearance of the leaf tip, and the number of leaf veins.   All were faithfully recorded by photographs.

A few -  helleborines - did not have regularly patterned edges, but rather, unequally sized papillae, together with areas of only small obvious edge cells, or indeed gaps.

There are definite differences between most species of marsh orchids (dactylorhiza), although common spotted and heath spotted were similar.   The 3 fragrant orchids showed subtle differences,  for example chalk fragrant had more crowded teeth then the other two, and upper leaf stomata were more likely.   The helleborines also had some differences but maybe not enough to safely attribute a species  name to a particular plant.  Nevertheless leaf edge characteristics might be useful in the mix of plant morphological characteristics in identification.

New Year Plant Hunt: an Improbable Calculation

For the fourth consecutive year, I took part in the BSBI organised, New Year Plant Hunt.   This year on my regular route plus an extension to include a fragment of woodland, I found 22 plants in flower.  Like for like, excluding the add-on, the total was 19, two species less than last year.   They were almost all perennial weeds, and there were no early flowering spring plants.  Because our weather station is mis-recording temperatures I have not made a weather comparison

Since the event I went looking for flowering plants around Greta Tew and Duns Tew, two villages south east of Hooky, finding another 8 species that were not on my Hooky NYPH list - again weeds, except for a few flowering snowdrops- bringing me up to a total of 30 species.   To find them I covered 30 Km in 10 hours, 1 species every Km or 3 species per hour. The rate has though dropped off to a meagre 0.36 species per Km, (8 additions in 21 Km and 10 hours) on the Great Tew and Duns Tew exploration.

The NYPH is becoming increasingly a competition, but in and around Hooky the leader board is but an abstract, surrounded as we are with arable and pasture, some of it quite joyless in winter especially the bridleways churned up by horses.  As far as I am aware the highest total is for Swanage, 120 species.   At the reduced capture rate of 0.36 species per Km I would have to cover another 247 Km to get to that number.   Swanage is in fact 220 Km from Hooky.

For what it is worth my list is:

Hooky NYPH

Annual Meadow Grass	Poa annua
Cock's-foot	Dactylis glomerata
Common Chickweed	Stellaria media
Common Field Speedwell	Veronica persica
Daisy	Belis perennis
Dandelion	Taraxacum officinale
Dog's Mercury	Mercuralis perennis
Dove's-foot Cranesbill	Geranium molle
Feverfew	Tanacetum parthenium
Gorse	Ulex europaeus
Greater Periwinkle	Vinca major
Groundsel	Senecio
Hazel	Corylus avellana
Hogweed	Heraclium sphondylium
Petty Spurge	Euphorbia peplus
Primrose	Primula vulgaris
Red Dead-nettle	Lamium purpureum
Shepherd's-purse	Capsella bursa-pastoris
Smooth Hawk's-beard	Crepis capillaris
Smooth sow-thistle	Sonchus oleraceus
White dead nettle	Lamium album
Yarrow	Achillea millefolium

Great Tew and Duns Tew additions

Snowdrop	Galanthus nivalis
Common Fumitory	Fumaria officinalis
Scentless Mayweed	Tripleurospermum inodorum
Pineappleweed	Matricaria matricarioides
Creeping Buttercup	Ranunculus repens
Herb Robert	Geranium robertianum
Rape	Brassica napus
Nipplewort	Lapsana communis

Morphology of Butterfly Orchids to the North and East of Stroud in Gloucestershire (for the specialist only!)

There are two butterfly orchids in Britain; only one, Greater Butterfly orchid can be found near to Hook Norton, though when we lived on Skye we had both on our croft and made a long term study of them.  One of these days we might get the results published.   Uncommonly the two can hybridise and one of the best places to see the two parents and the hybrid is near Stroud in Gloucestershire.

 I wrote a blog some months ago about a visit to Strawberry Bank in Gloucestershire to look at the butterfly orchids there - a site for both greater butterfly (Pc = Platanthera chlorantha), lesser butterfly (Pb = P. bifolia) and the hybrid between them (Ph = P. x hybrida).    In fact in late May and early June I went to 5 sites in all to the north and east of Stroud, finding evidence of hybridisation.   I have now had a chance to analyse the data I collected, and review some recent relevant literature.

Bull's Cross

Location Map

I made morphometric measurements of examples of the orchids I found at each site trying hopefully samples as representative  as possible.  Some measurements were made in the field such as height of the flowering spike, and width of the lowest leaf, but for measurements of the floral parts I photographed a flower from each plant alongside a scale, generally but not consistently from near the base of the inflorescence, and these were analysed at home.    From the pollinia positioning I identified them as Pc, Pb or for intermediates, Ph. 

Platanthera bifolia, Lesser Butterfly Orchid from Bull's Cross

Hybrid from Bull's Cross

Platanthera chlorantha Greater Butterfly Orchid from Bull's Cross

 In all I made up to 16 measurements on  66 plants (30 from Bull's Cross, 9 from Strawberry Bank, 12 from Cranham Common, 6 from Sheepscombe and 9 from Edge Common.  I found hybrids at all 5 locations, and in addition both parents at Bull's Cross and Strawberry Bank, but only one of the two parents was present at Cranham Common (Pb), Sheepscombe (Pb) and Edge Common (Pc).    Cranham does have a population of Pc and I was probably there before they were fully in flower.   I also have results for a 'pure' population of Pc at Greenfield farm in Oxfordshire, but this population is shaded by ash and beech whereas the Gloucestershire plants are ut in the open, and might not be fully comparable;  other work I have done suggests that shaded plants, or plants in greater sward are bigger.

A summary of the results follows, picking out those characteristics which discriminate between Pb and Pc (significant with a probability factor of less than 5% by a simple T test).  Beyond pollinia positioning (which was my a priori discrimination) the height to leaf width ratio was a discriminator as was the width of the dorsal sepal (the one at the top!) and the labellum length.

By contrast  none of the measures other than the pollinia positioning gave a good way of discriminating between Ph and the two parents.   Esposito et al (1) in a study of hybridisation in two Belgian populations found a clustering of the intermediates (my Ph) towards Pb, perhaps suggesting that there is variance in Pb, rather than hybridisation  but in my results there is scatter right across the centre ground between values for the two parents suggesting introgression is occurring, and there may be some F1 hybrids but there is also much back crossing.  (I collected a few seeds in October, from these hybrids and they appeared to be viable)  The average value for Ph plants of the subtended angle between pollinia is midway between values for the parents, as is the labellum length. Spur length and dorsal sepal width lie closer to Pb, but the ratio of spur length to labellum length is closer to Pc.

Esposito et al did find though that values for both parents growing side by side were closer and less extreme than where the two species were isolated from each other.   My results seem to be in agreement though I have not done a proper statistical study yet (I am still trying to get my head round Principal Component Analysis which seems to be the best technique).  An example is in the Pc spur lengths at Edge Common  where the average was 21.9mm, the same as for Pb plants elsewhere (rather, I would have expected it to be nearer 30mm) and compares to an average of 35.3mm for Pc growing alone at Greenfield Farm. 

Platanthera muelleri (Pm) was proposed as a new species in 2017 by Angela and Heinz Baum (2) arising out of a collaboration detailed in Durka et al,(3) undertaking morphometric and molecular studies of some Dutch populations; that name has been accepted on the World Checklist of Selected Plant Families (WCSP) maintained at Kew.   Essentially it is a long spurred plant resembling P. bifolia. A discriminator is the ratio of the spur length to labellum length; 2.8 +/- 0.3 for Pm, cf 2.3 +/- 0.3 for Pb, 2.2 +/- 0.3 for Pc and 2.1 +/- 0.3 for Ph.   All the plants I found in Gloucestershire agree to one of Pb, Pc, or Ph.  None resemble the type description of Pm.

In passing it is worth commenting on some other proposed species names.   P. fornicata was a name revived in 2011(4) but it appears to be a synonym of a Pb subspecies (P. bifolia subsp bifolia), a woodland form, I think.   WCSPF recognises this as a subspecies but not P. fornicata as a species, and it also gives three other subspecies: osca, subalpina, and laxiflora, but I have no details of their distinguishing characteristics.  

Refs:

1 Characterization of sympatric P. bifolia and P. chlorantha populations with intermediates - Esposito et al 2018; Peer J, DOI 10.7717
2 Platanthera muelleri - Baum, A and H 2017;  Journal of the European Orchid 49 133-152
3 Darwin's legacy in Platanthera - Durka, et al 2017;  Plant Systems Evolution 303 419-31
4 Revision von Platanthera bifolia sensu lato -  Buttler, 2012; J Wetterau Ges Naturkunde 159-61 93-108

More on fragrant orchids and their identification

Back in July I put down some thoughts on identifying the three fragrant orchids following a visit to Waitby Greenriggs in Cumbria; distinguishing between them is not easy. 

I have now done some further analysis because  over the summer I gathered data from another 7 locations where fragrant orchids occur, although not sadly from Clay Lane Meadow which is only 2 km away.  Frustratingly it  was closed off because of damage cause by irresponsible dog walkers. 

Marsh fragrant (Gymnadenia densiflora)

Chalk fragrant (Gymnadenia conopsea s.s.)

 

Heath fragrant (Gymnadenia borealis)

                                                                   The locations were:

• Helsington, Cumbria - Heath fragrant
• Rampsons Farm, South Stainmore - Heath and Marsh fragrant
• Garrow, Perthshire - Heath fragrant
• Keltneyburn, Perthshire - Heath fragrant
• Wye Down, Kent - Chalk fragrant
• Eades Meadow, Worcestershire - Chalk fragrant
• Sheepscombe, Gloucestershire - Chalk fragrant
• Waitby Greenriggs, Cumbria - Chalk, Marsh and Heath fragrant.

In all I made measurements on 40 plants: 

• Spur length (in most cases)
• Sepal length and width
• Angle of the sepal above or below the horizontal (+ or -)
• Number of flowers (in most cases)
• Leaf width (in most cases)
• Width and length of the labellum
• Width and length of the central lobe (except in most of the Heath fragrants where it is not readily distinguishable).

I then looked for measures that can be used for differentiation.  The first step was to decide what a particular plant was which immediately introduces bias because I relied mainly on the floral parts, such as the size of the labellum lobes.  The gold standard, DNA analysis, was not a possibility.  The keys in Stace 3rd edition were not wholly reliable, and with a revised edition of that reference on the way, hopefully there will  be more insights. The results in summary were as follows.

There was significant variation and the boundaries between species on a particular characteristic overlapped.   Nevertheless it does seem possible to separate out the three species.  Easiest is to pick out Heath fragrant.  I found statistically significant differences (using a T-test) between it and the others on labellum width and length, spur length, and the two ratios which I think are the most useful because they avoid absolutes which might be affected by environment and latitude, viz. labellum width to length and sepal length to width.   The former is not a surprising result because my initial allocation was probably weighted  towards this characteristic.

The other two were harder to separate, but on three parameters there was a statistically significant difference (using a T-test):  angle of sepals to the horizontal, labellum width to length and sepal length to width.  Given the variability and species overlap, I would look for best fit against each of them.

I did not measure the height of each plant nor the height of just the inflorescence.  A pity because it is probably worth looking at further, though I suspect there will be latitudinal effects, with northern plants on average shorter than those further south.   It was also noticeable that the inflorescence of Chalk fragrant plants had a mor lax appearance, and what might be very useful is the ratio of the number of flowers to the length of the inflorescence.  

I should also mention habitat and the common names define what is likely in marshes, downland and essentially north and west Scotland, but it is not 100% predictive.  The Helsington plants of heath fragrant for instance are overlying a base of limestone, so there is probably a cap of more peaty soil over it.

Looking forward there could be some significant  advances in fragrant orchid disaggregation.  I have mentioned the new edition of Stace which comes out in late January,  but Richard Bateman and Ian Denholm, BSBI's joint orchid referees, will publish a  note on identification in the January edition of BSBI News, to be followed later in the year by an academic paper on the subject.   So next summer fragrant orchids will become easy peasy, at last!

The Commonest Orchid in Britain and Ireland

It's November, miserable and grey.  We have sleet this morning.   Earlier this week I had just two moths in the light trap, and that is probably it for this year.  No other insects around.   Nada.

It's time for indoor botany.   I am preparing a little presentation for my village garden society on Britain's wild orchids and in doing so natural questions pop up such as  'how common / rare are they' and 'where do you find them'.   Reading newspaper headlines it the phrase is always 'rare orchid'.

I have been a member of BSBI for at least 25 years.  I have been on a few field trips, which are mainly plant recording events, but because my identification skills for anything other than orchids, gentians and saxifrages, are very limited my contributions are little better than nothing.   So why remain a member?   Well there are benefits and a significant one is access to the database derived in part ironically from those recording trips.   It is terrific, and it was to this database I turned  so that I could work out how common or otherwise orchids actually are.   I analysed records for this century for all the orchid species by vice county.   These vice counties have I think been unchanged since 1852 in Britain when they were introduced by Watson, (Praeger introduced the same concept for Ireland in 1901) unlike the administrative county boundaries, which change with every passing whim of politicians. 

There have been taxonomic changes this century.   I eliminated records for Dactylorhiza majalis, which is now mostly reclassified in the most part to D. kerryensis, an Irish species, and the fragrant orchid aggregate, Gymnadenia conopsea s.l (sensu lato = in the broadest sense) was split in the early 2000's  into 3 species, formerly recognised as sub-species, (Heath fragrant, G. borealis; Marsh fragrant, G. densiflora; Chalk fragrant, G. conopsea s.s (sensu stricto = in the narrowest sense)) 

Heath Fragrant Orchid

The results were surprising.   The database gives no hint as to plant abundance, and therefore one record on one day in one place is equivalent to 1000's in lots of places every year, so there are limitations.  Nevertheless it is the best we have, and the results were surprising.   The commonest orchid (i.e. the most widespread) was Common spotted orchid (Dactylorhiza fuchsii) which occurred in 152 of the 153 vice counties in Britain and Ireland.   Three other orchids, Common twayblade (Neottia ovata), Early purple (Orchis mascula) and Heath spotted (Dactylorhiza maculata) were not far behind and occurred in 150, 149 and 148 vice counties respectively.   By contrast 4 orchids were only found in one vice county; Ghost orchid (epipogium aphyllum) made its only appearance this century in 2009, and the Hebridean marsh orchid (Dactylorhiza evudensis) is not now recognized as a separate species and should not really be included.  (I saw it this year on North Uist in the sand dunes there.)

Common Spotted Orchid, Eades Meadow, Worcs

Common Twayblade, Cornbury, Oxfordshire

                                               

Early Purple Orchid, The Burren, Ireland

Increasing the granularity of the analysis by looking at the data by hectad (10km x 10km squares), in Britain only (excluding Ireland), there are around 3500 hectads.  Common spotted orchid is still the commonest, appearing in 2121 hectads (61%), but the top order ranking changes a little with the next three reshuffled.  Heath spotted orchid is in 1744 hectads (50%), Early purple is in 1308 (37%),  and Common twayblade in 1205 (34%).

The most orchid-rich vice county was Westmorland, where I was born and raised.   The administrative county disappeared years ago and is now remembered only as a service station on the M6, and this vice county name.  It has 31 species, and seems to be a cross over point between northern and southern species, amplified by the upland there.   My present home, the Oxfordshire vice county, comes third with 29 species.   Two small Scottish vice counties have only 6 species.  The 3 pictures below were of plants at Helsington near Kendal.

Dark-red Helleborine, Helsington 

Fly Orchid, Helsington

Fly Orchid, Helsington

But wherever you live in Britain and Ireland you are never far from an orchid of one sort or another

Leaf Miners

Things quieten down in autumn, and its a chance to look at wildlife that is easily overlooked.  Fitting into that category are leaf miners, flies and moths which at the caterpillar stage feed and grow in the leaves of trees and other plants.  I have seen a number of postings in blogs and on Facebook (eg Plants of Skye, Raasay & the Small Isles) describing leaf miners that have retarded leaf senescence leaving green islands in an otherwise brown leaf, so I started looking in the hedge in front of my house for similar examples and this widened out into a more general search for leaf miners.  To identify them I relied on British Leaf Miners - with a heavy dependence on miners being specific to host plants - and the captions on a few examples that follow may be entirely wrong, if that dependence is misplaced.  But it's a start, and the best thing is that they are easy to find in quiet moments.

 Phyllonorcyter cerasicollela, Wild Cherry

 

Phyllonorcyter cerasicollela, Wild Cherry

Phyllonocyter hostis on Crab Apple

Phyllonocyter hostis on Crab Apple

Stigmella tityrella on Beech

Stigmella microtheriella on Hazel

Butterfly Orchids in Oxfordshire; Bats Nearer Home

An eclectic combination.   

Over the last few days I compiled the results of surveys I did of the two butterfly orchid species in Oxfordshire for a status update for the Flora Guardians programme.

I had some site references from the Flora Guardians programme itself and then added in records (primarily from the year 2000 onwards) from the BSBI Distribution database, access to which and the analysis it allows is well worth the BSBI subscription alone.  Unfortunately there is a misalignment of boundaries because the administrative county contains all of BSBI's vice county 23 (which I used) but also bits of vice county 22.   No matter, I ended up with 26 localities where Greater butterfly orchid has been seen this century, and 5 where Lesser Butterfly orchid has been seen.  I also added a site which I have been studying over the last 2 summers, and probably holds the largest number of plants in the county but was on neither database.

I surveyed only 6 of the Greater butterfly orchid localities, though almost certainly these contain the bulk of the plants.  I also surveyed 2 of the 5 Lesser butterfly orchids locations.   Greenfield Farm (to which there is no public access) had over 500 plants of which 196 produced flowering spikes, an increase of on 2017.  The plants are in beech / ash woodland, and thus quite heavily shaded; there is no real herb layer, but there is regrowth of ash and beech and a tangle of brambles.  Habitat maintenance is selective; in the 2017/18 winter the bramble was cut back in three quarters of the site but left in the other quarter.  Dog's mercury is becoming a problem in one corner, developing a heavy smothering sward, perhaps encouraged by increasing light levels (which we are trying to monitor) because the ash is suffering from die back. 

BBOWT's  Warburg reserve has both species. An underestimate for certain,  I counted around 40 flowering plants of Greater butterfly orchid,  scattered quite thinly through the reserve apart from one hotspot.  Generally they were in scrub regrowth in areas cleared some years back.  Numbers have been stable since 2015, when comparable annual counts started.   Lesser butterfly orchid is rare and at Warburg there were only 4 flowering plants growing amongst scrub on the edge of rides.   There was also one flowering plant of the hybrid P. x hybrida, last seen (recognised?) in 2003.    Some modest scrub clearance around the existing plants might help.

Elsewhere I counted 28 scattered flowering Greater butterfly orchids at Bald Hill, English Nature's Aston Rowant reserve on top of the hill amongst low scrub, and a few plants on the Cornbury estate adjacent to a footpath.

So, a few sizeable colonies of Greater butterfly orchids, a species which is probably under-recorded because there may be more in some of the Chiltern woodlands.   By contrast, lesser butterfly orchid is just abut hanging on at Warburg, though there has only ever been a few plants there.  Unless the 3 locations that I have yet to look at have plants, it might be that those few remaining at Warburg are the only examples in Oxfordshire.

Leaves count! Cornbury

Greater butterfly orchid, Bald Hill

Lesser butterfly orchid with scale, Warburg

Closer to home I have been out bat detecting  having bought a new toy a  bat detector which works on my phone, an Echometer touch.  It constitutes a microphone which plugs into a smart phone and links to an app. Its clever, providing a chart of a bat echolation sound in what it calls real time expansion or as a heterodyne signal.   For lazy people like me who just want to know what bats are out there, it also gives an identification which is correct most of the time, and it gives a gps location.    We are in to November, yet even so, possibly because it was a mild evening  I picked a noctule and a common pipistrelle near home.  

Common pipistrelle sound wave