Getting the Most Out of Herbaria: Systematics and Chemistry

Murder Most Florid by Mark Spencer, London: Quadrille, 2019

As mentioned in the last post, herbaria, both real and virtual, are most frequently visited by taxonomists, who are usually studying particular plant taxa or preparing flora of areas ranging in size from city parks to entire countries.  These are the traditional uses of plant collections and are still crucial.  However, several things have changed.  Now the “visit” is often to digital portals rather than onsite, making it much easier for researchers to look at specimens from far-flung institutions, IF the material has been digitized, and particularly if it is available through aggregators such as iDigBio, GBIF or JSTOR Global Plants with their links to massive numbers of specimens.  Still, coverage is uneven, with some collections more fully digitized than others.  Also changed is the way taxonomic information, once generated, is distributed.  Many flora are now published virtually, with or without an accompanying paper format.  The 2012 International Code of Nomenclature for Algae, Fungi, and Plants made it acceptable to publish descriptions of new species digitally as long as they were responsibly published and properly archived.

Plant taxonomy is also changing because of its increasing links with genetics.  Most treatments of species and genera now include DNA sequencing data.  While this has been going on for decades, the last ten years or so have seen greater use of DNA data derived from samples taken from herbarium specimens, with NGS, next-generation sequencing (NSG) making this possible. NGS techniques utilize small pieces of degraded DNA found in dried plant material easier to sequence and to determine how such sequences fit together to provide meaningful results.  That this work has revolutionized taxonomy is hardly news.  Still, it is interesting to look at how the information has solved various puzzles, such as the origin of European potatoes or the origin of the pathogenic Phytophthora strain responsible for the Irish potato famine of the 1840s.  In a study of the genetics of grapes, researchers used over 200-year-old specimens from the herbarium at the Royal Botanical Garden in Madrid.  These plants were collected by Simón de Rojas Clemente y Rubio, considered one of the founders of the botanical study of grape vines, especially varieties used in wine-making.

DNA is not the only chemical being extracted from specimens to glean useful information about plants and also about their ecological relationships.  For example, researchers in Copenhagen tested specimens of four species of Salvia used for medicinal purposes for levels of terpenoids, known to have medicinal applications.  These plants were collected over the past 150 years.  While the terpenoid levels did decrease with the specimen’s age, the “chemical composition of four Salvia species are predominantly defined by species, and there was a substantially smaller effect of year of sampling.  Given these results, herbarium collections may well represent a considerably underused resource for chemical analyses.”  Also being investigated are secondary metabolites that plants produce to control herbivore damage.  In one study researchers were able to extract pyrrolizidine alkaloids from plants in the Apocynaceae family that includes milkweed.  The specimens were as much as 150 years old, and even in those treated with alcohol or mercuric chloride, alkaloids were detectable.

There has also been work on the presence of heavy metal pollutants in collections as a way of tracking contamination.  A study at Brown University in Providence, Rhode Island analyzed samples from specimens collected around the city from 1846 to 1916, compared with newly collected ones.  Levels of copper and zinc remained relatively consistent, but lead levels were much lower in plants growing in Providence today.  It was impossible to test accurately for another toxic heavy metal, mercury, because mercuric chloride was so often used to prevent insect damage to specimens.  While toxic metals in plants might make them seem less palatable as food sources, there is an emerging field of agromining:  growing plants that are hyper-accumulators of metals like lead and mercury to eventually reduce soil contamination.  Herbarium specimens can be used to discover how long areas have been contaminated and also to identify species that are particularly good at extracting metals.  There are even some who think that growing plants in nickel-rich soil could be a way to extract this metal for sale.

Such studies suggest that the possible uses of specimens are only limited by the ingenuity of researchers in coming up with them.  It is fun to see what they can ferret out.   The British botanist Mark Spencer recently published a book on his work as a forensic botanist.  It has a great title:  Murder Most Florid (2019).  He was at the herbarium at the Natural History Museum, London curating the British and European collections when he was first asked by the police to aid in a murder investigation.  Human remains have been found in a forested area and had apparently been there for several years.  Would he be able to determine the time more precisely by studying plants at the site?  I don’t want to spoil this story or the other great ones in the book, but I will say that Spencer explains why a herbarium is essential for the work he does, now that he has become much more involved in forensics.

Reference

Spencer, M. (2019). Murder Most Florid: Inside the Mind of a Forensic Botanist. London, UK: Quadrille.

The Myriad Uses of a Herbarium

Type specimen of Altamiranoa elongata from the US Herbarium, Smithsonian Institution

When it comes to discussing the uses for plant specimens, you can’t do better than refer to Vicki Funk’s classic 2004 article “100 Uses for an Herbarium (Well at Least 72).” It is thorough and succinct. So why am I bothering to write on the same topic, if Funk has covered it so brilliantly? Well, that would deprive me of the fun of exploring some of those uses in a little more detail and perhaps even unearthing one or two new ones. While service to botany may seem the obvious place to begin, I’m going to start with medicine, the field that prompted the creation of the first documented herbarium, that of the Italian botanist and professor of medical botany, Luca Ghini (1490-1556), founded the first botanical garden at Pisa. The garden was designed to introduce students to the plants that were used by apothecaries, so they would be able to recognize them. This was obviously the function of the herbarium as well, and several of Ghini’s students, including Andrea Cesalpino (1519-1603) and Ulisse Aldrovandi (1522-1605) created specimen collections that exist to this day. Apothecaries also took up the practice, with Hieronymus Harder (1523-1607) creating a dozen bound herbaria, eleven of which are still extant.

The link between herbaria and medicine continues to this day. Those doing research on herbal medicines are often required to create voucher specimens of the plants they are studying so others can verify the identification in the future (Eisenman et al., 2012). One problem in past research was the difficulty of getting consistent results from one batch of plant material to the next. Having some of the material itself preserved in a voucher makes it easier to check whether or not the same species, subspecies, or variety was used in both cases. Not only can the plants be visually examined, but chemical tests can be done on the material if necessary. Rainer Bussmann has a chapter on “Taxonomy—An Irreplaceable Tool for Validation of Herbal Medicine” if you would like to learn more on this topic.

Voucher herbarium specimens are required in almost all botanical research, because the plant itself is the best evidence for answering questions about what species was actually growing where at a particular time. The most valuable kind of voucher is the type specimen, the record of the plant that was used in describing the species for the first time. Since modern taxonomy dates back to the publication of names by Carl Linnaeus in Species Plantarum in 1753, it is no wonder that the material he studied is itself still being tracked down to give as full a record as possible of the plant specimens, and in some cases plant images, upon which he based his descriptions (Jarvis, 2007). To this day, journals such as Taxon require authors to list specimen numbers and the herbaria where they are located for all material studied in revising taxa.

With the advent of DNA sequencing, herbarium vouchers can now be used in an entirely new way. A small piece of a specimen may yield enough intact DNA for researchers to identify a species with DNA sequencing. There are a number of factors limiting success including age, method of preparation, and species, but there are now many papers documenting the reliability of the results. The rich information the studies can yield include improvement in the fungal phylogenetic tree (Dentinger et al., 2015), tentatively identifying plants on Linnaeus’s Hamerby’s estate as related to his type specimens (Andreasen et al., 2014), and employing herbarium specimens in large-scale genomics research.

There are also more conventional uses of specimens that are nonetheless critical to botany. The development of floras—lists of plants in a particular locale—are impossible without using herbaria for collecting information. And when botanists are looking for living specimens of a species, the logical place to begin is in the herbarium where they can discover the location of past collections for the plant in question. The Royal Botanic Gardens, Kew (Kew) produces seed collecting guides for botanists going to areas such as Mozambique. The guides include descriptions of each species with a photo of the herbarium sheet as well as of the live plant. Specimens also can be used to hunt for other things besides plants, like diamonds and gold. The palm-like plant, Pandanus candelabrum is rare because it only grows in soils containing kimberlite which is rich in magnesium, potassium, and phosphorus. Kimberlite is also the volcanic rock where diamonds are found. So P. candelabrum, which grows in Liberia, may make it a little easier to discover these gems. As for gold, particles of the metal in Eucalyptus specimens may indicate gold deposits in the area where the trees grew. Using this link to direct prospecting is still experimental. It is definitely a long way from panning for gold, but worth a try (Lintern et al., 2013). These are very specific examples of botanical biogeography, but this is a much broader topic that I’ll discuss in my next post.

References

Andreasen, K., Manktelow, M., Sehic, J., & Garkava-Gustavsson, L. (2014). Genetic identity of putative Linnaean plants: Successful DNA amplification of Linnaeus’s crab apple Malus baccata. Taxon, 63(2), 408–416.

Dentinger, B. T. M., Gaya, E., O’Brien, H., Suz, L. M., Lachlan, R., Díaz-Valderrama, J. R., … Aime, M. C. (2015). Tales from the crypt: genome mining from fungarium specimens improves resolution of the mushroom tree of life. Biological Journal of the Linnean Society, 117(1), 11-32.

Eisenman, S., Tucker, A., & Struwe, L. (2012). Voucher specimens are essential for documenting source material used in medicinal plant investigations. Journal of Medicinally Active Plants, 1(1), 30–43.

Jarvis, C. (2007). Order Out of Chaos: Linnaean Plant Names and Their Types. London, UK: Linnaean Society.

Lintern, M., Anand, R., Ryan, C., & Paterson, D. (2013). Natural gold particles in Eucalyptus leaves and their relevance to exploration for buried gold deposits. Nature Communications, 4, 2274.

Specimen Labels: Changing Practices

Abronia glabrifolia collected by Noel and Pat Holmgren, in the Steere Herbarium at New York Botanical Garden

While taxonomists are still heavy users of herbaria, there are more and more ecologists and environmentalists using collections to document environmental changes such as global warming, species extinctions, and ecosystem degradation. This is one reason why georeferencing of label data has become crucial. It is important to know not only when a specimen was collected for studies in phenology, but also precisely where it came from. Georeferencing essentially means identifying the latitudinal and longitudinal coordinates of a location, and estimating the probability of the collection within a certain radius of that point. The more accurately the location can be determined, the smaller the radius. There are now programs such as Geolocate that aid in determining the radii, but it is still a time-consuming process that explains why most recent collections report GPS coordinates. Other collection practices have also changed, with small pieces of living material often being packed in drying gel for future DNA sequencing. Dried specimens can be used, but fresh material is much easier to work with and provides more detailed results. However, DNA analysis is one of the unanticipated uses of older specimens as well. Analysis techniques have improved greatly over the past 10 years which means that more and more specimens that are 100 or even 200 years are yielding sequence information. This is a real boon for phylogenetics, evolutionary ecology, and also for pharmaceutical exploration. Since so many drug compounds were originally found in plants, the medicinal study of plants has seen a resurgence of an interest that was significant during the Renaissance.

This use of historical material brings up another issue. Many specimens from the 19th century are darkened. They simply look dirty, but the darkness is due to treatment with mercuric chloride to prevent insect damage. This destroys the DNA in the specimens but is also a risk for those handling the plants. Dealing with this problem is one of the ways that specimen preparation and handling have changed the most over the years. While mercuric chloride hasn’t been employed for decades, it’s presence is still felt in herbaria with historical material. Curators and herbarium managers are always on the lookout for ways to minimize staff exposure and to decontaminate cabinets and storage areas. However, these specimens are too valuable to discard, particularly now when they have been in a sense rediscovered as time capsules of historical ecological information.
Today, many herbaria control insects primarily by freezing specimens at -20° for seven days or more both when they enter the herbarium and again before they are placed in cabinets. Insect traps are used in monitoring, but usually chemical treatment isn’t necessary, especially if the cabinets are well-sealed. Some smaller herbaria still rely on camphor balls in each cabinet. This doesn’t help air quality, but is effective and much less dangerous than mercury. Bringing up this topic may seem like an aside in a discussion of labeling, but there are herbaria where old specimens are stamped as having been treated with mercuric chloride to warn handlers of the danger.

I would like to end with a reminder that no matter the amount or quality of the information on a herbarium label, there is always a great deal left unsaid, a rich tapestry that underlies every specimen. The great plant collector Frank Kingdon Ward (2003) writes in his book, In the Land of Blue Poppies, about a case where he had seen a promising rhododendron. When he returned to the site: “The flowering specimen of the rhododendron I had worked so hard to get [in a remote mountainous area] had set no seed. One truss of two flowers I had taken lightheartedly for the herbarium. The other two trusses, four flowers in all, hadn’t produced a seed. I nearly wept. And then as I was on the point of giving up, hidden away in a crevice of the cliff I found one more plant. It had one capsule, and it was full of fertile seed. It had become a point of honor to collect seed of the pink rhododendron. Had it been a diamond as big and blue as the Koh-i-noor, I couldn’t have taken more care of it. May it succeed! Yet, when the plant flowers in England, connoisseurs looking at it and reading the simple label, ‘Rhododendron sp. (K.W. 9413) Burma 1931,’ will think of none of these things” (p. 138).

My next series of posts describe efforts to deal with just this issue of linking the excitement of collecting with the rich scientific and historical data in herbarium sheets.