Light and Scanning Electron Microscopy in the Digitization of Arthropods’ Biological Collections

Abstract

Among biological museum specimens, many relatively small organisms are difficult to observe macroscopically. Therefore, their attractiveness to visitors is limited. To present such small objects, various magnifying techniques can be used in combination with the digital presentation of the specimens. In this conference report, we describe how we use amalgamating techniques, along with their advantages and limitations. Our strategy uses light and electron microscopy to present small biological specimens. Light microscopy enables the observation of organisms in their natural colors but does not allow precise observation of very small microstructures, such as setae, ommatidia, and cuticle sculpture. In turn, scanning electron microscopy, thanks to its very good resolution and the possibility of obtaining higher magnifications, allows the observation of fine details of structure, but the objects are presented in grayscale. We present the scheme we use to prepare arthropods that are stored as museum specimens for digitization and subsequent presentation to visitors. Our strategy enables the comprehensive use of the specimens, as well as limiting the number of museum specimens used to prepare digital collections and reducing the number of individuals captured in the wild.

1. Introduction

Arthropods kept in natural collections are peculiar specimens that are composed of organic substances. They are exposed to degradation due to decomposition, dehydration, or attack by other organisms that feed on dead organic matter. Moreover, most arthropods are organisms that are difficult to present to visitors or students, not only because of their fragility and the resulting high probability of destruction but also because of their small size, which makes the observation of their details impossible. These details are often important for scientific, didactic, and popular science reasons. Issues such as the differentiation of legs, mouthparts, antennae, wings, and other body parts play an important role in teaching zoology, evolutionary biology, and environmental protection. Therefore, the variety of arthropods’ morphological adaptations to diverse environments is a very useful tool in biological education at the various levels of educational systems. Digital files, showing details of their structures, are the perfect complement to courses focusing on evolution, entomology, taxonomy, and systematics, as well as zoogeography and ecology [1]. In such cases, museum collections operating at scientific units such as biological science departments at universities are very often used. The usage of specimens stored in museum biological collections also limits the sourcing of specimens from the natural environment, which is extremely important since arthropod species have decreased worldwide [2]. Proper storage of specimens may allow new research. Specimens collected in the last century can be examined using the latest, previously unavailable research techniques, which can contribute a lot of new scientific data to our knowledge. Among such methods are the latest biochemical and microscopic techniques, the digitization of objects, and the possibility of presenting them using 3D techniques. 3D techniques and virtual reality may provide impressive and valuable data, attractive for visitors and students [3,4,5,6]. However, these methods are not always available.

In this conference report, we present a method based on light and scanning electron microscopy—two microscopic techniques that are quite easily available and in standard use at our university. The Natural History Collections at the Faculty of Biology, Adam Mickiewicz University in Poznań, Poland, possesses many arthropod specimens of scientific, historical, and didactical value. They cannot always be made available to everyone who wants to study them. Due to their scientific value and risk of damage, it is also not possible for the specimens to be taken outside the museum, which limits their availability for research or educational purposes.

In such cases, the digitization of collections is very helpful [7,8,9]. It has many advantages. Firstly, it protects valuable biological specimens against destruction or infestation by other organisms transferred from recurring handling. Secondly, digitization enables the use of objects in remote learning. Thirdly, thanks to the ability to enlarge objects beyond the range of light microscopy, it is possible to present enlargements and draw attention to elements of the arthropod’s morphology that are poorly visible or not visible at all to the naked eye, especially for educational purposes. Digitization and remote observations also protect valuable specimens from being destroyed when they are repeatedly taken in and out from the collection.

The COVID pandemic that began in 2019 has made the whole world aware of how important the digitization of scientific sources is in the context of distance education. Also, this type of education enables teaching in areas subject to educational exclusion, in less developed countries, in areas where wars are taking place, or in those that are recovering from armed conflicts. Moreover, image recording makes it possible to leave a huge legacy of biological data to subsequent generations and use it in the future. After all, it is thanks to recordings—analog ones—that we can observe the extinct thylacines (Thylacinus cynocephalus, Tasmanian tiger/Tasmanian wolf, Marsupialia) recorded on film tapes [10,11]. The digitization of collections enables further progress in science and teaching. In recent years, the number of publications on digitization in biology has been increasing significantly (Figure 1).

In this paper, we present data we demonstrated at the VIII Forum BioGIS (29–30 November 2023, Poznań, Poland), and describe the way we combine the usage of both environmental and laboratory observations for museum educational purposes. In our opinion, such a holistic attitude toward organisms occurring in certain environments enables us to present students and non-biologically-educated visitors with the complexity of nature and to show the importance of particular species for the biosphere. Using various types of microscopes (stereomicroscope and scanning electron microscope, SEM), we try to show how arthropods adapt to their habitats. The database will be useful at various levels of education while teaching evolutionary topics related to arthropod morphology, taxonomy, and systematics, as well as convergence and divergence in animals. Often, educators focus on vertebrates. Only rarely are these phenomena described using invertebrates as examples. Meanwhile, arthropods, with particular emphasis on insects (as the most diverse group of animals), are organisms that can be used as examples illustrating evolutionary processes. Many of them are easy to obtain from the nearest environment, where they occur in great numbers and are always at hand. In the case of many species, they can be easily bred at home or in the laboratory. Also, arthropods possess a relatively hard exoskeleton that plays important roles for the animal: preventing dehydration, protecting against predators and parasites, stabilizing the shape of the body and its internal structures, and being a place of attachment of muscles [12]. The cuticle is also beneficial for our purpose because it increases the stability of the body and prevents the body from dessication in vacuum or during storage in the museum. We present arthropods that can be used as examples of convergence to show both the unity and the variety of life on our planet. In this paper, we describe the multi-microscopic strategy used by our faculty to digitalize arthropods to increase their availability for the audience as well as to use them in education and to show the complexity of their morphology in relation to their environment and life strategies.

2. Materials and Methods

The number of scientific publications referring to “digitization” and “biology” published yearly was calculated using the Scopus database, with (title–abstract–keywords (digitization)) as the first searching criterion, and (title–abstract–keyword (biology)) as the second searching criterion to limit results of the first search only to biological samples.

Our report focuses on the strategy of using both types of microscopes in the presentation of relatively small biological specimens. Hence, no particular taxa were considered but a wide variety of them were selected. Museum entomological specimens were photographed using various digital camera models, including DSLR cameras, an Olympus SZX16 stereo microscope with an Olympus DP74 digital camera, and a SteREO Lumar V12 stereomicroscope. For scanning electron microscopy (SEM) observations, the specimens were fixed using glutaraldehyde as a standard fixative for electron microscopy. Then, the specimens were mounted on stubs with double-sided sticky tape, coated with gold, and observed with a Zeiss Evo 40 Scanning Electron Microscope (Carl Zeiss, Jena, Germany).

3. Results

The Natural History Collections at the Faculty of Biology, Adam Mickiewicz University in Poznań, Poland, stores important specimens that are interesting from the scientific, popular science, or even artistic point of view (Figure 2). Usually, because of their scientific or historical value, the specimens cannot be taken out of the showcases and they cannot be subjected to microscopic procedures related to conducting teaching activities. For this purpose, we most often use individuals caught during ongoing teaching activities and scientific research, especially by using various types of traps. Then, the animals are fixed by the rules applicable to the preparation of individual taxa. After that, the specimens are subjected to light microscopic observations, and later on, the specimens are subjected to SEM procedures. Such a strategy enables us to document the colors of the specimens. In arthropods, colors play an important role in their adaptations to habitats, reproduction, or catching prey. The colorization may be produced by bioluminescence, chemical pigments, or structural colors. Very often, metallic, blue, or green colors are generated by fine microsculpture of the exoskeleton, and they appear when the dimensions of the microstructure are close to the wavelengths of light [13,14]. We begin with the full-object pictures, using digital cameras and stereomicroscopes (Figure 3), to document the natural colorization of arthropods (Figure 4). Using this strategy, we may first observe the natural colorization of the specimens, and then, electron microscopy may explain if the colors are structural or not. In other words, light microscopy shows the colors, and electron microscopy clarifies if the colors are structural or caused by pigments.

After collecting light stereomicroscopic documentation, the specimens are transferred to SEM. Here, the specimens lose their natural colors, but they can be observed with much higher resolution than in light microscopes. SEM enables observation of the fine structures of arthropods, like setae, eyes, the microsculpture of exoskeletons that produce structural colors, and the sculpture of legs, mandibles, or eggs (Figure 5). Observation of the ultrasculpture may bring information that is crucial for explaining important features of organisms. For example, in butterflies, the structural colors of wings depend on the very fine structure of wing scales that reflect light at various angles (Figure 6). Then, the gold-coated organisms are deposited in the Natural History Collections, as a specific collection of SEM samples (Figure 7). Although covered with gold, organisms treated as pests in museums, like skin beetles (Dermestidae), may destroy them (Figure 8). Therefore, they are subject to the same rigorous preservation regulations as the typical biological museum specimens.

4. Discussion

Museums, including natural history museums and biological collections, are not only places to preserve specimens of scientific importance, but they also play other, equally important roles: they are economically important tourist attractions, and they play a crucial role in teaching, at various levels of education, as sources of specimens illustrating a process of didactics. In this view, digitization (the process of converting physical artifacts into digital pictures), and digitalization (the process of using digitization in business or sharing digital objects with the audience), become a very important part of the activity of museums and collections [15].

Thanks to a combination of both microscopic methods, we can present various features either at the organismal or organ level. Digitization allows us to present specimens to a wide range of people. For this purpose, we strongly cooperate with AMUNATCOLL—AMU Nature Collections [16,17], a web page that is dedicated to making available open information about the nature collections in the Faculty of Biology at Adam Mickiewicz University in Poznań (AMUNATCOLL). Most arthropods are relatively small organisms and digital collections enable spectators to note fine but crucial details of their structure. For example, insect mouthparts or legs present a range of adaptations to various habitats, various ways of movement, as well as various types of feeding. These organs possess numerous adaptations that are not easily observed with the naked eye or even with light microscopes, like numerous setae on feet, tips of mouthparts, etc. SEM images are very important in the description of these fine structures but electron microscopy techniques are not very accessible, and the only way to present SEM images to a wide range of people—including pupils, students, and lifelong learners, either in professional courses or in the popularization of science—is to present digitized electronograms.

Such collections may be important tools in learning in the area of educational exclusions. For example, digital collections are easily accessible, and hence, they are important for breaking barriers in education and self-development, as in the case of disabled people, by making the resources inclusive [18]. Therefore, Yap et al. [15] point out that digital displays enhance the overall museum experience for museum visitors and promote the educational aspect of museums as an ‘interconnected space’. Digital and virtual natural collections may significantly change the experience of visitors and also induce competitive quality improvement between natural history museums and natural collections, just like in the case of art galleries [19]. Easy-to-access information about the collections is also important for the cooperation of scientific units worldwide. It is also noteworthy to add that digitalization that results from the digitization of museum objects is an important point of eco-innovation [20], which, in turn, reduces the impact of humankind on the environment, and—as emphasized in EU documents—is vital in supporting the transition to a circular economy and achieving the objectives of the European Green Deal [21]. Therefore, modern biological collections play an important role in science and education nowadays. As pointed out by Massari and Netti [22], thanks to digitalization, museums become “innovation laboratories” and may become effective tools for connecting scientists and society as well as successfully promoting their collections. It is hard to estimate the time of digitization of whole biological collections. It depends on the number of people involved in the process and the number of microscopes used. The standard procedure takes approximately two days: observation and digitization using light microscopy, gold-coating (many samples at the same time) and observation using SEM. Of course, the time spent on specific structures also varies. Some more complex structures need more images than others.

The destruction of gold-covered insects by other organisms was not a full surprise, although the nanometric layer of metal may cause difficulty for Dermestidae beetles to get into the organic matter of the specimen. As we could observe, the beetles were able to destroy the metal surface and then ”dig into” the object. This suggests that the SEM collections must be subjected to the same rigorous museum preservation procedures as typical collections. Perhaps keeping them in professional SEM stub storage boxes or closed in museum vitrines, together with regular monitoring of their condition and freezing procedures, is inevitable for the preservation of samples.

5. Conclusions

We strongly suggest magnification and digitization, followed by digitalization, as a method of collecting and presenting invertebrate collections. We believe that, first of all, documentation should be conducted using a light microscope to present the image on a larger scale and preserve its natural colors, and then images should be taken using a scanning electron microscope to present details at high magnifications and in better resolution than available with light microscopy. The specimens must be stored properly so that they can be used in the future and so that they do not get damaged.