Understanding habitat heterogeneity and habitat preference are central tenets of undergraduate ecology courses, but many urban campuses, particularly community colleges and others with large student enrollments, may lack the means for extensive field sampling and monitoring. We outline the use of diatoms living in decorative water fountains as a proxy for field sample collection, data analysis, and ecological interpretation. These methods are amenable to undergraduate laboratory courses and independent student research projects.
The teaching recommendations outlined in Vision and Change in Undergraduate Biology Education (AAAS, 2011) call for stronger emphasis on applying the process of science, developing quantitative reasoning, and providing meaningful research experiences for students. However, many urban campuses, like ours, are predominated by commuter and nontraditional students and may service hundreds or thousands of declared biology majors every semester. Additionally, metropolitan campuses may not have access to natural environments. These conditions present challenges when coordinating field-based sampling for a variety of ecology-themed courses. One method that we have employed for teaching students field collection, data analysis, and quantitative reasoning is to introduce the concepts of biodiversity, species richness, and species evenness using cars in the commuter parking lots as proxies for organisms (see Hodgson, 2015).
More recently, to provide meaningful experience in sampling living organisms in the field within a guided inquiry on habitat heterogeneity and habitat preference, we developed a protocol that uses diatoms living in decorative water fountains (located on our campus) for independent student research projects. Urban locations are often analogous to natural environments as habitats and satisfy niche requirements for a variety of organisms; likewise, their structural complexity creates habitat mosaics that can be used to understand habitat use and patchiness, population ecology, and metapopulations (Lundholm & Richardson, 2010). The methods described herein are also applicable for classroom exercises ranging from a few days to several weeks in length.
Diatoms (Bacillariophyceae) are unicellular algae characterized by their silica cell walls. They are nearly ubiquitous, with assemblages of both cosmopolitan and fastidious species (Wehr & Sheath, 2003). In fact, their autecology is frequently used to determine environmental quality and to analyze habitat preference and suitability (Wetzel, 2001). Additionally, their silica cell walls greatly aid in sample preparation for microscopy and species identification (Battarbee, 1986).
Biofilm samples of diatoms collected from natural and undisturbed environments often contain a very large number of species, including many non-diatoms, and require extensive cleaning and preparation prior to microscopy. However, the samples we collected from decorative fountains on our campus possessed only a few species and required minimal preparation, making them more suitable for undergraduate inquiry. Additionally, the smaller number of diatom species living in the fountains (compared with natural environments that may have >100 species) makes identification and the use of taxonomic keys easier for students and helps build their confidence.
If decorative water fountains are unavailable for sampling, diatoms are often abundant in other campus and urban settings. Our exploratory sampling revealed that diatoms are present on leaking water valves, rooftop drains, downspouts, air-conditioning cooling towers, and birdbaths (Figure 1). Essentially, diatoms are usually found anywhere there is a quasi-consistent source of water and light. Many diatoms have subaerial niches, grow attached to moist environments called “wetwalls,” and survive periods of desiccation (Lowe et al., 2007). We ultimately chose fountains for our methods because they were more environmentally stable from day to day, but sampling the other environments listed will provide similar experiences for students.
Having students repeatedly sample diatoms from the fountains helps them understand the scientific process of specimen collection, sample preparation for microscopy, data analysis, and interpretation. In turn, this process elucidates habitat preference and habitat heterogeneity. Depending on environmental factors and the length of the study, it may be possible for students to analyze the concepts of colonization and succession. Furthermore, they gain practical experience in field ecology, practice using taxonomic keys, and develop identification skills. This model can be applied by non-diatom biologists and by a variety of biology instructors versed in the basic mechanics of taxonomic keys.
We selected two water fountains that had different constructions (Figures 2 and 3) and flow patterns. They were within 300 m of each other and were supplied by the same municipal water source. Our analyses showed that pH and water temperature were similar between sites (pH consistently 8.0 ± 0.2; temperature range: 20–28°C). However, substrate and water flow differed. This provided an opportunity to determine habitat suitability for species common to the area, with fewer confounding environmental variables. Both fountains were plumbed primarily with PVC, and no copper was visible. During our sample period, the fountains were not cleaned, chlorinated, or altered in any way by maintenance staff.
Fountain 1 had central jets; water flowed through channels leading away from the jets and then circled the entire fountain. This setup provided two sampling environments; we sampled diatoms attached to the rocks nearest the jets, which simulated an epilithic environment (“rocks”), and also from within the outer metallic channels, which simulated a lotic environment (“stream”). Water inundated the spaces between the rocks, and there was no mortar or sealant between them.
Fountain 2 had central jets and nonflowing water contained within a pool. This setup provided two sampling environments; we sampled diatoms attached to the inner metallic ring containing the jets, which simulated a waterfall and/or scouring environment (“inner ring”), and also from the outer metallic shelf, which simulated a mist zone with temporal periods of desiccation (“outer metal wetwall”; cf. Lowe et al., 2007).
We scraped visible biofilms with a metal spatula and used these smears to make microscope slides; exploratory analysis did not detect diatoms living suspended in the water column, so we discontinued open-water sampling. Over the course of two summer months, spanning Julian days 130–179, we took one sample from each “habitat” within each fountain once per week. We sampled the “stream” and “rocks” habitats in Fountain 1 and the “outer metal wetwall” and “inner ring” in Fountain 2. Diatoms often have fast generation times (e.g., 15–18 hours), so samples could alternatively be taken daily over a shorter total period (Wetzel, 2001).
Two slide-preparation techniques are available, depending on your teaching needs. If the students will be able to view the slides immediately, a wet mount can be made with plastic or glass coverslips. When making wet mounts, we found that using 0.25 g of biofilm wet mass smeared on a coverslip and then diluted with one drop of water from a disposable transfer pipette worked well. This method is also recommended if 400× bright-field microscopy will be used. Alternatively, if the slides will be analyzed later, and/or if 1000× oil-immersion bright-field or phase-contrast microscopy will be used, the 0.25 g wet mass smears should be thinly spread onto glass coverslips, dried in an oven, and then permanently mounted on slides using a resin with a high refractive index, such as Permount or Naphrax (Battarbee, 1986). Phase-contrast microscopy with permanent mounting is preferred because water and diatoms have similar refractive indices (Battarbee, 1986).
The goal of the sampling methods is to elucidate habitat suitability by using proportions of diatoms living in each location. Presence, absence, and differing proportions of diatoms between habitats are sufficient for making broad comparisons in autecology and niche ecology (Batarbee, 1986; Wetzel, 2001). We counted 300 total diatom individuals from each sample “habitat” per Julian day and used those raw numbers to calculate the relative abundance of each species. Relative abundance was calculated by dividing the count of each species by 300. For example, the “stream” in Fountain 1 had 269 Nitzschia palea on Julian day 130, giving it a relative abundance of 0.897 (Figure 4). Usually, accumulating 300 total individuals was achieved from a single slide, but additional replicate slides were occasionally needed from each “habitat.”
To facilitate and streamline this learning exercise, we examined the proportions of diatoms per sample, and not standardized numbers per area or habitat. This method eliminates the need to quantify sample biomass, dilution factors, and total sample area when using diatoms to make ecological interpretations (Battarbee, 1986). There will be sampling error when not quantifying the sample area or habitat, but Battarbee (1986) indicates that relative abundances calculated from a minimum of 300 diatoms are sufficient for broad conclusions. Given that millions of algal cells can inhabit a cubic centimeter, the relative abundance of diatom species from each location is far more important than establishing a ratio of total cells to total habitat area. If instructors wish to provide their students with more in-depth analyses, including biovolume or species per area in differing habitats, Wetzel & Likens (2000) provide appropriate protocols.
Diatom species were identified with standard taxonomic keys, but recommended ones for inexperienced undergraduate students and non-diatom experts include Vinyard (1979), Wehr & Sheath (2003), and Spaulding et al. (2010). These particular keys have many pictures and visual descriptions of the morphological attributes that will reinforce identifications and build student confidence. Identifying diatoms to the species level is time consuming, but the reduced number of species in urban settings facilitates the process. Students and instructors can usually expect to learn the taxonomy of local diatom species in a few days or weeks. Based on our experience, a previously unencountered species can be keyed out in a few hours or less. If instructors are able to identify most species ahead of time, guided inquiry and instruction will expedite the activity. Furthermore, instructors may choose to construct their own picture-based dichotomous keys using only local species. Alternatively, identifying diatoms to the genus level would still be useful for making more generalized habitat and environmental determinations applicable to the teaching objectives.
Suggested Approach & Points of Emphasis
The overall objective of this teaching exercise is to emphasize the process of science and quantitative reasoning, in order to understand the ecological concepts of field sampling and habitat preference. Therefore, when teaching this to your students, we recommend reinforcing the importance of creating a consistent sampling schedule, uniform sampling protocols, and careful documentation of all methods within a field journal. Students should also measure water temperature and pH every time diatoms are collected. These probes are inexpensive and easy to use. Salinity measurements require more expensive meters, and their results may not be important in freshwater fountains, but salinity could be measured as well. Samples should not be taken from the same scrape zone each time, so it is valuable to have the students draw a map of the fountains and document where samples were collected.
Students should understand the habitat heterogeneity of their fountains and the habitat preference of the diatoms living in the fountains. Most diatoms have well-documented autecologies that list their preferred substrates, temperature, salinity, and pH (Battarbee, 1986; Wetzel, 2001). For example, Round et al. (1990) have extensively categorized diatoms as planktonic (floating in water), epilithic (attached to rock), epipsammic (attached to sand), epipelic (attached to silt), or epiphytic (attached to plants). Additionally, some diatoms are capable of directed movement. Therefore, guided inquiry should include students making predictions of which diatom autecologies they expect to find in the fountains, based on the natural habitat analogues the fountains approximate. Furthermore, different areas of the fountains will provide different growing conditions, and students should be able to make comparisons of diatoms within the fountains. Another excellent opportunity that may be available if fountains are ever cleaned and/or turned off is to study colonization and succession.
Ecological interpretations depend on statistical treatments, so instructors will need to determine the level of appropriateness for their students. Statistical analyses, particularly analysis of variance (ANOVA) and other iterations of the general linear model, may be beyond the expertise of lower-level students. It may be helpful for the instructor to calculate ANOVA for the students and describe the meaning and significance to them. Alternatively, the instructor could create a template spreadsheet that automatically calculates statistics for the students as part of a guided analysis. Furthermore, simply creating scatter plots or histograms showing changes in diatom relative abundance will still be effective for the students in elucidating community patterns and changes through time.
We present results from one sampling regime as a demonstration and template for future exercises based on this protocol. Results will undoubtedly be different across fountains and campuses. However, the overall objective is to predict which diatom autecologies will be abundant, based on the abiotic environment of the fountains; identify the diatoms present; and compare the data to ecological predictions.
Our two fountains had four “habitat” regions (Figures 2 and 3). We expected benthic and planktonic diatoms to be present in both. Within Fountain 1, we predicted that epilithic and stream species would be abundant. Within Fountain 2, we expected species tolerant of scouring near the inner jets and species tolerant of desiccation to be present on the outer metal wetwall.
In total, three species were identified; all three were present in Fountain 1, and only two were found in Fountain 2 (Figure 4). Planothidium lanceolatum is a benthic diatom that lives attached (nonmotile) to a variety of substrates and prefers alkaline pH (Wehr & Sheath, 2003; Spaulding et al., 2010). Also, Planothidium and its basionym genus Achnanthes have several subaerial species (Lowe et al., 2007). Pinnularia saprophila is benthic but does not attach to substrates and is capable of moderate motility (Spaulding et al., 2010). Additionally, P. saprophila prefers acidic water but is tolerant of basic pH. Nitzschia palea is benthic and attaches to rock substrates, is capable of motility, and prefers alkaline pH (Wehr & Sheath, 2003; Spaulding et al., 2010).
Diatom relative abundance corresponded with predicted habitat preference. The water in the fountains was consistently alkaline (pH = 8.0 ± 0.2), providing suitable habitat for P. lanceolatum and N. palea, which were the two most abundant species. Furthermore, P. lanceolatum can tolerate subaerial environments, which explains the abundance of individuals growing on the metallic wetwall of Fountain 2. In contrast, P. saprophila is tolerant of alkaline water but prefers acidic conditions, which explains its consistently lower abundance. Nitzschia palea is capable of attaching to rocks, which explains its presence in Fountain 1 and its absence in Fountain 2, since rocks were present in the former and absent in the latter. We cannot explain why N. palea was more abundant than P. lanceolatum in Fountain 1, but we hypothesize that the motility of N. palea allowed it to colonize more of the fountain and/or escape areas of turbulence that the nonmotile P. lanceolatum could not. No planktonic species were present, which suggests that the turbulent water in the fountains was unsuitable. We were unable to elucidate any relationships with temperature, scouring, stream channel, or season with the small number of species present, but future studies over a longer period may do so.
The species abundance in these four areas within two water fountains provided an opportunity for students to use quantitative reasoning to analyze habitat preference and habitat heterogeneity. These simplified systems reduce the number of variables and streamline the inquiry and analysis procedures but give students practical skills in conducting ecological research.
Assessment of student performance should focus on evaluating their application of the scientific process. Instructors should grade the students' habitat descriptions, sampling methods, field-journal accuracy and thoroughness, slide preparation, taxonomic identification, data management, and data analysis. For example, if preliminary diatom identifications do not match their cited habitat preference, students should double check their taxonomic process, site descriptions, and abiotic measurements. Students should understand the connection between physical habitat, habitat preference, and species presence.
We encourage formal presentation of the results. It is appropriate for students to write a research manuscript with an introduction, predictions, methods, results, discussion, and relevant citations defending the interpretations. This allows for grading them in a way similar to a peer review of a scientific manuscript. Alternatively, students could present their findings via poster or oral presentations to the class or at a student research conference. The grading rubrics created by the Center for Teaching Excellence at Cornell University (http://www.cte.cornell.edu/documents/Science%20Rubrics.pdf) for assessment of research papers, laboratory reports, and oral presentations are effective and appropriate for this project.