Learning Geomicrobiology as a Team Using Microbial Mats, a Multidisciplinary Approach

Abstract

Microbial mats are one of the best suited laminar organo-sedimentary ecosystems for students from different educational backgrounds to visualize the direct relationship between microbes and minerals. We have used tropical hypersaline microbial mats from Puerto Rico as educational tools to promote active learning of geomicrobiology introductory concepts for undergraduate students organized in multidisciplinary teams with biological and geological backgrounds. Besides field trips and independent research projects focused on microbial mats, four intensive workshops and one capstone activity were designed to expose students to the different geomicrobiology subdisciplines (microbiology, molecular biology, geology, and geochemistry). The teaching-learning process was assessed using pre- and posttests, group discussions, activities including Gallery Walks and exquisite cadaver’s, case studies, and focal interviews. While the posttest showed a significant difference in conceptual understanding, the Gallery Walk and the capstone activities demonstrated increase in the depth, coherence, and thoughtfulness in answering questions, including a clear integration of the different subdisciplines during their presentations. Finally, the main themes described by the students as important outcomes of their participation in the Research at Undergraduate Institutions: Microbial Observatory (RUI-MO) program were: (i) the opportunity to study and learn new and different science disciplines, (ii) the microbial mats were excellent tools to learn from and integrate different science disciplines, and (iii) working in multidisciplinary teams gave them the opportunity to learn from their peers’ discipline backgrounds. To our knowledge this is the first educational initiative that uses tropical hypersaline microbial mats to teach geomicrobiology in a multidisciplinary fashion.

Geomicrobiology focuses on the role of microorganisms in the geological transformations that take place at the interface of Earth’s biosphere and lithosphere in space and time (5). Specific transformations in which microorganisms serve as mediators of geological processes include: alteration of geochemical microenvironments through redox reactions (e.g., using O₂ and producing H₂S), precipitation and dissolution of minerals (e.g., the CaCO₃ deposits), and solubilization and dispersion of insoluble metals (e.g., Fe(III)-oxide reduction) (7, 12). Microbes also produce biomass and exopolymeric substances (EPS) that help stabilize sediments (12). Production of EPS is particularly important in the cyanobacteria-dominated biofilms that we commonly refer to as “microbial mats.” These organosedimentary structures are not only considered analogs of the oldest (2) but also the most productive ecosystems on Earth (9).

Microbial mats are indeed the quintessential systems for important geomicrobiological studies (2, 9, 12). As these studies are based on interactions of microorganisms and their geochemical environment through the Earth’s history, investigations in geomicrobiology span vast temporal and spatial scales. It is indispensable that students introduced to this emerging field first understand the specific physicochemical conditions of the ecosystem (such as light and nutrient availability, temperature, pH and oxygen content, and sediment composition). Only when students fully understand the range of these environmental conditions, can they identify the role of the microbial component and its potential to alter, precipitate, dissolve, and accrete the different minerals. Studies in geomicrobiology are interdisciplinary in nature and students need to learn how to work as part of a multidisciplinary team. Indeed that is the main educational goal of our work, to use mats as tools to promote active and effective learning of basic concepts in geomicrobiology within a multidisciplinary team. If our students are able to visualize the link between the geochemical and biological components of a microbial mat, then they will be able to use the mats as a tool to conduct a successful multidisciplinary study with peers from other disciplines. This strategy of team work using a multidiciplinary approach has been recommended by the National Research Council (NRC) as one of the main priorities in the new educational reforms at the undergraduate level (11).

To promote the interaction of our students, we assembled a series of multidisciplinary teams that were requested to perform a series of activities. These activities ranged from traditional (i.e., participation in workshops and oral presentations) to nontraditional such as field approaches to one specific site in Puerto Rico named Cabo Rojo. This location contains various types of interesting microorganisms as well as microbial mats (3, 4). Students comprising the multidisciplinary teams were requested to investigate these ecosystems by conducting extensive microbiological, geochemical, and geological analysis of them using the main strengths of each member of the team.

Description of the program. One of the main goals of the Cabo Rojo Salterns (CRS) NSF-sponsored Research at Undergraduate Institutions (RUI) : Microbial Observatory(MO) program is the training of undergraduate students in geomicrobiology (http://www1.uprh.edu/salterns/). For four consecutive years we have used the microbial mats from these salterns as a “living” laboratory. Undergraduate students from the biology and geology programs of the University of Puerto Rico (UPR) and Turabo University (TU) have interacted here to understand basic concepts of geomicrobiology. One of the main strengths of the program is the collaboration with the geosciences and marine sciences programs of the University of Connecticut through which our students interact with faculty and graduate students during two field trips each year. The rest of the academic year, our undergraduate students perform a series of individual research projects in their main universities until they receive their degree. Most students enter into a graduate program and pursue a master or doctoral degree in different fields of the biological sciences.

METHODOLOGY

Fourteen students from the Biology departments of the UPR-Humacao, Mayagüez, Bayamón, as well as from TU, and the Geology department of the UPR-Mayagüez were trained in the field of geomicrobiology during academic year 2005–2006 under the CRS-RUI-MO program. Four multidisciplinary teams were formed, and it was imperative that each team included students from at least three different fields. The training consisted of four intensive workshops and one capstone activity designed to expose students to the different subdisciplines that span the geomicrobiology science field: microbiology, molecular biology, geology, and geochemistry. All these activities used the Cabo Rojo saltern tropical hypersaline (CRth) microbial mats as a central educational and research tool (Fig. 1, summarizes the activities and aims of the program).

FIG. 1.

Summary of the educational backgrounds of the Microbial Observatories program participants and the main workshops offered during 2005 and 2006. The letters M, G, and B represent students from microbiology and marine biology, geology and biology backgrounds respectively. Please note that microbial mats are the central educational tool of all the educational activities.

Geomicrobiology multidiscipinary workshops

Workshop 1. Cultivation of microbial communities from the mats using general microbiology culture techniques. During the first field trip the students received a 30-minute talk about general microbial physiology concepts and the influence of physicochemical parameters such as light, oxygen, and salinity on the growth of the different populations in the CRth-microbial mats (3). The main geomicrobiology subdiscipline question of this workshop was: “which specific microbial communities can be isolated from the CRth-microbial mats based on the environment where they are located?” After the talk, multidisciplinary teams were assigned to sample specific CRth-microbial mats and determine environmental physicochemical factors such as temperature, light, pH, as well as dissolve oxygen and nutrient contents. The teams were also responsible for dissecting and culturing CRth-microbial mat samples in petri plates using culture media with different salinities and pH. Some plates were incubated at different temperatures or put into an anaerobic jar (13). As the different organisms take time to grow, results from this workshop were presented at workshop 2.

Workshop 2. Using molecular techniques for the identification of microbial groups in the CRth-microbial mats. Our second workshop consisted of two intensive days where the students were first introduced to the organisms able to grow under such conditions and then received a short talk about DNA extraction methods, electrophoresis, restriction enzyme analysis, cloning, and PCR. In addition to introducing the general concepts of genetic engineering, the talks focused on how to apply those technologies and techniques in order to identify the nonculturable CRth-microbial communities.The main geomicrobiology subdiscipline question of this workshop was: “can we determine and/or detect specific microbial groups present in the different layers of the CRth-microbial mats using molecular techniques?” The students were organized into four multidisciplinary teams and received sections of the top (green) and bottom (black) layers from the CRth-microbial mats with a set of primers specific to cyanobacteria (8), archaea (7) and bacterial 16S rDNA (1). Each team was responsible for extracting DNA from the mats and determining the presence of members from the different groups in the distinct mat layers.

Workshop 3. Mineral characterization in the organo-sedimentary layers of the CRth-microbial mats: their sources and genesis. Teams of students were exposed to some of the minerals present in the mats, and how microorganisms play an important role in mineral formation (4, 5, 12). The main geomicrobiology subdiscipline question of this workshop was: “Are minerals present in the CRth-microbial mats the result of specific biological activities, binding and trapping of the mats, or other geological processes?” The workshop started with a 1-hour talk, then the students were able to manipulate different types of rocks and minerals commonly found in the CRth-microbial mats.

Workshop 4. What is it that we need to know and how do we measure it? In situ geochemical assessment of geomicrobiological processes in the CRth-microbial mats. In this workshop the teams were introduced to a holistic approach typical for an in situ geomicrobiological investigation. The concepts of coupled microbial populations, metabolic rates, and mineral products were introduced, and as a snapshot of the system, students learned to measure how the oxygen and sulfide profiles change across the different CRth-microbial mats (3, 12). The main geomicrobiological subdiscipline question of this workshop was: “Can oxygen and sulfide depth profiles help us assess the microbial communities and their activities in each layer of the CRth-microbial mats?” After learning how to operate the microelectrodes and conduct in situ analyses, each multidisciplinary team measured the geochemical gradients (including physico-chemical conditions, pH, salinity, and light), constructed depth profiles of oxygen and sulfide for the various mats, and plotted and analyzed the data (14).

Geomicrobiology capstone activity—case studies: how to perform a geomicrobiological investigation in other microbial mats, putting it all together. After being introduced to many concepts basic to a geomicrobiology study ranging from molecular microbiology to geology, multidisciplinary teams were asked to do a specific case study. They had to formulate a project description for the following field sites: (i) microbial mats in the Dry Valleys, Antarctica, (ii) microbial mats in the Iron Mountain Mine, California, and, (iii) biofilms in the deep biosphere, ca. 400 m below the sediment surface, off the coast of Perú.

Each group of students was provided with an information sheet that included some useful facts about each study site including pictures, specific instructions, and main goals of the activity. In addition, colored markers and poster boards were given to each group to prepare their oral presentation. The main goal of the activity was for the groups to produce hypothesis-driven predictions about the relevant geochemical factors present in the environment and the microbial communities that could be present due to those factors. The students needed to describe strategies for successful enrichments of these microorganisms and how to analyze specific biogeochemical signatures expected to be present in the ecosystem. Students were assisted by the CRS-RUI-MO research investigators.

Assessment at the Microbial Observatory. Several assessment strategies were used to determine the student learning process during participation in the Microbial Observatory projects. All assessment measures were performed by both investigators associated with this project and the educational coordinator of the program. Assessment for the first and second workshops was conducted through a pre- and posttest, exquisite cadavers, and group discussions in which the students shared their knowledge about the organisms per layer and their associated physiologies. A modified Gallery Walk was employed in the third workshop and in the fourth we used group discussions. A case study was used as the final approach to coherently measure integration of the knowledge acquired by students. In addition a focus group interview helped to explore the perception of the students regarding their learning process through the program.

Pre-and posttest. The tests were given to assess student’s knowledge and understanding about molecular ecology, particularly the molecular techniques needed for the identification of the most relevant microbial groups in the mats. The pre- and posttests were identical and consisted of 21 questions which included multiple-choice, matching, true-or-false, and short answer questions for a total of 29 points. For the true-or-false questions, students had to identify the word or phrase that made the false statements false. A paired t test was performed to compare student’s knowledge before (pretest) and after the workshop experience (posttest) and to determine any significant differences. The results are shown with box and whisker plots.

Exquisite cadaver. Exquisite cadaver, an educational tool, is a game in which students summarize their ideas about one specific topic. In our case, the general theme was microbial mats, and the students were organized in four multidisciplinary teams. Initially we requested that one student from each team write a sentence on whatever came to mind about one specific topic (e.g., Cabo Rojo mats). Once the student finished writing, he folded the paper and wrote the last word of his statement on a new space. The next student used this word to start a new sentence. After all the students on the team finished writing their sentences, the whole document was edited in order to make a paragraph. The final document is named the exquisite cadaver and shared with the others at the end of the activity. The exquisite cadaver is a collective of several sentences that are all linked by one main idea. The activity was derived from surrealistic painters and the technique they use to put different ideas into one specific work of art. Our students used this activity before and after workshop 2.

Individual versus group Gallery Walk. Gallery Walk is a discussion technique that promotes active engagement while students work in teams to synthesize information written from a variety of perspectives (6, http://serc.carleton.edu/introgeo/gallerywalk/index.html). We modified this technique by first asking our students four questions after workshop 3. Then, the multidisciplinary teams rotated around the conference room to answer the same questions, which were posted on pieces of paper located at different stations (6). Each group had 3 minutes to answer each question and after rotating through the different stations the students returned to their original station to write a summary. The activity ended with an informal oral report in which each group synthesized comments to the particular question assigned. All questions were framed according to Bloom’s hierarchy into knowledge, comprehension, and application (10). Answers given by students were compared with those given by the mentors according to their depth, coherence, and completeness.

Group discussion after generating the profiles. Once depth profiles for oxygen and sulfide from the CRth-microbial mats were generated, each multidisciplinary team gave an oral presentation to discuss their findings. Microbial Observatory mentors evaluated how the students related the knowledge obtained from the profiles that they generated with the microbial communities and their physiologies in the mats.

Geomicrobiology capstone activity: case studies. Two different strategies were used for the assessment of student learning through the case studies, after the oral presentations and construction of the posters. The first one measured the team performance based on a rubric (Table 1). The second was based on open-ended questions answered by the MO mentors and used to analyze and evaluate their perception of team integration of the workshops into geomicrobiology, and performance during the capstone activity.

TABLE 1.

Scoring rubric developed by the Microbial Observatories mentors to evaluate the performance of the students after their poster presentations of the case studies

Components	Scoring criteria
	3	2	1
Knowledge	The concepts and principles used for the experimental design are appropriate to the study. The student’s response reflects thorough understanding.	The concepts and principles used for the experimental design are appropriate to the study with no significant errors.	The student explains the experimental design but misapplies some concepts or principles, or omits some facts that are important for understanding the study.
Scientific method	Use scientific method to pose the project’s hypothesis. The experimental design addresses all important questions raised by the prediction.	Use scientific method to pose the project’s hypothesis. The experimental design addresses the most important questions raised by the prediction.	Partially use scientific method to pose the project’s hypothesis. The experimental design addresses some important aspects of the prediction, but omits others.
Integration of different subdisciplines	The experimental approach to understand the field site is truly multidisciplinary.	The experimental approach to understand the field site crosses the disciplinary boundaries to some extent.	The experimental approach to understand the field site does not show a multidisciplinary character at all.
Creativity in the experimental design	The students present the information in a way that reflects creativity in the experimental design.	The students present the information accurately but with minimal creativity.	The students present the information with little or no originality.
Clarity of the presentation	Presentation is clear and to the point. Ideas and opinions are well prepared.	Presentation is somewhat clear. Some ideas and opinions need to be refined.	Presentation is not well organized. Thoughts and opinions are unclear.

Focus group. Upon completion of all MO workshops and activities, a focus group interview was held to collect and explore shared understanding from students about their views and learning process during participation in the program. One of the main objectives of the focus group was to determine the students’ perceptions upon completion of the different geomicrobiology multidisciplinary workshops. Five students participated in the focus group interview. All interviewees completed an informed consent form before beginning the interview. This consent form was approved by the University Development Office, Institutional Investigation Area, UPRHumacao. The interview was organized and conducted by the educational coordinator of the Microbial Observatory.

RESULTS

Assessments of the Microbial Observatory workshops

Pre- and posttest and group discussion. There was a significant difference in conceptual understanding about the microbial community of the mats and the techniques to study them after student’s participation in the first workshops. Results from the pre- and posttest conducted during workshop 2- (paired student’s t test (9) = −6.48, p < .001) indicated a difference mean value of −11.3 and an increase in median values (of correct answers) from the 15th to the 25th percentile (Fig. 2).

FIG. 2.

Box-and-whisker plot showing median of 15th and 25th percentile of positive responses in the pre- and posttests given during Workshop 2. Please notice the differences in the vertical lines indicating a more uniform distribution in the responses of the posttest.

Our exquisite cadaver activity also revealed an increase in students’ knowledge. Before the second workshop students were more oriented to microorganisms and the geologic content of the mats. The documents after the workshop were more oriented toward the molecular aspects of the mats (data not shown).

After completion of the workshop, the discussion of the interdisciplinary teams was evaluated using a rubric (not shown) with a scale from 1 (poor) to 3 (good) based on three criteria: (i) technique proficiency, (ii) relevance to the CRth-microbial mats, and (iii) overall presentation clarity. After their presentations, the average scores for the teams were 2.5, 2.5, 3.0 and 2.25. The global presentation average score was 2.6.

Individual versus team Gallery Walk. Each investigator compared the answers provided by the students once they went through the gallery individually and as a team. The CRS-RUI-MO research investigators detected an increase in the depth, coherence, and completeness of the answers given by the teams over those given by individuals. Such improvements were a direct result of the active discussions of the students on the team. Contrary to their individual assignments, students participating in the Gallery Walk were able to defend, exchange, and rethink their final answers. Improvement in the final answers provided was independent of the level of abstraction used (i.e., knowledge, comprehension or application). For example, for a knowledge-level question individual students were unable to discuss specific concepts, they just copied information from the written presentation. During the team Gallery Walk students not only mentioned specific concepts but were capable of discussing them. Similar results were reported in the comprehension-level question where students listed more reasons for their analysis; their answers were more coherent and thoughtful as teams. Also, when students were asked to apply the knowledge they had acquired, only during the team Gallery Walk were the relevant techniques required for mineralogy analyses and the detection of biogenesis in mat samples mentioned.

Measuring and constructing geochemical depth profiles. All teams measured and constructed vertical depth profiles for oxygen and sulfide from the CRth-microbial mats (3, 14). In order to accomplish this, the students were first taught the underlying electrochemical principles of microsensor construction and operation, then learned how microbiology affects geochemistry and that key biogeochemical characteristics of sediments can be determined by depth profiles of oxygen and sulfide (12, 14). All of the teams performed an abbreviated system calibration, measured the geochemical profiles, learned the major factors (microbiological composition, light, salinity, and temperature) that determine the oxygen and sulfide profiles, and converted the raw data into actual values. During the exercise, students were able to assess the key factors, including microbial and physicochemical characteristics of the mats. At the end of this workshop, the students could explain all of the observations, and also had a clear understanding of the significance of the geochemical snapshot in the context of the microbial mat functioning.

Geomicrobiology capstone activity: case studies. The oral presentations given by the students at the end of the capstone activity were summarized in three main posters. Each presentation consisted of a hyphothesis, predictions, and tests needed to conduct a full geomicrobiology study in each location (Fig. 3). The individual team’s rubric global average scores (Table 1) after evaluation of the posters from each team ranged from 1.9 (63%) to 2.66 (89%). The results obtained globally by category indicated that the highest scores were given to the integration of the different subdisciplines (89%) and the lowest to the knowledge (74%).

FIG. 3.

Schematic of the poster presentation for case study 2 in which a multidisciplinary team described the communities and biogeochemical signatures expected from the microbial mats present in the Iron Mountain Mine, California. As assigned on the information sheet, the team divided their presentation into observations at the site, hypothesis, predictions, and tests to be conducted to prove the hypothesis.

Based on the second assessment strategy, the overall perception of the MO mentors was that the multidisciplinary teams worked well, as evidenced by the quality of their poster presentations. Each team member contributed information and insights from his area of specialization while at the same time understanding enough of the basics from the fields outside of theirs.

Focus group. During the focus group, students shared their experiences as participants of the CRS-RUI-MO program. From the six brief open-ended interview questions, three emerging themes were related to learning a multi-disciplinary discipline, geomicrobiology, as they work in multidisciplinary teams: (i) the CRS-RUI-MO program gave the opportunity to study and learn new and different science disciplines, (ii) the microbial mats were excellent tools to learn from and integrate different science disciplines, and (iii) working in multidisciplinary teams provided an opportunity to learn from their peers.

DISCUSSION

We have presented evidence that microbial mats are excellent educational tools that can be used by multidisciplinary teams of students from biology and geology backgrounds to learn more of the geomicrobiology field. A series of teaching and learning strategies were assembled using these mats as the central piece. Initially we took advantage of the incredible microbial diversity within these productive ecosystems to teach the students how to grow organisms from the mats by placing them under different growth conditions. Once the students understood how to culture some of the communities from the mats, we taught them how they are able to determine the identity of cultivatable and noncultivatable organisms by generating genomic libraries using group-specific primers.

The importance of the mats as platforms for several geological transformations was addressed in the third workshop. The students were exposed to the main techniques used by geologists to determine the biogenicity of specific minerals in sediments, such as the ones surrounding the Cabo Rojo salterns, and their implications not only in the formation of the early Earth but also in the possibility of extraterrestrial life. These enticing topics served to captivate the students who at the end of the talk manipulated some of the most abundant minerals on the planet. However, one of our most interactive learning techniques was the generation of oxygen profiles by the students. Each student was in charge of taking in situ measurements of the oxygen content within the mats at different depths. Once they assembled all the data needed, students generated their own profiles and gave an oral presentation describing their findings. A final capstone activity challenged the students to assemble all of their acquired knowledge and provide hypothesis-driven proposals for how the metabolisms associated with the microorganisms in the location and how to analyze them in order to gain more knowledge about the environment. Students had to apply what they already knew about the Cabo Rojo ecosystem and the microbial mats, which environmental conditions, organisms, climate, seasons, day/night, etc., play a role to understand the field sites assigned to work on and then formulate their project description. The students were motivated by the different investigators to mentally revisit previously performed experiments and the workshop taken at the observatory, and based on the knowledge acquired through the CRth-microbial mats, use them as dissecting tools to critically analyze and understand the given environmental system. The effectiveness of this activity was also evident and sustained by the fact that the teams communicated ideas related to the different subdisciplines that comprise geomicrobiology (molecular science, microbiology, geology, etc.) in terms that both their peers and research investigators understood.

According to our qualitative and quantitative data, the students were able to understand the CRth-microbial mat system and integrate the knowledge from the different sub-discipline perspectives. The improvement observed in the posttest and team discussion scores, and the generation of coherent answers at the Gallery Walk proves their acquisition of knowledge and skills development during the workshops. Also, the students were able to verbalize during the focus group interview how important the mats were as an integrative ecological system and as an educational tool. Some of the main facts described by the students after these pedagogical activities included the relevance of the CRth-microbial mats as a tool to understand microbial diversity and the importance of team work.

Promotion of discussion among students from different disciplines was indeed a main goal of our study. To specifically determine how group discussions were linked to a meaningful learning of the multidisciplinary teams, we compared the answers provided by individual students and groups during a Gallery Walk activity. As previously reported, this technique promoted discussion among the students who were then able to provide more complete and coherent answers. To our knowledge this is the first time the technique was applied to discuss topics related to the geomicrobiology field.

The ultimate goal of our program was to expose our students to novel educational challenges as future professional alternatives by exploring emerging disciplines such as geomicrobiology. Although such statistics are beyond the goals of this publication, we currently have five former students of our program that are pursuing a graduate degree in geomicrobiology. In addition we have offered the first undergraduate course in topics of geomicrobiology within Puerto Rico. Both indicators of the success of the CRS-RUIMO program have in common a single location: the Cabo Rojo tropical hypersaline microbial mats.