Bacteria and protozoa are critical components of aquatic and terrestrial ecosystems, driving biogeochemical processes including carbon fixation, oxygen production, nutrient cycling, and break-down of anthropogenic contaminants. In many habitats, the bacterial community structure and net production are controlled in a top-down fashion by protozoan predation; in turn, predation of protozoa by larger animals mobilizes nutrients and contaminants to higher trophic levels. Despite their importance, relatively little is known about the diversity, biogeography, and ecosystem function of microbial eukaryotes including protozoa, due to limitations of both traditional culturing methods and genetic techniques. New instrumentation is needed to sample, study and understand a broader range of protozoa in natural systems. The objective of this research is to develop a microfluidic field sampling and analysis tool to study the biogeography and function of microbial eukaryotes in natural habitats. Microfluidic samplers will be fabricated with micron-scale physical features in polydimethylsiloxane (PDMS) and glass. Samplers will be selectively baited and placed in different natural environments in order to select for microorganisms based on morphology, behavior, habitat, and prey source, and will enable direct observation of entrapped live protozoa via light microscopy. Genetic analysis of collected eukaryotes and prokaryotes will help provide a unified framework for taxonomic, functional, and genomic information. Together with co-located micro-scale measurements of environmental variables such as pH, temperature, and nutrient concentrations, this work will enable researchers to place microbial eukaryotes within the context of their immediate physical and chemical microhabitats. This work will enable development and validation of a completely new microbial community analysis tool that will be of immediate and practical use to microbial ecologists, aqueous geochemists, engineers, and marine scientists. It may help to answer fundamental questions in biology including what is the relevant physical scale of community structure variations, and to what extent are protozoan species cosmopolitan vs. endemic in natural habitats? Also, a better understanding of microbial community structure and function will permit better predictions of biogeochemical feedbacks as a function of climate change; improved understand the impacts of microbial community structure and function on contaminant uptake and mobilization, and enhanced risk models for environmental reservoirs of microbial pathogens. Future work will integrate a comprehensive suite of sensing capabilities and will deploy devices in a broader range of natural and engineered microbial habitats. The results of this project will be disseminated through peer-reviewed journals and conference presentations that target the biological user community. Implementation of the technology will be enabled by web-posted demonstration videos that illustrate how samplers are designed, created, tested, deployed, and studied back in the lab. All materials will be available online at www.cmbe.engr.uconn.edu/facultyshor.html.
. [PI Shor] (5/1/10 to 4/30/12, no-cost extension to 4/30/14). $160k. The goal of this project was to develop microfluidic passive samplers to capture, concentrate, and observe live aquatic protists collected from field sites. INTELLECTUAL MERIT: Passive samplers were created on microscope slides using PDMS-cast microfluidic devices. Several different sampler geometries were created and tested in freshwater and estuarine field sites. Testing showed many different protist forms were successfully trapped in observation galleries arrayed on chips based on the organism’s own motility and behavior. The geometry of individual galleries limits the ability of protists to swim or crawl back out, similar to an array of miniature lobster traps. BROADER IMPACTS: This research resulted in a new protist sampling tool of broad use to biologists. K-12 students have already used the tool. New high-content and high-throughput methods were developed to image protists in trap arrays. Five conference papers were presented at national meetings including ACS, ASM, and AEESP, one manuscript is in review, and two others are in preparation. Two graduate students, one teacher and three undergraduate students were cross-trained in microbiology and engineering (four of these six participants are from under-represented groups).
