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This report reviews the highlights and evaluates the immediate outcomes of my 2009-10 sabbatical year at the California Institute of Technology in Pasadena, California.  I worked in the laboratories of Professor Jared Leadbetter, a microbial ecologist, located in the Division of Engineering and Applied Science.  The sabbatical was an outstanding success intellectually and in terms of experimental productivity.  While some experiments led to setbacks and others took some unexpected turns during the sabbatical, the understanding that I gained ultimately proved invaluable resulting in a productive summer with my Occidental College research group in June through August 2010. 

Fundamental chemical measurements were proposed to understand biosurfactant synthesis in surface-bound bacteria.  Biosurfactants are surface-active compounds synthesized by a wide variety of microorganisms.  Like chemical surfactants, these compounds have hydrophobic and hydrophilic domains, respectively, and lower surface tension when enriched at interfaces.  Two critical uses of surfactants are found in the petroleum industry, and biosurfactants may prove better choices than synthetic surfactants to both remediate spills as well as increase recovery from natural oil sources.  Biosurfactants generally exhibit lower toxicity, higher biodegradability, and biosurfactants can be produced under milder conditions.  Thus, economical production of biosurfactants from bacteria grown on surfaces with sustainable feedstocks either natively or more likely in a genetically engineered host, could prove beneficial. 

To expand the range of compounds produced with genetically engineered microbes, a larger library of biosynthetic pathways from various native hosts is needed.   To address this need, I proposed to elucidate the isoprenoid biosynthetic pathways of a biosurfactant in the bacterium Bdellovibrio bacteriovorus.  Many prokaryotes have a common pathway to make isoprenoid-based compounds from C₅ based building blocks, but to my knowledge no detailed mechanisms have been developed for B. bacteriovorus.   Bdellovibrio bacteriovorus is a small, motile, Gram-negative bacterium that preys on a wide variety of other Gram-negative bacteria.  In earlier studies, my collaborators and I investigated the growth and lifestyle of this bacterium at interfaces.   About 4 years ago, my laboratory discovered a prey-independent phenotype of this bacterium when host-dependent cells are trapped at an interface; these host-independent cells possess vastly different morphologies and exhibit a pigment.  We extracted and isolated a biosurfactant from these trapped cells, and have nearly completed its chemical characterization from measurements with LC-MS and high-resolution tandem mass spectrometry at Caltech.  More refined separations are needed to unambiguously determine the hydrophilic head region with NMR.  At this writing, we continue to assign our biosurfactant to a member of the isoprenoid family, and additional LC data collected during my sabbatical suggest that the bacterium synthesizes a large number of chemically similar compounds to thrive in a host-independent (without prey) lifestyle at an interface. 

We hypothesized that a quorum of surface-bound B. bacteriovorus synthesizes this surfactant to lower interfacial tension and allow efficient uptake of nutrients.  To begin to test this hypothesis, I originally proposed to use my sabbatical to complete the chemical characterization of our biosurfactant and to identify the chemical cues that lead to this host-independent phenotypic switch with accompanying biosynthesis.  To accomplish these goals, I started by first learning to cultivate and handle the bacterium in the Leadbetter lab.  This seemingly straightforward task took more time than allotted, and I was fortunate to have access to Professor Leadbetter and his graduate students and postdoctoral fellows to assist me with all the modern as well as traditional microbiological laboratory techniques and assays. 

Once the cultivation issues were solved, I investigated the rate and morphological changes of the phenotypic change from host-dependent (HD) to host-independent (HI) lifestyle of Bdellovibrio at an interface.  The conversion is rapid (perhaps less than 1 day) between HD and HI Bdellovibrio phenotypes at high cellular densities, and indicates that quorum sensing may involved.  Quorum sensing (QS) is a mechanism by which bacteria use the concentration of secreted small molecules to determine cell density; at high cell densities these small molecules re-enter the cells and induce phenotypic lifestyle changes such as biofilm formation, gliding motility, adhesion, and virulence.  While elucidating any QS mechanisms in this bacterium was a proposed goal, I realized that I first had to collect additional convincing data to show that the observed morphological changes in cells from HD to HI phenotypes observed by high-resolution microscopy in Leadbetter’s laboratory were in fact Bdellovibrio and not prey cells or contaminants.  Thus, new techniques including FISH (Fluorescence In-Situ Hybridization) have been employed recently to study this phenotypic switch with promising preliminary findings.  Working in a lab of microbial ecologists pushed my group at Occidental to determine if this phenotypic switch, observed in a lab-cultivated strain, is relevant in the environment.  This summer two of my students isolated a strain (as yet unknown) of Bdellovibrio and showed that indeed this phenotypic switch occurs quite readily.  These results will lead to publication in the next one to two years.

All in all, I was immersed in the intellectual atmosphere of a vibrant and productive research lab, attending group meetings and seminars and participating in the life of the lab.  In addition, Professor Leadbetter provided scientific guidance as well as ample bench space, equipment, instruments, and consumable supplies.  Such space and resources were critically important for this sabbatical year as one of my primary research goals was to run all my experiments on my own.  I did that lab bench work for 9 months, and I now possess first-hand knowledge of growing and handling this fascinating microbial system.  Moreover, the intellectual and collegial interactions with Leadbetter and his group were of the highest quality.  Professor Leadbetter and I continue to collaborate following the conclusion of my sabbatical.  Finally, from this sabbatical year, I am feel equipped to purposefully (1) attract students to study forefront chemical questions in an interdisciplinary context, (2) continue to write competitively for external funding to fund research with undergraduate collaborators, and (3) better probe this novel microbial system in my own lab with a variety of modern techniques.