Project Overview

Summary:

     In January 2016, Dr. Jacinta Conrad at the University of Houston was awarded an RFP-V grant of $1,818,344 to lead the GoMRI project entitled, “Role of Microbial Motility for Degradation of Dispersed Oil” consisted of 3 collaborative institutions and approximately 17 research team members (including students).   

     After a catastrophic oil spill, quickly remediating, removing, or cleaning the spilled oil can prevent damage to fragile coastal, open-ocean, and deep-water ecosystems and minimize public health and economic impacts for nearby human communities. Improved understanding of both natural and human-engineered processes that generate rapid remediation is therefore a top priority. Microbial biodegradation processes are thought to have played a substantial role in the surprisingly swift disappearance of oil and gas released into the Gulf of Mexico after the catastrophic Deepwater Horizon MC252 blowout. Although previous GoMRI-supported work investigated the composition of the coastal, open-open, and deepwater microbial communities that degraded this oil, much remains poorly understood regarding the impact of physical factors in heterogeneous ocean and coastal environments on the rate of microbial biodegradation. These factors include both steady ocean currents and intermittent flows due to turbulence; gradients in temperature, pressure, and salinity; and variations in local organic matter and soil content. These physical factors are expected to affect the ability of motile hydrocarbon-utilizing bacteria to move towards oil via chemotaxis, the directed motility of bacteria towards a chemical attractant, and hence the rate at which they degrade oil. Understanding the effects of physical factors on microbial biodegradation is additionally complicated by human-engineered strategies to clean spilled oil. For example, dispersants applied to break large oil slicks into smaller droplets can render oil more accessible to degradation by bacteria. Previous GoMRI-supported work identified the effects of dispersants on the viability of microbial populations, but their impact on microbial motility and attachment to oil-water interfaces remains poorly understood. Furthermore, dispersants alter the size of oil droplets and the surface tension and compliance of oil-water interfaces, which may in turn modulate microbial motility and chemotaxis. Hence there is a pressing yet unmet need to understand how (a) nearby liquid oil/liquid water or gaseous oil/liquid water interfaces, (b) fluid flow, and (c) dispersants affect microbial motility towards dispersed oil. Moreover, this need must be addressed for bacteria living in each type of ecosystem impacted by catastrophic oil spills.

     The objective of this proposal is to elucidate the effects of oil-water interfaces on motility of marine bacteria in the initial stage in biodegradation, as microbes move towards and attach to dispersed oil. This proposal addresses Theme 2 in the GoMRI RFP-V, "Chemical evolution and biological degradation of the petroleum/dispersant systems and subsequent interaction with coastal, open-ocean, and deep-water ecosystems." The underlying idea driving the proposed work is that bacterial chemotaxis will enhance the rate of biodegradation and can be modulated by physical factors in the environment and by properties of oil-water interfaces. The four scientific questions to be addressed through this proposal are:

(1) How does bacterial chemotaxis affect the rate at which microbes degrade oil?
(2) How do the elevated pressures found in the ocean and in the Deepwater Horizon spill modify the motility, chemotaxis, and hydrocarbon utilization?
(3) How do dispersants alter microbial motility mechanisms and adhesion to oil-water interfaces during biodegradation?
(4) How do viscoelastic interfaces (characteristic of bacteria- and dispersant-coated oil drops) and flows (characteristic of the ocean environment) alter microbial motility?

These questions will be addressed by a team of four investigators with complementary expertise in microbiology, functional genomics, microscopy, interfacial science, high-throughput image analysis, mathematical modeling, and simulation. Using state-of-the-art experimental and computational methods, the team will address four hypotheses answering the scientific questions. The expected outcome of this plan of work is new understanding of how dispersed oil modifies bacterial motility and the efficacy of biodegradation, which in turn will address two ultimate goals that directly relate to the project objective and to GoMRI Theme 2. First, this plan of work will generate new insight into strategies for dispersant use that optimize microbial biodegradation rates, allowing spilled oil to be more rapidly cleaned. Second, this knowledge will contribute to improved models to predict the amount of oil degraded by microbes. Successfully achieving these goals will improve the ability to respond to and mitigate catastrophic oil spills in the Gulf of Mexico and hence the proposed work is strongly aligned with the mission of GoMRI.

Research Highlights

     As of December 31, 2019, this project’s research resulted in 16 peer-reviewed publications, 41 scientific presentations, and 21 datasets being submitted to the GoMRI Information and Data Cooperative (GRIIDC), which are/will be made available to the public. The project also engaged 2 Master’s level and 12 PhD level students over its award period. Significant outcomes of this project’s research according to GoMRI Research Theme are highlighted below.

Theme Two:

PhD student Vaseem Shaik and Dr. Arezoo Ardekani (Purdue) obtained solutions of the governing equations for the motion of the swimmer near a weakly-deforming plane interface (such as an oil-water interface), published in the Journal of Fluid Mechanics. The long-time dynamics of a time-reversible swimmer are such that it either moves towards or away from the interface. Thus, its phase space can be divided into regions of attraction (repulsion) towards (from) the interface. The long-time orientation of a time-reversible swimmer that is moving towards the interface depends on its initial orientation.

PhD student Nikhil Desai and Ardekani (Purdue) authored an invited review article in the journal Soft Matter, detailing the modeling of micro-organism suspensions. They reviewed different mathematical models and experimental studies on bacterial chemotaxis in conjunction with fluid flow. They highlighted the importance, and reviewed the formulations, of bacteria-fluid interactions in the context of marine chemotaxis, the fundamental mechanism of subsurface bioremediation. This review thus summarizes the fluid mechanics relevant for understanding interactions of bacteria under flow.

Shaik, Desai, and Ardekani derived expressions for the translational and rotational velocities of a swimming micro-organism (modeled as a force-dipole) located outside an oil drop covered with surfactant. Using these results, they analyzed the swimming dynamics of micro-organisms outside surfactant-laden-drops, and studied the attraction and possible attachment of the micro-organisms to the oil drop. This study was published in Physical Review Fluids.

Motivated by bacterial bio-remediation of insoluble hydrocarbons (HCs) released during oil spills, Desai, Shaik, and Ardekani evaluated the critical trapping radius and trapping time distribution of swimmers in Soft Matter. Addition of surfactant reduces the critical trapping radius of a drop by ∼30%. For hydrodynamics combined with diffusion-based motion, the interface-retention times are greater by ∼5–25% for surfactant-laden drops as compared to clean drops. These differences occur for low values of surface viscosity, and saturate rapidly as the surface viscosity increases. This study reveals that additives such as dispersants can alter the hydrodynamics of bacteria-drop interactions.

Ardekani and collaborators performed probabilistic simulations to ascertain the impact of environmental and motility characteristics on the spatial distribution of microorganisms around oil drops, published in Physical Review E. They described a host of bacterial behaviors near the nutrient source, dictated mostly by their morphology and chemotactic ability. They showed that hydrodynamic trapping can significantly amplify (by ≈40%) the nutrient exposure of bacteria, both chemotactic and non-chemotactic. They also showed that larger aggregates (marine snow particles with radii greater than 1 mm) proved too fast for the bacteria to get trapped onto, thus diminishing the role played by hydrodynamics in those regimes. Rising crude oil drops are not amenable to hydrodynamic trapping. Thus, while hydrodynamics is significant in dictating bacterial accumulation around fixed and neutrally buoyant drops, the same cannot be said for rising oil drops.

PhD student Adib Ahmadzadegan and Ardeknai conducted experiments to study the hydrodynamic attraction of motile bacteria to liquid-liquid interfaces. A mathematical model was developed to rationalize the experimental observations. The bacterial distributions within the suspension were used to measure important features of the forces exerted by the bacteria on their surrounding fluid. For example, it is known that bacteria exert equal and opposite forces and torques on their surrounding fluids. The near-interface concentration of bacteria and the radius of the circular trajectories traced by the bacteria are directly related to the magnitude of these forces and torques, respectively. Thus, by measuring these quantities and relating them to the parameters of our mathematical model, we can estimate the forces and torques exerted by bacteria on their surroundings. A measure of these forces and torques can inform our understanding of flow fields produced by bacteria and their effects on bacterial motion in the bulk and near liquid-liquid interfaces. This manuscript is in preparation.

MS student Ryan McLay and Dr. Jacinta Conrad (Univ. Houston) studied the adhesion of E. coli bacteria with tunable expression of type 1 fimbriae, a surface adhesin, to oil/water interfaces. Increasing the level of fimbriation (by increasing the expression level of an inducible fim operon) increased the ability of these bacteria to adhere to the oil/water interface, as characterized through single-cell direct imaging and a bulk MATH assay. Without enhanced fimbriation, wild-type and fimbriae-deficient E. coli bacteria rarely adhered to oil/water interfaces. This study, published in Langmuir, showed that changes in surface properties can significantly alter how bacteria interact with dispersed oil.

PhD student Narendra Dewangan and Conrad characterized the adhesion of Marinobacter hydrocarbonoclasticus, a nonmotile hydrocarbon degrader, to dodecane droplets that were stabilized by various surfactants, including a component of Corexit dispersants used in oil-spill response. The adsorption of bacteria at the oil/water interface followed kinetic models used for passive colloids, with the greatest areal number density of bacteria adhered to the smallest droplets. For a fixed droplet size, the long-time areal density of bacteria at the interface decreased with increasing surfactant concentration because of a reduction in oil/water interfacial tension that increases the free energy of adhesion of the bacterium. This knowledge, published in Langmuir, provides insight into physicochemical interactions between bacteria and dispersed oil. In a follow-up study recently published in Soft Matter, Dewangan and Conrad showed that motility increased the rate at which bacteria adhered to droplets and the surface density at which they packed there. This study used the motile hydrocarbon-degrader Halomonas titanicae.

Dr. Roseanne Ford(Univ. Virginia) and collaborators published in AIChE Journal a manuscript describing similarity solutions of transport equations for bacterial chemotaxis under shear flow near an oil-water interface. This study is a first step towards incorporating bacterial behaviors into models for biodegradation in marine environments.

PhD student and GoMRI scholar Kelli Mullane, MS student Tanya Xu, and Dr. Douglas Bartlett (UCSD) carried out a thorough set of experiments to explore the effects of hydrostatic pressures of the magnitude present in the Deepwater Horizon oil plume on motility and chemotaxis of hydrocarbon-degrading marine microbes. Using three Gulf Strains from various depths, Mullane characterized the effects of pressure and temperature on the motility, growth, and hydrocarbon degradation of selected hydrocarbon degrading Gulf of Mexico bacteria. Xu examined the growth of the strains as a function of pressure on hexadecane with Corexit, performed experiments to demonstrate that the study strains can degrade hexadecane pressure using gas chromatography, and measured the protein synthesis activity of bacterial cells using the fluorescent BONCAT assay coupled with flow cytometry.  This was done to assess the transient effect of pressure on the activity of the three strains during grown with hexadecane under anaerobic respiratory conditions with nitrate as an electron acceptor. This study is in preparation.

 Proposal Abstract - RFP-V PI Conrad