Center for Biofilm Engineering News Update:  December, 2001 Volume 4, Issue 12 __________________________________________   Research Highlights   Paper Spotlight by Dr. Paul Stoodley   This paper deals with the detachment of cells from bacterial biofilms. Detachment is arguably the least studied biofilm process and yet plays a critical role in the dissemination of infection and industrial contamination. In this paper we directly observed the growth and detachment of cell clusters from two mixed species biofilms, both of which contained opportunistic pathogens. By using digital time-lapse microscopy we were able to subtract each frame from the subsequent frame to quantify and size the clusters that had detached during the interval. We found that although smaller cell clusters detached more frequently than larger cell clusters, the larger cell clusters contained a disproportionately high percentage of the total detached biomass. The largest cluster that detached during the 19 h monitoring period had a diameter of approximately 500 µm, and we estimated that it could have contained up to 1x105 CFU. To compliment these results we also microscopically examined detached biomass in the effluent. Effluent was collected and filtered without vortexing or homogenization. Cells were stained with Molecular Probes live/dead kit. Confocal microscopy showed that the clumps formed flat monolayers on the filters allowing us to calibrate particle area with cell number for automated counting. These results showed a similar pattern to those from direct observation of detaching biofilm particulates. Although single cells and clusters of up to 10 cells made up over 90% of the total number of detached particles, they only contained approximately 50% of the total detached biomass. Conversely, particles containing over 1000 cells made up less than 1% of the total number of detached particulates but contained almost 10% of the total number of detached cells. The particle size distribution showed a continuum between single cells and cell clusters of up to 0.5 mm diameter calling into question the division of detachment into two distinct processes, erosion and sloughing. Finally, information on biofilm detachment is usually inferred from monitoring the cell concentration in the liquid phase after sample homogenization. A minimum infectious dose of cells concentrated in a biofilm particulate may be overlooked if such an aggregate is disrupted and diluted in the test-tube. We concluded that in public health issues it might not only be the cell concentration which is of concern but also the manner in which the biomass is distributed.   Stoodley, P., Wilson, S., Hall-Stoodley, L., Boyle, J.D., Lappin-Scott, H.M., and Costerton, J.W. 2001. Growth and detachment of cell clusters from mature mixed species biofilms. Appl. Environ. Microbiol. 67(12):5608-5613.   _______   Collaborative $1.2 Million Grant to Study Clean-Up of Uranium   Dr. Brent Peyton, a researcher at the Center for Multiphase Environmental Research, Washington State University and also a CBE alumni, has recently received a $1.2 million grant from the Department of Energy's Natural and Accelerated Bioremediation Program (NABIR) to study the use of bacteria in stopping the underground movement of contaminants, such as uranium, in groundwater.   The work is important for designing treatment strategies and for improving clean-up of contaminants from soils around aging nuclear facilities, such as the DOE’s site at Hanford, Washington. If the uranium can be precipitated out of the groundwater and locked up in the soil, sensitive areas such as rivers and wetlands can be protected. Although bacteria have been used for years to clean-up organic contaminants such as oil spills and chlorinated solvents, their use in clean-up of pollution from heavy metals is a new area in bioremediation.   Working in collaboration with a multi-disciplinary team of researchers, including Gill Geesey and Zbigniew Lewandowski from Montana State University and James Amonette at the Pacific Northwest National Laboratory, Brent Peyton, assistant professor in chemical engineering, hopes to determine how different types of bacteria and soil compositions affect the ability to immobilize uranium in contaminated aquifers. Previous work has shown that bacteria that are found naturally in soils and groundwater may be used to stop the spread of toxic heavy metals and radionuclides, including uranium.   Novel techniques will be used in the three-year project, including biological reactors that simulate groundwater flow, micro-sized probes to measure chemical concentrations near bacterial surfaces, and specially designed field sampling equipment to better relate laboratory tests to field conditions. _______   Dow Chemical Contribution   Dow Chemical recently contributed laboratory equipment and supplies to the CBE. Their generous contribution and continued support of the CBE's research is greatly appreciated by the faculty, staff and students.   _______     Welcome New Industrial Associate Members   Kurita Water Industries Designated Representative: Minoru Yagi, Chief Researcher Kurita's research focuses on membrane separation, ion-exchange, and organic treatment technologies that are core technologies for removing impurities and toxic elements from water. They also conduct R&D activities relating to pollution purification technologies for soil and underground water, as well as corrosion and scale-prevention technologies for the development of water treatment chemicals. Website: http://www.kurita.co.jp/english/e-index.htm   Philips Oral Healthcare, Inc. Designated Representative: Greg Peterson, Scientist Philips Oral Healthcare makes the Sonicare toothbrush and their research interests are in plaque removal and cavity prevention. Website: http://www.optiva.com   _______   New Staff Member   Jennifer Sestrich, Lab Specialist, recently joined Mark Pasmore's research group where she will be working on medical projects in the Biofilm Behavior Laboratory. Jennifer graduated with a microbiology degree from Montana State University in 1998. She completed a one-year medical technology internship at the Sacred Heart Medical Center in Spokane, Washington and then was hired as a medical technologist at the Marcus Daly Hospital in Hamilton, Montana.   _________________________________________   Upcoming CBE Workshops and Meetings   See the Events Calendar at http://www.erc.montana.edu/CBEssentials-SW/whats_new/index.htm   _________________________________________   CBE People in Action   Drs. Haluk Beyenal and Zbigniew Lewandowski presented "Structure and Activity of Sulfate Reducing Biofilms and Mineral - Biofilm Interactions During Lead Immobilization," at the Advances in Environmental Biotechnology # 2, American Institute of Chemical Engineers’ 2001 Annual Meeting, Reno, Nevada, November 4-9, 2001.   Nurdan Yurt and Drs. John Sears and Zbigniew Lewandowski presented "Quantifying Biokinetic Parameters for Microbial Growth in Biofilms from Substrate Concentration Profiles," at the Thermodynamics and Transport in Biological Systems, American Institute of Chemical Engineers’ 2001 Annual Meeting, Reno, Nevada, November 4-9, 2001.   Drs. William G. Pitt and John D. Pruitt presented "Temporary Stabilization of Pluronic P-105 Micelles for Drug Sequestration, Advances in Structure, Function and Stability for Biologics and Pharmaceuticals #2, American Institute of Chemical Engineers’ 2001 Annual Meeting, Reno, Nevada, November 4-9, 2001.   Dr. Bill Costerton was an invited visitor to DuPont Fluoroproducts in Wilmington, DE. He met with a group of research personnel and gave a seminar on the CBE and biofilms, November 28, 2001.   Dr. Bill Costerton presented "Biofilm Infections in the ICU Patient" at the Surgery Grand Rounds meeting, University of Louisville, KY, December 1, 2001.   Dr. Bill Costerton as an invited speaker presented "Bacterial Biofilms: A Common Cause of Persistent Infections," at the Center for Devices and Radiological Health's (CDRH) Antimicrobial Device Efficacy Testing Seminar at the Food and Drug Administration (FDA) in Rockville, MD, December 3-4, 2001.   Dr. Bill Costerton as an invited speaker will present "Biofilms and Signaling," at the International Symposium - New Frontiers in Microbial Ecology and meeting on The International Activities of ASM in Barcelona, Spain, December 11-12, 2001.   __________________________________________   Theses   See theses abstracts at http://www.erc.montana.edu/Res-Lib99-SW/pubs/Theses/default.htm     __________________________________________   Web Watch     Subscribe to the BiofilmsOnline.com Newsletter   A newsletter subscription will keep you notified of recent postings in the Biofilm News, Patents and Research Digest. The Subscribe link is located under Current Contents.  See the BiofilmsOnline.com December issue at http://www.BiofilmsOnline.com.   _______   Industry Sponsored Research   The CBE has developed new web pages with information about Industry Sponsored Research. Check the Recent Web Updates pages to locate the latest information posted on the CBE web. See http://www.erc.montana.edu/Recent%20Web%20Updates/default.htm     __________________________________________   Newsletter Listserve   The CBE News Update is a listserve newsletter. If you need to subscribe or unsubscribe from the listserve, follow the directions at the following CBE website.   http://www.erc.montana.edu/Res-Lib99-SW/newsarchives/subscribe.htm   An alternative to subscribing to the listserve is to view the CBE News Update on our web page at http://www.erc.montana.edu/Ind-Col99-SW/Current_Newsletter/default.htm. Newsletter archives can be found at http://www.erc.montana.edu/Res-Lib99-SW/newsarchives/index.htm.   __________________________________________   Diane Williams  editor of the CBE News Update