S297: Soil Microbial Taxonomic and Functional Diversity as Affected by Land Use and Management

(Multistate Research Project)

Status: Inactive/Terminating

Project Menu

SAES-422 Reports

Annual/Termination Reports:

[07/18/2001] [08/15/2002] [07/26/2004] [10/03/2005]

Date of Annual Report: 07/18/2001

Report Information

Annual Meeting Dates: 06/11/2001 - 06/13/2001
Period the Report Covers: 10/01/2000 - 05/01/2001

Participants

S. Albrecht, USDA-ARS, Oregon, Chair-elect;
J.S. Angle, University of Maryland;
J.A. Entry, USDA-ARS, Idaho;
S. Deng, Oklahoma State University;
Y. Feng, Chair, Auburn University;
A. Franzleubbers, USDA-ARS, Georgia;
J.J. Fuhrmann, University of Delaware;
J. Germida, University of Saskatchewan;
J.H. Graham, University of Florida;
C. Hagedorn, Virginia Polytechnic Institute and State University;
P.G. Hartel, Secretary, University of Georgia;
W.J. Hickey, University of Wisconsin;
K. Jayachrandan, Florida International University;
M.B. Jenkins, USDA-ARS, Georgia;
R. Klucas, University of Nebraska;
T. McGonigle, Idaho State University;
J.B. Morton, West Virginia University;
M.D. Mullen, University of Tennessee;
C.M. Reynolds, U.S. Army (CRREL);
E.C. Schroder, University of Puerto Rico;
H.D. Skipper, Clemson University;
D.M. Sylvia, University of Florida;
J. Thies, Cornell University;
D.C. Wolf, University of Arkansas;
A.G. Wollum, North Carolina State University;
D.A. Zuberer, Texas A&M University

Brief Summary of Minutes

The annual meeting minutes can be found at the S-297 project web site (http://dmsylvia.ifas.ufl.edu/msp/minutes.htm).

Minutes Attachment:

Attachment

Accomplishments

Objective 1: To determine the geographic and temporal variability of E. coli ribotypes in the United States. Eight experiments were conducted under this objective. In Experiment #1, ribotyping was used to determine a source of fecal contamination. Ribotyping results suggested that the point source of the E. coli contamination was a pond, sinkhole, or two streams, and not the septic systems. In Experiment #2, ribotyping was used to determine the biogeographic variability of the subspecies of the bacterium Escherichia coli from one location Idaho and three locations in Georgia for four animals, cattle (Bos taurus), horse (Equus caballus), swine (Sus scrofa), and poultry (Gallus gallus domesticus). The data suggest that there is good ribotype separation among host species at one location, and that the distance to obtain good host origin matching depends on a distance <175 km and a large sample size. In Experiment #3, ribotyping was to determine the temporal variability of E. coli ribotypes at one location in Georgia. A sample of 25 E. coli isolates were obtained from six steers at four sampling times over a one-year period. A total of 347 ribotypes were identified from a 547 E. coli isolates.  No ribotype was observed at all all sampling times. In Experiment #4, ribotyping was used to determine how much sampling was needed to obtain a representative sample of E. coli ribotypes in a watershed with a complex land use patterns and varied flow conditions. The study area was a 77-kilometer reach of the Chattahoochee River and its tributaries in metropolitan Atlanta. The large number of unique ribotypes obtained suggests that considerable ribotype heteogeneity exists, and that a large number of E. coli isolates is needed to obtain a representative sample of ribotypes from watersheds with complex land use patterns and varied flow conditions. In Experiment #5, ribotyping was used to determine the source of fecal contamination in a watershed of moderate complexity. In this case, Rock Creek, a 66-km creek in Idaho that empties into the Snake River, was selected. Of 46 E. coli isolates that have been ribotyped so far, there were matches between environmental isolates and humans. More isolates need to be ribotyped before any firm conclusions are possible. In Experiment #6, ribotyping was used to identify the host origin of fecal contamination in a forested watershed that contained penned deer but no humans or domestic animals. The results suggest that ribotyping can identify the source of fecal contamination to such an extent that it may be possible to delist a watershed impaired only by wildlife. In Experiment #7, research was conducted to determine if it was possible to use the ribotyping in conjunction with the restricted host range of Enterococcus faecalis, a fecal streptococcus, to distinguish between human and nonhuman sources of fecal contamination in water. Only a few of the isolates have been ribotyped, but preliminary results suggest that isolates from humans and nonhumans were different. Therefore, if the restricted host range of Ent. faecalis is combined with ribotyping, then it may be possible to distinguish between human and nonhuman sources of fecal contamination easily. In Experiment #8, antibiotic resistance analysis (ARA), carbon source utilization (Biolog), and pulsed-field gel electrophoresis (PFGE) were used for comparative purposes to classify a collection of 60 E. coli isolates from known sources (10 each from human, cow, horse, deer, goose, and sheep) (Virginia). Objective 2: To determine relationships among microbial taxonomic and functional diversity, contaminant bioavailability, and remediation rates for different organic-contaminated soils. Eight experiments were carried out under this objective. Two experiments involved the phytoremediation of heavy metals. The influences of metal hyperaccumulator plants, Thlaspi caerulescens and Trifolium pratense L., on soil microbes in the rhizosphere were compared (Maryland). The distribution of bacteria in heavy metal contaminated and uncontaminated soils was evaluated to explain the presence of metal sensitive bacteria in metal contaminated soils and the presence of metal resistant bacteria in uncontaminated soils (Maryland). Experiment#3 is a collaboration between Arkansas and New Hampshire, dealing with rhizosphere-enhanced remediation of PAH-contaminated soils. Field studies were continued at Alaska and Korea sites. There was a positive vegetation effect in reducing extractable PAHs relative to controls, as PAH molecular weight increased. The fertilizer without vegetation treatment resulted in the least reduction in heavier PAHs. New field plots were established in Experiment#4 to determine the rhizosphere effects on remediation of a crude oil-contaminated site in El Dorado, AR. In Experiment#5, PAH-degrading bacteria were isolated using standard enrichment culture techniques and procedures involving a sorbed substrate (Wisconsin). Experiments were carried out to analyze the phylogenetic diversity of these isolates, and examine the organisms? sorptive characteristics. Experiment#6 determined the effects of the characteristics of bacteria on the differential accessibility of soil-sorbed biphenyl (Alabama). Experiment#7 has been initiated in Florida?Miami to evaluate the effect of soil amendments (compost) on soil water balance, agro-chemical retention, and soil hydraulic properties in order to improve crop water use efficiency and reduce the potential for movement of atrazine and nutrients (P and N) into surface and subsurface water. In Experiment#8, pure cultures of bacteria that were able to use 2,4,6-trinitrotoluene (TNT), or hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) as the sole nitrogen source were isolated in Oklahoma. Objective 3: To characterize taxonomic and functional diversity of bacteria and mycorrhizae in disturbed lands and urban landscapes. Fourteen experiments were undertaken under this objective. Seven experiments examined mycorrhizae under different landuse conditions. Experiment#1 was conducted to study the impact of AM fungi and phosphorus on tomato competition with bahiagrass (Florida-Gainesville). The results indicated that the role of mycorrhizae in plant competition for nutrients was markedly impacted by soil nutrient status. Experiment#2 examined the occurrence of AM fungi in roots under combinations of cover crops, plastic mulch and solarization as alternatives for control of soil-borne pathogens and weeds in tomato/pepper systems (Florida-Lake Alfred). In addition, the ‘fallow effect‘ after long cycle cropping of sugarcane was investigated (Florida-Lake Alfred) (Experiment#3). In Experiment #4, isolation of fungi from moderate to high organic matter and phosphorus soils in sugarcane fields was carried out through the collaborative effort between Florida-Lake Alfred and West Virginia. Investigations on the mycorrhizal ecology in nonagricultural ecosystems were carried out in Florida (Gainesville) and West Virginia. In Experiment #5, ecotypic variation among populations of AM fungi associated with sea oats obtained from four divergent locations in Florida was characterized and a nested PCR approach was being developed to genetically characterize AM fungal isolates (Florida-Gainesville). In Experiment #6, comparative analyses of fungal communities in the southwestern deserts of the U.S. (Arizona, New Mexico, Texas) and Mexico with those of Namibia, Africa were also conducted (West Virginia). Experiment #7 was carried out to investigate the bias in using roots as inoculum in trap pot cultures to assess fungal species actively forming mycorrhizal associations (West Virginia). Seven other experiments under Objective 3 emphasize the bacterial communities in soils. In Experiment#8, the ecological shifts or stability of rhizobacteria from yellow nutsedge, Palmer amaranth, large crabgrass, common ragweed, and tropic croton over time in Norfolk soil were determined (South Carolina). In Experiment#9, whole soil fatty acid methyl ester (FAME) analysis was used to assess changes in microbial community structure of a soil planted with juniper (Georgia). In Experiment#10, field plots were established on a coffee farm to study pesticide effects on soil microbial composition and activity (Puerto Rico). Experiment#11 was initiated in Tennessee to evaluate the impacts of Bt-corn on microbial community structure of the rhizosphere. The study of the microbial biodiversity of surface mined lands reclaimed using mixed overburden was completed in Texas (Experiment#12). Total fatty acids (an estimator of biomass) showed a linear increase with time in reclamation, consistent with earlier studies using chloroform fumigation incubation or extraction methods as well as substrate-induced respiration. Biolog community-level physiological profiles indicated that there is a high degree of similarity in carbon-source utilization at all the study sites suggesting a high degree of redundancy in substrate utilization among the microbial communities regardless of time in reclamation. In Experiment#13, monitoring the abundance of water-extractable C in sand-based root zones under dwarf bermudagrass was continued (Texas). In Experiment#14, two methods, phospholipid fatty acid (PLFA) and whole-soil fatty acid (WSFA) profile analyses, were evaluated for assessing soil microbial community structure (Alabama). In addition to the above projects, work is beginning on a second edition of the textbook, Principles and Applications of Soil Microbiology (now tentatively re-named Soil Microbiology: Environmental and Agricultural Perspectives) and a new laboratory manual to accompany it. In the case of the textbook, this project involves at least 13 multistate collaborators. Investigators from Delaware, Georgia, and Texas are collaborating on this project.

Publications

Refereed Articles: Angle, J.S., R.L. Chaney, A. J.M. Baker, Y.-M. Li, R. Reeves, V. Volk, R. Roseberg, E. Brewer, S.Burke, and J.P. Nelkin. 2000. Developing commercial phytoextraction technologies: Practical considerations. South African J. Sci. (In press). Bressan, W., C.H.S. de Carvalho, and D.M. Sylvia. 2000. Inoculation of somatic embryos of sweet potato with an arbuscular mycorrhizal fungus improves embryo survival and plantlet formation. Can. J. Microbiol. 46:741-743.; Bever, J. D., P. A. Schultz, A. Pringle, and J. B. Morton. 2001. Arbuscular mycorrhizal fungi: More diverse than meets the eye and the ecological tale of why. BioScience (In press) Burke, S., J.S. Angle, R.L. Chaney and S. Cunningham. 2000. Arbuscular Mycorrhizae effects on heavy metal uptake by corn. Inter. J. of Phytoremediation. 2:23-29. Chaney, R.L., Y.-M. Li, J.S. Angle, A.J.M. Baker, R.D. Reeves, S.L. Brown, F.A. Homer, M. Malik, and M. Chin. 2000. Improving Metal Hyperaccumulator Wild Plants to Develop Commercial Phytoextraction Systems: Approaches and Progress. pp. 131-160. In N Terry and G.S. Banuelos (eds.) Phytoremediation of Contaminated Soil and Water. CRC Press, Boca Raton, FL. Chaney, R.L., J.A. Ryan, U. Kukier, S.L. Brown, G. Siebielec, M. Malik and, J.S. Angle. 2000. Heavy Metal Aspects of Compost Use. Pp. In P.J. Stofella and B.A. Kahn (eds). Compost Utilization in Horticultural Cropping Systems. CRC Press, Boca Raton, FL. Coffin, R. B., P. H. Miyares, C.A. Kelly, L. A. Cifuentes, and C. M. Reynolds. 2001. 13C and 15N Isotope Analysis of TNT; Two Dimensional Source Identification. Env. Tox. Chem. (In press) Delorme, T., J.S. Angle, F. Coale, R. Chaney. 2001. Phosphorus accumulation by select plant species. Inter. J. Phytoremediation (In press). Feng, Y., D. M. Stoeckel, E. van Santen, and R. H. Walker. 2001. Effects of subsurface aeration and trinexapac-ethyl application on soil microbial communities in a creeping bentgrass putting green. Biol.Fertil. Soils (In review) Franke-Snyder, M., D. D. Douds, L. Galvez, J. G. Phillips, P. Wagoner, L. Drinkwater, and J. B. Morton. 2001. Diversity of communities of arbuscular mycorrhizal (AM) fungi present in conventional versus low-input agricultural sites in eastern Pennsylvania, USA. Applied Soil Ecol. 16:35-48. Gagliardi, J.V., J.S. Angle, J.J. Germida, R.C. Wyndham, C.P. Chanway, R.J. Watson, C. Greer, H.H. Yu, T. McIntyre, M.A. Levin, E. Russek-Choen, S. Rosolen, J. Nairn, A. Seib, T. Martin-Heller, and G. Wisse. 2001 Intact soil-core microcosms for pre-release testing of introduced microbes: Comparison with multi-site field releases in diverse soils and climates. Can. J. Microbiol. (In press). Gagliardi, J.U., J. Buyer, J.S. Angle, and E. Russek-Cohen. 2001. Comparison of soil microbial community structural and functional analyses following inoculation of a genetically engineered and a non-engineered pseudomonas to wheat roots in diverse soils. Soil Sci. Soc. Am. J. (In Press). Graham, J. H. 2001. What do root pathogens see in mycorrhizas? New Phytologist 148:357-359. Graves, Alexandria K., Charles Hagedorn, Alison Teetor, Michelle Mahal, Amy M. Bowman, and Raymond B. Reneau, Jr. Determining sources of fecal pollution in water for a rural Virginia community. Appl. Environ. Microbiol. (In review) Haney, R.L., A.J. Franzluebbers, F.M. Hons and D.A. Zuberer. 2001. Molar concentration of K2SO4 and soil pH affect estimation of extractable C with chloroform fumigation extraction. Soil Biol. Biochem. (In press) Haney, R.L., S.A. Senseman, F.M. Hons, D.A. Zuberer. 2000. Effect of Glyphosate on soil microbial activity and biomass. Weed Science. 48: 89?93.Hartel, P. G., W. I. Segars, J. D. Summer, J. V. Collins, A. T. Phillips, and E. Whittle. 2000. Survival of fecal coliforms in fresh and stacked broiler litter. J. Appl. Poultry Res. 9: 505-512. Janstfer, A. G. and D. M. Sylvia. Isolation, culture and detection of arbuscular mycorrhizal fungi. In: Manual of environmental microbiology. C. J. Hurst et al. (eds.). 2nd Edition. American Society of Microbiology. Washington D.C. (In press) Lanfranco, L., V. Bianciotto, E. Lumini, M. Souza, J. B. Morton, and P. Bonfante. 2001. A combined morphological and molecular approach to characterize isolates of arbuscular mycorrhizal fungal in Gigaspora (Glomales). New Phytology (In press) Malik, M., R.L. Chaney, E. Brewer, Y. Li and J. Angle. 2000. Phytoextraction of soil cobalt using hyperaccumulator plants. Inter. J. Phytoremediation 2:19-33. Malik, M., R.L. Chaney, E.P. Brewer and J.S. Angle. 2001. Phytoextraction of soil cobalt using hyperaccumulator plants. Int. J. Phytoremediation. (In press). Morton, J. B. and D. Redecker. 2001. Two new families of Glomales, Archaeosporaceae and Paraglomaceae, with two new genera Archaeospora and Paraglomus, based on concordant molecular and morphological characters. Mycologia 93:181-195 Oka, N., P.G. Hartel, O. Finley-Moore, J. Gagliardi, D. Zuberer, J. Fuhrmann, J.S. Angle and H. Skipper. 2000. Misidentification of soil bacteria by fatty acid methyl ester (FAME) and BIOLOG. Biol. Fertil. Soils 32: 256-258. Olexa, T. J., P. G. Hartel, J. J. Fuhrmann, and H. A. Mills. 2001. Whole soil FAME analysis to assess benomyl effects on microbial community structure in juniper. Plant Health Prog. (In review) Peacock, A.D., M.D. Mullen, D.B. Ringelberg, D.D. Tyler, D.B. Hedrick, P.M. Gale, and D.C. White. 2001. Soil Microbial Community Response to Dairy Manure or Ammonium Nitrate Applications. Soil Biol. Biochem. 33:1011-1019.; Redecker, D., J. B. Morton, and T. D. Bruns. 2000. Ancestral lineages of arbuscular mycorrhizal fungi (Glomales). Molecular Phylogenetics and Evolution 14:276-284. Redecker, D., J. B. Morton, and T. D. Bruns. 2000. Molecular phylogeny of Glomus sinuosum and Sclerocystis coremioides places both taxa firmly in Glomus. Mycologia 92:282-285. Ryan, M. H. and Graham, J. H. 2002. Arbuscular mycorrhizal fungi in agriculture: from theory to practice. Plant Soil (special issue): (In review). Schrvder, E. C. 2001. Importance of Symbiotic Nitrogen Fixation in Tropical Forage Legume Production. In: Tropical Forage Plants: Development and Use. Eds. Sotomayor-Rmos, A. and Pitman, W. D., CRC Press, Boca Raton. Chapter 15, p 251-268. Stutz, J. C., R. Copeman, C. A. Martin, and J. B. Morton. 2000. Patterns of species composition and distribution of arbuscular mycorrhizal fungi in arid regions of southwestern North America and Namibia, Africa. Canad. J. Bot. 78:237-245 Saunders, J. A., M.-K. Lee, C. M. Morton, Y. Feng, J. C. Rutherford, V. A. Bennett, and I. Thompson. Behavior of metal(loids) during bacterial sulphate reduction in contaminated groundwater. Applied Geochemistry (In review) Sylvia, D.M., A.K. Alagely, D.O. Chellemi, and L.W. Demchenko. Impact of AM fungi on tomato competition with bahiagrass. Biol. Fertil. Soils (In review) Sylvia, D.M. and D.O. Chellemi. 2001. Interactions among root-inhabiting fungi and their implications for biological control of root pathogens. Advances in Agronomy 73:1-133. White, K. D., C. M. Reynolds, D. B. Ringelberg, J.P. Laible, K. L. Foley, and L. B. Perry. 200_. Low Temperature Microbial Activity in River Systems. (Accepted, revision as ERDC report) Zuberer, D.A., 2001. Nitrogen fixation (nonsymbiotic) in soils. Encyclopedia of Environmental Microbiology. John Wiley & Sons, NY. In press. Conference Proceedings and Abstracts: Bowman, A. M., C. Hagedorn, and K. Hix. 2000. Determining sources of fecal pollution in the Blackwater River watershed. p. 44-54. In T. Younos and J. Poff (ed.), Abstracts, Virginia Water Research Symposium 2000, VWRRC Special Report SR-19-2000, Blacksburg. Chaney, R.L., S.L. Brown, Y.-M Li, J.S. Angle, T.I. Stucaynksi, W.L. Daniels, C.L. Henry, G. Siebielec, M. Malik, James A. Ryan and Harry Compton. 2001. Progress in Risk Assessment for Soil Metals, and In situ ?Phytoremediation: State of the Science.? May 1-2, 2000. Boston, MA. (In Press). Chaney, R.L., S.L. Brown, J.S. Angle, T.I. Stuczynski, W.L. Daniels, C.L. Henry, G. Siebielec, Y.-M. Li, M. Malik, J.A. Ryan and H. Compton. 2000 In situ Remediation/ReclamationRestoration of Metals Contaminated Soils using Tailor-Made Biosolids Mixtures. Chapter 2; 24 pp. In Proc. Symp. Mining, Forest and Land Restoration: The Successful Use of Residuals/Biosolids/Organic Matter for Reclamation Activities (Denver, CO, July 17-20, 2000). Rocky Mountain Water Environment Association, Denver, CO. Chaney, R.L., J.A. Ryan, Y.-M.Li, and J.S. Angle. 2001. Transfer of cadmium through plants to the food chain. In J.K. Syers and M. Gochfeld (eds.) Proceedings of the SCOPE Workshop on Environmental Cadmium in the Food Chain: Sources, Pathways, and Risks, (13-16 Sept., 2000) Belgian Academy of Sciences, Brussels. Feng, Y., D. W. Reeves, C. H. Burmester, A. C. Motta, and G. Wu. 2000. Comparison of phospholipid and whole-soil fatty acid profiles of soil microbial communities. Abstracts of 2000 annual meetings of ASA-CSSA-SSSA, p.261. Funk, A. L., M. B. Gregory, E. A. Frick, and P. G. Hartel. 2000. Microbial source tracking using ribosomal RNA typing in the Chattahoochee River National Recreation Area Watershed, Metropolitan Atlanta, Georgia?Study design and preliminary results. Building capabilities for monitoring and assessment in public health microbiology, U. S. Geological Survey, March 14-16, Columbus, OH. Entry, J. A., P. G. Hartel, J. L. Hill, and J. D. Summer. 2001. Ribotyping of Escherichia coli isolates from Rock Creek Watershed, Idaho. American Society for Microbiology, May 20-24, Orlando, FL. Godfrey, D. G., and P. G. Hartel. 2001. Geographic variability of Escherichia coli ribotypes from swine feces in Georgia and Idaho. National Conference on Undergraduate Research, March 15-17, Lexington, KY. Graham, J. H., Jifon, J. L., Drouillard, and Syvertsen, J. P. 2001. Growth depression of mycorrhizal ctrus seedlings grown at high P supply is mitigated by elevated CO2. Proc. 3rd Intern. Conf. On Mycorrhizas (ICOM3). Univ. Adelaide, South Australia (In press). Haney, R.L., A.J. Franzluebbers, F.M. Hons and D.A. Zuberer. 2000. Soil CO2 evolution: field moist vs. dried and rewetted soils. Agron. Abst. Pg. 256. Minneapolis, MN. Hartel, P. G., J. D. Summer, W. I. Segars, and J. Entry. 2000. Geographic variability of Escherichia coli from cattle and swine. American Society of Agronomy Meetings, November 5-9, Minneapolis, MN. Hartel, P. G., A. L. Funk, J. D. Summer, J. L. Hill, E. A. Frick, and M. B. Gregory. 2001. Ribotype diversity of Escherichia coli isolates from the Upper Chattahoochee River Watershed, Georgia. American Society for Microbiology, May 20-24, Orlando, FL. Hill, J. L., P. G. Hartel, W. I. Segars, and P. Bush. 2001. Ribotyping to determine the source of fecal coliform contamination in three household wells near Cochran, Georgia. p. 743-746. In: K. J. Hatcher (ed.) Proceedings of the 2001 Georgia Water Resources Conference, March 26-27, University of Georgia, Athens. Jenkins, M. B., P. G. Hartel, T. J. Olexa, and J. A. Stuedemann. 2000. Temporal variability of Escherichia coli ribotypes from cattle. American Society of Agronomy Meetings, November 5-9, Minneapolis, MN. Kern, J., B. Petrauskas, P. McClellan, V. O. Shanholtz, and C. Hagedorn. 2000. Bacterial source tracking: a tool for total maximum daily load development. p.157-172. In T. Younos and J. Poff (ed.), Abstracts, Virginia Water Research Symposium 2000, VWRRC Special Report SR-19-2000, Blacksburg. Lee, M.-K., Saunders, J. A., Morton, C., and Feng, Y., 2000. The biogeochemistry of heavy metals under artificial sulfate-reducing conditions, American Geophysical Union Fall Meeting Abstracts, p. 285, San Francisco, California, December 15-19. Peach, A.E., J.J. Fuhrmann and D.A. Zuberer. 2000. Influence of mine reclamation on microbial biodiversity. Agron. Abst. Pg. 261. Minneapolis, MN. Reynolds, C. M., B. A. Koenen, L. B. Perry, K. L Foley, D. B. Ringelberg, and K. J. McCarthy. 2001 . Phytoremediation in Korea: Evidence for Rhizosphere-Enhanced PAH Degradation Sixth International Symposium, In-Situ and Onsite Bioreclamation. June 4-7, San Diego CA. Battelle Press. (In Press) Reynolds C. M, L. B. Perry, B. A. Koenen, K. L. Foley, D.C Wolf, and K. J. McCarthy 2000. Phytoremediation in Alaska and Korea. EPA - Phytoremediation: State of Science Conference. Proceedings of a Conference held in Boston MA: May 1-2. Reynolds, C.M. and B.A. Koenen, 2000. Rhizosphere-Enhanced Remediation Project at Osan Air Base, South Korea, April 2000. CONXXX. Reynolds, C.M. and B.A. Koenen, 2000. Rhizosphere-Enhanced Remediation Project at Kunsan Air Base, South Korea, April 2000. CONXXX. Skipper, H. D., J. H. Kim, K. Xiong, D. T. Gooden, and T. L. Lalande. 2001. Diversity of Rhizobacteria from Weeds. Abstracts, p. 26. Weed Science Society of America, Lawrence, KS. Thoma, G.J., T.B. Lam, E.N. Dempsey, D.C. Wolf, and C.A. Beyrouty. 2000. A mathematical model of phytoremediation of oil contaminated soil. In Proc. Seventh International Petroleum Environ. Conf. Albuquerque, NM. 7-10 Nov. 2000. Integrated Petroleum Environmental Consortium, Tulsa, OK. White, Jr., P.M., D.C. Wolf, and G.J. Thoma. 2000. Influence of soil amendments on plant growth in a petroleum-contaminated soil. p. 10. In Volume 4 Abstracts Arkansas Crop Protection Assoc. 30 Nov. - 1 Dec. 2000. Fayetteville, AR. Zuberer, D.A., J. Brien and A. Fincher. 2000. Water-extractable carbon in sand-based rootzones of dwarf bermudagrasses. Agron. Abst. Pg. 256. Minneapolis, MN.

Impact Statements

Studying biogeographic aspects of land-applied wastes would be useful to the state and federal agencies trying to determine the host origin of nonpoint pollution sources and would be useful to water resource managers writing TMDL implementation plans.
Characterization of microbial diversity of rhizosphere microorganisms involved in bioremediation of contaminated soils will improve our ability to use plant-microbial systems to give optimal in situ decomposition at contaminated sites with minimum cost and management.
Research on microbial diversity in disturbed landscapes and urban environment will provide a basis for achieving better plant-microbial systems to sustain ecosystem functions.

Date of Annual Report: 08/15/2002

Report Information

Annual Meeting Dates: 06/12/2002 - 06/14/2002
Period the Report Covers: 10/01/2001 - 09/01/2002

Participants

Albrecht, Steve (Stephan.Albrecht@orst.edu)- USDA-ARS, Oregon, chair; Angle, Scott (ja35@UMAIL.UMD.EDU) - University of Maryland; Bryant, Tamara (TAMBOKITTY@aol.com) - University of New Hampshire, guest; Deng, Shiping (deng@mail.pss.okstate.edu) - Oklahoma State University; Entry, Jim (JEntry@KIMBERLY.ARS.PN.USBR.GOV) - USDA-ARS, Idaho; Godfrey Dominique (epidemic0072001@yahoo.com) - University of Georgia, guest; Hagedorn, Charles (Chuck) (chagedor@vt.edu) - Virginia Tech; Hartel, Peter (pghartel@imap.arches.uga.edu) - University of Georgia; Jubb, Gerald (Skip) (jubbg@vt.edu) - Virginia Tech/Virginia Ag Experiment Station, acting Administrative Advisor; Hickey, William (wjhickey@facstaff.wisc.edu) - University of Wisconsin; Jenkins, Michael (mjenkins@imap.arches.uga.edu) - USDA-ARS, Georgia; Kuntz, Robin (rlkuntz@arches.uga.edu) - University of Georgia, guest; Nakatsu, Cindy (cnakatsu@purdue.edu) - Purdue University; Reilly, John (jpr11@cornell.edu) - Cornell University, guest; Savin, Mary (msavin@uark.edu) - University of Arkansas; Sylvia, David (DMSylvia@MAIL.IFAS.UFL.EDU) - University of Florida; Thies, Janice (jet25@cornell.edu) - Cornell University, secretary and host; Wang, Ping (wang.566@osu.edu) - Ohio State University, guest; Zuberer, David (dzuberer@taexgw.tamu.edu) - Texas A&M University

Participants Attachment:

Attachment

Brief Summary of Minutes

Skaneateles, NY

Steve Albrecht, 2002 Chair, welcomed all participants to the meeting and brought the meeting to order.

Janice Thies, local organizer, presented the agenda and explained the local arrangements.

Administrative Report - Gerald (Skip) Jubb

Skip Jubb presented the Administrative Report. Unfortunately, Kriton was unable to join us due to a recent illness. As funding is getting tighter all the time, Skip advised us to emphasize high project quality and tangible outcomes. We are to work only on our defined objectives and to stay focused. Our annual report must be posted within 60 days of this meeting on the new National Information Management Support System (NIMSS). Format of final report is responsibility of the Chair (Steve Albrecht). David Sylvia manages the web page and Mike Mullen hosts the listserv for email.

Questions/Discussion:

It was apparent to all participants that funding for travel to the Annual Meeting is not managed equitably between institutions. The formula, however, is set by the states; therefore there is no way to ‘normalize‘ it.

The group acknowledged with sadness the passing of Bob Klucas, who joined the group in the early 1980s and contributed to it for over 20 years.

The remainder of the day was given over to group reports from all three objectives.

Peter Hartel - Obj #1

Peter provided an update of the work in Georgia on bacterial source tracking. Key points were:

1. The Hartel group has a laser focus on Obj #1 and has been very successful in obtaining extramural funding.

2. GA/ID collaborative E. coli work will appear in the Jour. of Environmental Quality.

3. Microbial source tracking -focus has been on E. coli, however, enormous total diversity and temporal variability is forcing them to look for another indicator - E. faecalis is looking promising as it is essentially only found in humans, wild birds and poultry.

Dominique Godfrey - Obj #1

Dominique presented a detailed report of this year‘s outcomes by the Georgia group. The primary data are contained in their Annual Report. Two key points were:

1. The greatest weakness for microbial source tracking is SAMPLING. The Georgia group is now undertaking intense targeted sampling. E. faecalis is the focus organism.

2. The fly in the ointment is that the fundamental premise for the use of indicator organisms is that they do not persist in the environment. This is likely not true for E. coli.

Questions and discussion for ALL reports are given in detail in the Meeting Minutes located on the S-297 website.

BREAK - Discussion on European Collaboration

The group was in favor of increasing collaboration overseas and developing new links. It was suggested that the Annual Meeting might be held in Europe in 2 years to facilitate integration and collaboration with European colleagues.

Jim Entry - Obj #1

Jim reported on his collaborative work with Peter Hartel on tracking E. coli in the Rock Creek catchment in ID. Limited data are available, however, contamination appears of human origin, not cattle. They will now apply intense targeted sampling. Jim has 3 years funding to do this. He will be sending the ID samples to the riboprinter in GA.

Chuck Hagedorn- Obj #1

Chuck reported on their work in the Chesapeake Bay. Antibiotic resistance, Biolog and PFGE are being used as tracking tools. They have found that some species of Enterococcus are related to certain hosts and not others and that the use of phenotypic markers only yields broad categories. They have been doing monthly sampling of E. coli and Enterococcus for one year, and have picked up several storm events. A human signature is seen during the wet season. Their aim is to contribute to TMDLs. To date, only 1/3 of state‘s waters have been evaluated. Chuck presented a poster at ASM and has a paper coming out in JEQ shortly on this work.

Scott Angle - Obj #2

Scott spent 5 mos. at Univ. of Melbourne with Alan Baker. He taught a joint class (MD and Aus) on phytoremediation, which was recorded on a CD. Scott is studying rhizosphere ecology of hyperaccumulators and looking for potential inoculant organisms. Different plants are being examined. He has isolated 250 bacteria - mostly pseudomonads, and has found 4 that increased uptake of Ni considerably. One that increases uptake by over 30%. Scott also reported on his work in constructed Rain Guard systems in which plant heavy metal hyperaccumulators are planted to take up Zn from tires. Anaerobic denitrification, C decomposition and decreased erosion are other benefits of the Rain Guards.

Cindy Nakatsu - Obj #2

At Pudue, remediation work is focusing on site assessment. Real-time PCR is being used to target aerobic oxygenase genes. A remediation method is put in place and then evaluated for increasing degradation. They are following gene copy number to see if it increases over time. They are also monitoring wells. This work is soon to be published.

David Zuberer - Obj #2

David reported on their work on water-soluble C in turf grass (urban Agriculture), where they are particularly interested in C dynamics and bioclogging in sand-based root zones. Although the greens‘ design is aeration and drainage friendly, they find a black zone developing after a few years. ‘Snake oil‘ products, that are being sold to greens-keepers as cure-alls, were discussed extensively.

Janice Thies - Obj #1 and #3

Cornell University hosted a workshop to train S-297 participants in the denaturing gradient gel electrophoresis (DGGE) and terminal restriction fragment length polymorphism (TRFLP) analyses for soil microbial community analysis 6/8-11/02.

Cornell report - Federal Formula Funds funding has been obtained to begin collaborative work in Obj #1 with Hartel/Hagedorn/Emerson et al., beginning 10/1/02.

Bill Hickey - Obj #2

Bill reported on his work in characterizing PAH degrading bacteria isolated from humic acid (HA) solutions. He is studying the bioavailability of phenanthrene, which is sorbed by OM and has a low aqueous solubility. The hypothesis being tested is that enrichment systems with sorbed PAH may yield organisms relevant to biodegradation in soil. REP PCR fingerprints and 16S sequence analysis are being used to characterize PAH degraders. Mechanisms of degradation are also being studied.

David Sylvia - Obj #3

Dave reported on studies in dune stabilization. They have examined compatible plant/mycorrhizal fungus ecotype combinations from micro-propagated sea oats planted on FL beaches. Fungal diversity is being characterized by large subunit rRNA gene amplification and sequencing at four geographically diverse sites. They have found:

1. as much intra-site variability as inter-site variability.

2. a range of symbiotic effectiveness within the beach dune system, which affords the opportunity for selection of superior AM fungal-host combinations.

3. the origin of host and AM fungi has little predictive value in screening these combinations.

Shiping Deng - Obj #2

Shiping is studying nitroaromatic contamination and associated degrading bacteria in TNT contaminated soils. She has been conducting bioreactor studies with engineers. TNT-RDX-HMX - remediation sometimes works, sometimes not. The impact of contamination on soil N pools and the microbial community are being investigated. Microbial biomass, enzyme activity and total recoverable bacteria are all seriously affected in contaminated soils. Controlled studies are also in progress in which TNT has been added to healthy soils and the community response measured. DGGE and 16S sequencing of isolates are being conducted.

Textbook Committee - David Sylvia

Principles and Application of Soil Microbiology - 2nd Edition. David reviewed for us the proposed chapters and authors. These can be found on the S-297 website.

Steve Albrecht -

Steve is studying organic matter (OM) dynamics in varying tillage systems in Eastern OR, where low soil OM is the primary problem. They have found that:

1. Tillage that returns residue and low-till practices increase soil OM.

2. Light fraction (sodium iodide or sodium tungstate) centrifugation is correlated with reduced tillage operations.

Steve has also been working on:

1. building a model for C sequestration,

2. examining microbial diversity in different tillage operations, and

3. N mineralization studies post manure application in sandy soils in areas new to cattle production.

Ping Wang-

Ping reported on tillage trials in OH (long term no-till for 45 y). TRFLP is being used to compare community structure between tillage treatments. They are also monitoring animal pathogens by examining leachate for E. coli O157:H7 and Cryptosporidium. NOTE: Warren Dick - message delivered by Ping Wang. Warren would be glad to develop a proposal or cooperate with like-minded researchers. His projects can be linked into all 3 objectives.

Chuck Hagedorn-

Announced the newsletter devoted to bacterial source tracking "Environmental Detection News." Copies of the first issue were distributed. The newsletter is meant to be a more informal information-sharing medium.

Textbook discussion

David Sylvia reported that they expect to have the new edition on the shelf for Fall 2003. It should be to Prentice Hall by end of this year. The Laboratory manual is also in the works. Prentice Hall will not be the publisher. Dave requested sharing of exercises in our courses that might be useful, including molecular exercises. The 2nd edition of the textbook will be dedicated to the memory of Bob Klucas.

Election of officers and selection of next year‘s meeting site

Janice Thies is the chair for 2002-03. The chair-elect position was open for election. There were two self-nominations: Jim Entry and Mary Savin. These nominations were both seconded by Cindy Nakatsu. The vote was taken by a show of hands. Mary Savin was elected by a vote of 10 to 0.

Next year‘s meeting site:

Next year‘s meeting will be held in Puerto Rico, June 2-4, 2003. Eduardo Schroeder will be the local host in concert with Peter Hartel. Other members of the planning committee are: Jim Entry and Robin Kuntz.

Discussion on future collaborative grants - whole group

The following ideas and funding sources were discussed:

Bacterial source tracking:

1. National Park Service, Yellowstone, is interested.

2. Coastal Zone management, the Sea Grant Program , 319 Funds and EPA.

Biosafety issues:

1. The Military (DOD) - Camps need baselines established.

2. The STAR program

3. Local/State Water Quality initiatives.

Other funding sources: NSF - Microbial Observatories Program.

We all wish Kriton the very best. On behalf of group we wish him a speedy recovery.

Where does S-297 stand in the Multistate program?

We would appreciate if the station directors were to provide feedback of a substantive nature. We would also deeply appreciate some encouragement and support for the work we are doing - if they think our research is worthy of it. Administratively, how do we remove participants that are no longer active? Peter Hartel proposed that if no progress report has been received in the last 2 years, which is current project‘s duration, then remove the participant from the list. In order to stay on, they must submit a progress report within the next 30 days. Steve Albrecht will communicate this in conjunction to soliciting input to the annual report. He will also contact Skip Jubb to clarify the membership list of record and email the current list to all participants.

The chair, Steve Albrecht, thanked our host, Janice Thies, for making the arrangements for this year‘s meeting.

Move to adjourn: J. Thies

Second: D. Zuberer

Meeting was adjourned at 12:01 pm

Accomplishments

A short-course on ‘Molecular Techniques for the Analysis of Microbial Community Structure in Soil and the Rhizosphere‘ (hosted by J. Theis in collaboration with Cindy Nakatsu [Purdue University] and Terry Marsh [Michigan State University] at Cornell Univ., 6/8-11/02) was taught for the S-297 principal investigators, their students and technicians. Objective 1: In order to improve water quality where high fecal coliform numbers have been detected, Escherichia coli has been commonly used as a target organism to determine the potential sources. In addition to investigations of E. coli, we are also studying other bacteria (e.g. Enterococcus faecalis) found in sewage and manure. Ribotyping the microorganisms is used by many laboratories participating in this Project to differentiate between microorganisms found in human sewage and microorganisms found in either wild or domestic animals. Research in Georgia found that human sewage samples contained Ent. faecalis, but high percentage of broiler litters contained no Ent. faecalis (some contained low numbers). Percentages of Ent. faecalis were high in screech owl and wild turkey, but were low or zero in great horned owl, pelican, robin, and sea gull. No enterococci were observed in Canada goose, dove, and pigeon. The initial isolation media had no effect on isolating Ent. faecalis. Chickens had Ent. faecalis, but the litter strongly selected against the organism, suggesting that broiler litter is not an important source of Ent. faecalis to the environment. The presence of Ent. faecalis in individual species of wild birds was variable. Two watersheds in Western Puerto Rico are being examined for fecal coliforms. Initial sampling of a portion of the Yaguez River (PR) indicated high numbers of enterobacteria. Work is continuing to determine the source of E. coli contamination in Rock Creek area and several other streams in southern Idaho. Two hundred and forty ribotypes were identified from 451 E. coli isolates. fecal samples of two herds of Black Angus cattle in Georgia. Isolates from Idaho and Georgia will establish temporal variability, whereas isolates from Georgia and Puerto Rico will establish geographic variability. An E. coli ribotype database is being developed for the state of Delaware using isolates recovered from various hosts (wildlife, domesticated animals, and humans) A large project on the Mill Creek Watershed (Montgomery County, VA) to determine source-tracking classifications (human vs. livestock vs. wildlife) as a function of sampling frequency was completed. The project was performed on both E. coli and Enterococcus, with antibiotic resistance analysis, carbon source utilization, and pulsed-field gel electrophoresis as source-tracking tools. Cooperative work (J. Harwood) will compare results to those obtained with ribotyping. The high ARCC of the large library demonstrates that isolates from different regions can be put together and it is not necessary to build a new large library with every new watershed. There is the distinct possibility that livestock so overwhelmed all other sources, when the cattle inventory in Mill Creek (approx. 3,800 animals) was considered, that the lack of obvious seasonal variation regarding livestock was simply a function of the predominant land use pattern devoted to pasture and cattle in the Mill Creek watershed. Two east Tennessee watersheds have generally characterized with respect to total coliforms, E. coli, fecal coliforms and enterococcus. Upcoming plans are to look at impacts of Best Management Practices on stream bacteriological quality. In addition to ribotyping, other genetic fingerprinting methods to differentiate microorganisms, including E. coli, in the environment are being compared. Methods being tested are: rep-PCR (repetitive sequences, using both REP and BOXA1R primers, using the Polymerase Chain Reaction), AFLP (Amplified Fragment Length Polymorphism), PFGE (Pulsed-Field Gel Electrophoresis), and ribotyping. The accuracy of these various methods is now being tested more extensively on animal fecal sources. E. coli populations of animals from different facilities in Indiana are being examined to determine genetic variation between isolates from known hosts collected at different facilities in a limited geographic region. And differences in genetic fingerprint patterns of isolates collected from pig manure applied to agricultural fields and those collected from underlying tile drains after transport through the soil profile will be examined. Objective 2: Research in Maryland showed that several microbial isolates from the rhizosphere of Alyssum murale were capable of increasing Ni uptake into the plant. Field trails using these associations will be conducted next year. The impact of 2,4,6-trinitrotoluene (TNT), hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), and octahydrol-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) contaminations on the chemical and microbiological properties of soils in Oklahoma was evaluated. A technique based on real time PCR amplification of aromatic oxygenase genes was developed to detect and quantify a number of aromatic catabolic genes in environmental samples. Groundwater samples from gasoline-contaminated sites in Indiana were studied. In field samples, aromatic oxygenase genes were detected in groundwater monitoring wells with current or recent petroleum contamination but not in wells without a history of contamination, confirming that this technology is appropriate for monitoring pollutant biodegradation. In Alabama strains of Pseudomonas putida and Rhodococcus erythropolis were found to degrade soil-sorbed biphenyl. Results suggest that cell attachment ability was important for the access of soil-sorbed biphenyl and that chemotaxic ability was significant in scavenging sorbed biphenyl. The effect of fertilizer addition and vegetation establishment on remediation of crude oil-contaminated soil was studied in Arkansas. Fertilizer addition and vegetation establishment significantly increased total bacterial and polycyclic aromatic hydrocarbon (PAH) degrader numbers. The fertilized-vegetated plots had significantly greater reductions in TPH levels as compared to the control. The plots are established and will continue to be monitored over the next several years to determine the effects of nutrient additions and vegetation on remediation and microbial ecology. Greenhouse studies are continuing to provide useful information related to selection of plant species suitable for establishment at crude oil-contaminated sites. The effect of compost layers on the retention and transport of atrazine and P were determined in several Homestead, FL area soils. A column study demonstrated that atrazine and P leached out slower in soil with compost than in soil without compost. Objective 3: The effect of crop rotation on rhizobacterial diversity in a Norfolk soil (South Carolina) was studied for three years. Root bacteria were identified by GC-FAME analysis. Biochemical assay and PCR technology were used to evaluate the denitrification potential for 200 rhizobacteria. Results from research in Indiana have demonstrated that microbial community fingerprinting techniques (16S rDNA PCR and DGGE) can be used to show differences in the microbial community structure influenced by the environment. The rhizosphere microbial community of corn plants was investigated in order to determine if the same populations are affected by different agronomic treatments over different years. Cluster analysis, using principal components analysis (PCA), revealed that in all three years the microbial community profiles grouped according to agronomic treatment. This indicates that agronomic treatment can influence the microbial ecology of the rhizosphere in corn plants. Experiments in Idaho evaluated genetic bacterial diversity and determined that the amount of microbial DNA extracted from soil correlated with soil C in a positive curvilinear relationship. Also, the amount of microbial DNA extracted from soil correlated with active bacterial biomass, active fungal biomass and active microbial biomass in positive curvilinear relationships. A research project is underway in Puerto Rico, in cooperation with a farmer, to study the effect of herbicides on soil biomass. Preliminary data indicate that the most probable number (MPN) determination of Rhizobium is affected by the plant species that predominates in the plots. Further data on MPN populations in these plots will be continued. Differential responses in citrus genotypes grown in Florida suggest that C expenditure on arbuscular mycorrhizal (AM) fungus - Glomus intraradices symbiosis is more tightly regulated in genotypes of low mycorrhizal dependency (MD), such as sweet orange, than in genotypes of high MD (e.g. sour orange). Further characterization of mycorrhizal effects on the carbon economy of citrus and the release of C from roots into the soil is planned. Also in Florida, research was conducted to test the level of functional diversity that exists among communities of AM fungi that are present in divergent Florida dunes. One study evaluated intra-location responses of plant and fungal genotypes on growth response of sea oats while a second study evaluated inter-location responses of plant and fungal ecotypes using the best combinations from experiment one. The research showed that there was a degree of specificity among hosts and their symbiotic fungi; however, relative to plant growth response, one fungal community (SG-1) was superior across plant ecotypes from all locations. Conversely, one plant ecotype (EK-17-3-2) tended to have the best response across all fungal communities. Progress on a second edition of the textbook, Principles and Applications of Soil Microbiology, and a laboratory manual to accompany it, has continued.

Publications

Angle, JS, et al. 2001. Developing commercial phytoextraction technologies: Practical considerations. S Afr J Sci 97:619-623. Chang, Y-J, et al. 2001. Impact of herbicides on abundance & structure of indigenous _-subgroup ammonia oxidizer communities in soil microcosms. Environ Toxicol Chem 20:2462-2468. Delorme, T, et al. 2000. P accumulation by select plant species. J Phytoremediation. 2:344-349. Gagliardi, JV, et al. 2001. Structural & functional analysis of whole-soil microbial communities for risk & efficacy testing following microbial inoculation of wheat roots in diverse soils. Soil Biol Biochem 33:25-40. Haney, RL, et al. 2001. Molar concentration of K2SO4 & soil pH affect estima of extractable C with chloroform fumigation-extraction. Soil Biol Biochem 33:1501-1507. Hartel, PG, et al. 2002. Biogeographic variability of E. coli ribotypes from ID & GA. J Environ Qual 31:1273-1278. Jarstfer, AG, DM Sylvia. 2001. Isolation, culture & detection of arbuscular mycorrhizal fungi. p 535-542. Man of Environ Microbiol. 2nd Ed ASM Wash, DC. Jifon, JL, et al. 2002. Growth depression of mycorrhizal citrus seedlings grown at high P supply is mitigated by elevated CO2. New Phytologist 153:133-142. Kubikova, et al. 2001. Mycorrhizal impact on osmotic adjustment in Ocimum basilicum during a lethal drying episode. J Plant Physiol 158:1227-1230. LaPara, TM, et al. 2001. Aerobic biolog treatment of pharmaceutical wastewater: Effect of temperature on COD removal & bacterial community development. Water Res 35:4417-4425. Maldonado, JD, et al. 2001. Arbuscular mycorrhiza inoculum potential in natural & managed tropical montane soils in Costa Rica. Trop Agric 77:27-32. Rickman, RW, et al. 2001. CQESTR: A model to estimate carbon sequestration in agric soils. J Soil Water Conser 56:237-246. Schutter, ME, JJ Fuhrmann. 2001. Soil microbial community responses to fly ash amendment as revealed by analyses of whole soils & bacterial isolates. Soil Biol Biochem 33:1947-1958. Sigler, WV, et al. 2001. Fate of biolog control agent Pseudomonas aureofaciens after application to turgrass. Appl Environ Microbiol 67:3542-3548. Sylvia, DM, et al. 2001. Arbuscular mycorrhizal fungi influence tomato competition with bahiagrass. Biol Fertil Soils 34:448-452. Sylvia, DM, DO Chellemi. 2001. Interactions among root-inhabiting fungi & their implications for biolog control of root pathogens. Adv Agron 73:1-33. van Berkum, P, JJ Fuhrmann. 2001. Characteristics of soybean bradyrhizobia for which serogroup affinities have not been identified. Can J Microbiol 47:519-525. Wagner, SC, et al. 2001. Long-term survival of Glomus claroideum propagules from soil pot cultures under simulated conditions. Mycologia 93:815-820. Wilbur, JD, et al. 2001. Statistical issues in the analysis of microbial communities in soil. Proc Appl Stat in Agric. KSU, Manhattan, KS. Wheeler, AL, et al. 2002. Potential of Enterococcus faecalis as a human fecal indicator for microbial source tracking. J Environ Qual 31:1286-1293. Zuberer, DA 2001. N fixation (nonsymbiotic) in soils. Ency of Envir Micro. John Wiley & Sons, NY. Becker, J, et al. 2001. Bacterial community diversity & activity at a mixed waste contaminated site. ASM Abst. Bezdicek, D, M Fauci, S Albrecht. 2002. Soil carbon & C sequestration under different cropping & tillage practices in the Pacific NW. p 101-107 Proc NW Direct Seed Cropping Systems Conf. WA. Booth, AM, C Hagedorn. 2001. Determining sources of fecal pollution in Blackwater River watershed, Franklin Co, Va. Crop & Soil Environ News. HTTP://www.ext.vt.edu/news/periodicals/cses/2001-11/franklincounty.html. Bowman, AM, K Mentz, C Hagedorn. 2001. Determining sources of fecal pollution in Blackwater River, Franklin Co, VA. ASM Abst. Bulinski, DA, et al. 2001. Rhizosphere microbial population dynamics over 3 field seasons. ASA Abst. Carrero-Colsn, et al. 2001. Microbial community response to perturbation of a model wastewater treatment system. ASM Abst. Chaney, RL, et al. 2001. Heavy metal aspects of compost use. Compost Utilization in Hort Cropping Syst. CRC Press, FL. Curtis, P, CH Nakatsu, A Konopka. 2001. Acid tolerance of chemoheterotrophic bacteria isolated from an acidic (pH 3) soil. ASM Abst. DeQueiroz, GA, et al. 2001. Viability & nodulation with Bradyrhizobium cultures after 10.5 years storage in sterile distilled water. ASA Abst. Entry, JA, et al. 2001. Ribotyping of E. coli isolates from Rock Creek Watershed, ID. ASA Abst. Entry, JA, RE Sojka. 2002. Polyacrylamide removes coliform bacteria from animal wastewater. p 1-3. Res & Ext Regional Water Qual Conf. WA. Feng, Y, et al. 2001. Tillage effects on soil microbial community composition in continuous cotton syst. ASA Abst. Frederick, JR, et al. 2001. Rhizobacteria from soybean & corn in rotation. ASA Abst. Graham, JH, et al. 2002. Growth depression of mycorrhizal citrus seedlings grown at high P supply is mitigated by elevated CO2. 3rd Int Conf Mycorrhizas. Australia. Gooden, DT, et al. 2001. Rhizobacteria from peanuts in rotation. ASA Abst. Graves, AK, M Mahal, C Hagedorn. 2001. Determining sources of fecal pollution for a rural unsewered community. ASM Abst. Hartel, PG, JD Summer, WI Segars. 2002. Potential for ribotyping to delist watersheds exceeding their TMDLs for fecal coliforms. Watershed Mgmnt to Meet Emerging TMDL Environ Regs. TX. Hartel, PG. 2002. Ribotyping enterococci. p 11-15. Microbiol Source Tracking Wkshop. CA. Hagedorn, C, et al. 2002. Carbon source utilization profiles as a method to identify sources of fecal pollution in water. ASM Abst. Hagedorn, C. 2002. Carbon source profiles, pulsed-field gel electrophoresis & antibiotic resistance analysis. p 55-58. Microbiol Source Tracking Wkshop. CA. Haines, SK, SP Deng. 2001. Isolation & characterization of bacteria capable of biodegrading explosives characterization of nitroaromatic-degrading bacteria isolated from explosive contaminated soils. ASA Abst. Haney, RL, AJ Franzluebbers, FM Hons, DA Zuberer. 2001. Is K2SO4 extraction of microbial carbon pH dependent? ASA Abst. Haney, RL, et al. 2001. Soil CO2 evolution: Is drying & rewetting substrate induced respiration? ASA Abst. Haney, RL, et al. 2001. The flush of CO2: drying & rewetting vs. chloroform fumigation. ASA Abst. Homhaul, W, J Fuhrmann, P van Berkum. 2001. Polyphasic analysis of soybean & cowpea bradyrhizobia isolated from US and Thai soils. ASA Abst. Hons, F, J Thomas, D Zuberer. 2001. Chem, phys & biol properties of turfgrass soils. CD ROM. Soil & Crop Sci, Tx A&M U. Jerke, K, CH Nakatsu, A Konopka. 2001. Genetic & physiological analyses of lead resistance in Arthrobacter sp. isolated from contaminated soil. ASM Abst . Kirkpatrick, WD, PM White, Jr, GJ Thoma, EE Gbur, CM Reynolds, DC Wolf. 2001. Plant response to N addition in a crude oil-contaminated soil. ASA Abst. Konopka, A, et al. 2001. Microbial community structure & function in a biomass-recycle bioreactor. 9th Int Symp Microbial Ecol. Netherlands. Kuntz, RL, et al. 2002. Field-testing Enterococcus faecalis for microbial source tracking. ASM Abst. Mesa, C, J Fuhrmann, P van Berkum. 2001. Phenotypic & genetic analysis of soybean bradyrhizobia isolated from DE soils. ASA Abst. Motta, ACV, et al. 2001. Mgmnt systems to improve soil quality for cotton production on a degraded silt loam soil in AL (USA). p 219-222. Proc 1st World Cong on Conserv Agric. Spain. Motta, ACV, et al. 2001. Enhancement of soil microbial biomass in cotton production systems with conserv tillage. 24th Ann So Conserv Tillage Conf Sust Agr. Nakatsu, CH, J Joynt, A Konopka. 2001. Impact of metal & organic waste on a microbial community. 9th Int Symp Microbial Ecol. Netherlands. Nakatsu, CH, et al. 2001. Chromium resistance by Arthrobacter sp. cr15 isolated from metal contaminated soil. ASM Abst. Ravnskov, S, Y Wu, JH Graham. 2001. The influence of different arbuscular fungi & P on gene expression of invertase & sucrose synthase in roots of maize. 3rd Int Conf on Mycorrhizas. Australia. Reynolds, CM, et al. 2001. Microbial populations during remediation at 3 petroleum-contaminated Alaskan sites. In Situ and On-Site Bioremediation. 6th Int Symp. CA. Rommesser, J, JJ Fuhrmann. 2002. A method to assess quorum sensing potential of microbial communities. Pat (prov) Appl #60/346,531. Thies, JE, T Marsh, C Nakatsu. 2002. Workshop Manual: Molecular Techniques for Analysis of Microbial Community Structure in Soil & the Rhizosphere. Cornell U, Ithaca, NY. Thoma, GJ, et al. 2001. Mathematical modeling of phytoremediation of petroleum contaminated soils. 8th Ann Int Petro Environ Conf TX. Thoma, GJ, et al. 2001. A mathematical model of phytoremediation of crude oil-contaminated soils. In Situ & On-Site Bioremediation. 6th Int Symp. CA. Wang, G, HD Skipper. 2001. Denitrifying rhizobacteria from bentgrass & bermudagrass USGA greens. ASA Abst. Wheeler, AL, et al. 2001. Ribotyping fecal streptococci to differentiate between human & nonhuman sources. ASA Abst. White Jr, PM, et al. 2001. Field study to evaluate phytoremediation of crude oil-contaminated soil. ASA Abst. White Jr, PM, WD Kirkpatrick, DC Wolf, GJ Thoma, RM Reynolds. 2001. Phytoremediation of crude oil-contaminated soil. 8th Ann Int Petro Environ Conf. TX. Wilbur, JD, et al. 2001. A bayesian approach to modeling & variable selection in high-dimensional multivariate binary data. 2001 Joint Stat Mtgs. Wilbur, JD, et al. 2001. Variable selection in high-dimensional multivariate binary data. New Eng Stat Symp. Wu, G, Y Feng, SA Boyd. 2001. Effects of biphenyl-degrading bacteria characteristics on bioavailability of sorbed biphenyl. ASA Abst. Zurdo-Piqeiro, JL, et al. 2002. Identificacisn de cepas de rhizobia que nodulan leguminosas tropicales en suelos de Puerto Rico. IX Nat Mtg on N Fixation. Spain.

Impact Statements

Training in modern molecular techniques, that can be used to characterize soil microbial population structure, was offered to participants of S-297 Multistate Project. This will significantly enhance the analytical capacity and capability of 8 laboratories that participated.
Obj 1: Research on Enterococcus faecalis suggests that the restricted host range of this organism may provide an easier and better way to distinguish human versus non-human fecal contamination than Escherichia coli. This information will be useful to water resource managers writing TMDL (Total Maximum Daily Load) implementation plans.
Both the apparent turnover of E. coli ribotypes and a clonal diversity index of 0.97 indicates an extensive ribotype variability implying the necessity of ribotyping a large number E. coli isolates per host to establish a host origin database that is independent of temporal variability, or complete enough to be effective.
Regulatory agencies are interested in the sampling number and frequency required to accurately determine sources of fecal pollution, as that determines project cost. While statistical analyses are incomplete, they indicate that the monthly sampling conducted at the Mill Creek Watershed was adequate, that seasonality is present and sampling must account for seasonal high and low flow conditions. This information will be very useful to water resource managers writing TMDL implementation plans.
Developing bacterial TMDLs for Delaware's surface waters is a high priority item for DNREC. The E. coli database for Delaware is a key milestone in meeting UDEPA mandates for ameliorating bacterial contamination of waters.
Canal systems transfer irrigation water from the Snake River to crop producing fields and dairy operations in the area and then empty into the river as
Early results of genetic fingerprint analyses using REP primers indicate that the majority of strains (~70%) collected from a single animal-type have identical patterns that are visually distinct. This information will contribute to the identification of sources of fecal coliform organisms.
Obj 2: Information on the utilization of compost to calcareous soils to reduce the leaching potential of atrazine and P should help reduce groundwater contamination in Florida and improve water quality.
Increasing Nickel uptake by the plant Alyssum via microbial inoculation is significant and should lead to increased profits of phytomining.
Understanding the bacterial characteristics that facilitate the degradation of soil sorbed contaminants will help us elucidate the mechanisms of the accessibility of sorbed contaminants and design unconventional enrichment culture conditions that favor the isolation of effective strains for use in bioremediation.
Obj 3: The use of culture-independent methods, such as PLFA profile analysis, being developed or modified in this project will offer improved techniques to characterize the changes of microbial community under different management systems and should provide insights into how conservation tillage improves soil quality and sustainability.
There may exist a widespread potential for mycorrhizas to produce negative crop responses because the fungi are no longer behaving as mutualists. The information gathered should allow for the evaluation of the potential association facilitate the establishment of symbiotic associations and reduce the possibility of negative crop responses.
Long-term storage of bradyrhizobia in sterile-distilled water, a straightforward technology, can be used as a very effective and inexpensive alternative to other more expensive methods of storage.
A database being developed on rhizobacteria isolated from soybean, related crops, and weeds that will assist in ecological studies in sustainable agriculture. For example denitrification may account for 20 to 30% loss of fertilizer nitrogen in turf. The identification of rhizobacteria responsible for denitrification is essential to develop management strategies to conserve nitrogen.
Results from research in Florida to help stabilize beaches utilizing sea oats confirm that a range of symbiotic effectiveness exists within the beach dune system and afford the opportunity for selection of superior AM fungal-host combinations from each location. It is possible that while it is beneficial to select superior AM fungal-host combinations for outplanting, origin of host and AM fungi have little predictive value in screening these combinations.
Agronomic treatments can influence the microbial ecology of the rhizosphere of corn plants. This information will be insightful in identifying factors influencing monoculture yield decline in corn and should contribute to the development of a comprehensive means to identify soil biota that influence crop productivity under different management conditions.

Date of Annual Report: 07/26/2004

Report Information

Annual Meeting Dates: 06/06/2004 - 06/07/2004
Period the Report Covers: 06/01/2003 - 06/01/2004

Participants

Mary Savin, University of Arkansas, chair;
Peter Hartel, University of Georgia, chair-elect/secretary;
Steve Leath, North Carolina State University, Administrator;
Steve Albrecht, USDA-ARS, Pendleton, OR, meeting host;
Jim Graham, University of Florida;
Mike Jenkins, USDA-ARS, Watkinsville, GA;
Rick Weaver, Texas A & M;
Peter Tomlinson, University of Arkansas;
Janice Thies, Cornell University, NY;
Shiping Deng, Oklahoma State University

Brief Summary of Minutes

See listserv (http://s-297.okstate.edu) for minutes to annual meeting.

Minutes Attachment:

Attachment

Accomplishments

Objective 1. Research for the past year has focused on: identifying sources of fecal pollution in impaired waters; geographic and temporal variability of Enterococcus spp. and subspecies; testing fundamental assumptions of microbial source tracking-survival in sediment and secondary habitats; and validating applications of the various methodologies for BST and determining ecological data that can be arrived from such information. Targeted sampling and bacterial source tracking in New York, Puerto Rico and Alabama have worked to narrow and identify sources of contamination. Like E. coli, Ent. faecalis isolates appear to be geographically and temporally variable. Because establishing a permanent host origin database to encompass this geographic and temporal variability is likely to be expensive, alternative methods of BST need to be considered. Enterococcus species and subspecies in nonsterile sediments did not survive sufficiently well in moist sediment to serve as a reservoir of long-past human and nonhuman fecal contamination. However, fecal enterococci survived 2, 30, and 60 days of desiccation in sediment. Enterococcal survival and potential regrowth during conditions of desiccation may cause problems for BST. Project evaluating different methods have shown promise. In Virginia, research has shown that the use of fluorometry in estuarine and coastal zone environments to determine the presence of optical brighteners can be used as a surrogate to detect human sources of fecal pollution. Using molecular PCR-based techniques, in Indiana, REP primers were found not to be ideal for use in microbial source tracking. The BoxAR1 primer produced a wider variety of fingerprint types than the REP primers, but most animals still had one dominant fingerprint type. A species-specific ribosomal DNA gene biosensor for rapid detection of Enterococcus faecalis from environmental samples is being developed and tested in New York. Objective 2. Surfactants could potentially increase solubility, hence bioavailability and bioremediation of organic and inorganic contaminants in soil. Research at Oklahoma showed that nonionic surfactants had little effect, while anionic surfactants generally inhibited growth of bacterial isolates tested. (Wisconsin and Iowa) Studies examined how sorption by humic acids affects bioavailability of polynuclear aromatic hydrocarbons (PAH) to PAH-degrading microbes, and assessed how PAH-degraders may adapt to overcome bioavailability restrictions. Reduced diversity of bacteria was isolated from humic acid-sorbed phenanthrene (HASP) enrichments compared with non-sorbed phenanthrene (NSP). In a study at Purdue University, an array of primers and a real-time PCR protocol was used to enumerate aromatic oxygenase genes at gasoline-contaminated sites undergoing bioremediation. Overall, enumeration of aromatic oxygenase genes indicated changes in the magnitude of oxygenase gene copy numbers and the catabolic genotypes detected as a result of common treatment technologies. Phytoremediation was used to evaluate remediation of a weathered crude oil-contaminated soil in Arkansas. Bermuda, fescue, and ryegrass were established at a field site contaminated with 2.5% by weight weathered crude oil. Vegetation and fertilizer increased total bacterial and fungal numbers compared to the control. Isolates from three long-term nitroaromatic contaminated soils have been characterized in Oklahoma. Polymerase chain reaction amplification and denaturing gradient gel electrophoresis (PCR-DGGE) results of TNT-contaminated soil indicated a shift of microbial community structure and identification of Achromobacter spp. Field studies in Alabama indicate that accelerated degradation of aldicarb seems to contribute to the loss of effectiveness in nematode control. Objective 3. Research for the past year has focused on: antibiotic resistance in environments receiving animal wastes, microbial dynamics in disturbed lands and urban landscapes, and ecological associations. On-going collaboration between the University of Arkansas, Cornell University, Purdue University and Oklahoma State University has led to investigations of how land application of organic wastes affects microbial functioning, diversity and spread of antibiotic resistance in the environment. Research in Texas includes elucidating the seasonal dynamics of microbial populations in managed sports turfs and compost-amended common bermudagrass plots, effects of slow-release nitrogen sources on biology, measuring hydrophobicity, and investigating several microbiological parameters in a cotton crop-management system. Ranges of microbial counts (7.5 to 8.0 for bacteria and 4.5 to 5.5 for fungi) appear to be quite common across all turfgrass environments investigated. Water-extractable carbon (WEOC) showed trends very similar to those for microbial biomass and respiration suggesting it may be a good proxy for these parameters in the soils managed for cotton. Researchers in Alabama and Indiana have been studying the dynamics of resident microbial communities in order to advance understanding of the influence of microbes in productivity of perturbed ecosystems. PLFA profiles and an approach using PCR-DGGE with cultivation appear to provide a stream-lined approach to analyze agronomic treatments. Tree performance in the first two years of the apple replant disease (ARD) trial in New York indicates that use of rootstocks tolerant of ARD and planting in previous grass drive lanes have the greatest benefit. Multi-year field trials with Cry3Bb rootworm resistant corn indicate there is no direct adverse affect on soil ecology by cultivating this crop. Vermicompost benefits were found to far outweigh that of thermophilic compost in another study. Multiple studies are being conducted on disturbed soil ecosystems in Colorado: a rangeland soil impacted by biosolids amendments; a forest soil impacted by a wildfire event; and dryland agronomic soils under alternative management practices along an evapotranspiration gradient. Arbuscular mycorrhizal fungi have been shown to be sensitive to the various disturbances. The relationships between symbiotic fungi and a biotrophic root pathogen are being studied in Florida. Mycorrhizal ferns that are hyperaccumulators of arsenic took up significantly more As than nonmycorrhizal plants, and a significant interaction was found with P level. Mechanisms of enhanced carbon sequestration in tall fescue infected with the fungal endophyte Neotyphodium coenophialum as compared to tall fescue free of the endophyte is underway. In addition to these projects, a second edition of the textbook, Principles and Applications of Soil Microbiology has been published.

Publications

Augé, R.M., J.L Moore, D.M Sylvia, and K. Cho. 2004. Mycorrhizal promotion of host stomatal conductance in relation to irradiance and temperature. Mycorrhiza 14:85-92. Augé, R.M., J.L Moore, K. Cho, J.C. Stutz, D.M Sylvia, A.K. Al-Agely, A.M. Saxton. 2003. Relating drought resistance of Phaseolus vulgaris to soil and root colonization by mycorrhizal hyphae. J. Plant Physiol. 160: 1147-1156 Baldwin, B., C. H. Nakatsu, and L. Nies. 2003. Detection and enumeration of aromatic oxygenase genes by multiplex and real-time PCR. Appl. Environ. Microbiol. 69:3350-3358. Bray, S.R., K. Kitajima, and D.M. Sylvia. 2003. Mycorrhizae differentially alter growth, physiology, and competitive ability of an invasive shrub. Functional Ecology 13:565-574. Darnault, C.J.G., P. Garnier, Y.-J. Kim, K. Oveson, T.S. Steenhuis, J.Y. Parlange, M.B. Jenkins, W.C. Ghiorse, and P.C. Baveye. 2003. Transport of Cryptosporidium parvum oocysts in the subsurface environment. Wat. Environ. Res. 75:113-120. Darnault, C.J.G., T.S. Steenhuis, P. Garnier, Y.-J. Kim, M.B. Jenkins, W.C. Ghiorse, P.C. Baveye, and J.-Y. Parlange. 2004. Preferential flow and transport of Cryptosporidium parvum oocysts through the vadose zone: experiments and modeling. Vadose Zone J. 3:262-270. Davis, K. C., C. H. Nakatsu, R. Turco, S. Weagant, and A. K. Bhunia. 2003. Analysis of environmental Escherichia coli isolates for virulence genes using the TaqManâ PCR system. J. Appl. Microbiol. 95: 612-620. Devare, M, CM Jones, and JE Thies. 2004. Effects of CRW transgenic corn and tefluthrin on the soil microbial community: biomass, activity, and diversity. J. Environ. Qual. 33 (3): 837-843 and Cover photos. Feng, Y., A.C. Motta, D.W. Reeves, C.H. Burmester, E. van Santen, and J.A. Osborne. 2003. Soil microbial communities under conventional-till and no-till continuous cotton systems. Soil Biol. Biochem. 35: 1693-1703. Hahm, B.K., Y. Maldonado, E. Schreiber, A. K.Bhunia, and C. H.Nakatsu. 2003. Subtyping of clinical and environmental isolates of Escherichia coli by multiplex PCR, AFLP, rep-PCR, PFGE and ribotyping. J. Microbiol. Meth. 53:387-399. Hartel, P. G., J. D. Summer, and W. I. Segars. 2003. Deer diet affects ribotype diversity of Escherichia coli for bacterial source tracking. Water Res. 37:3263-3268. Also selected for inclusion in the ?Virtual Journal of Environmental Sustainability? (<http:// www.elsevier.com/vj/sustainability>). Harwood, V. J., B. Wiggins, C. Hagedorn, R. D. Ellender, J. Gooch, J. Kern, M. Samadpour, A. H. Chapman and B. J. Robinson. 2003. Phenotypic library-based microbial source tracking methods: efficacy in the California collaborative study. J. Water & Health 01:153-156. Jenkins, M.B. 2003. Rhizobial and Bradyrhizobial symbionts of mesquite from the Sonoran Desert: salt tolerance, facultative halophily and nitrate respiration. Soil Biol. Biochem. 35:1675-1682. Kato, S., M. Jenkins, E. Fogarty, and D. Bowman. 2004. Cryptosporidium parvum oocyst inactivation in field soil and its relation to soil characteristics: analysis using the geographic information system. Sci. Total Environ. 321:47-58. Krutz, L.J., S.A. Senseman, K.J. McInnes, D.A. Zuberer and D.P. Tierney. 2003. Adsorption and desorption of atrazine, desethylatrazine, deisopropylatrazine and hydroxyatrazine in vegetated filter strip and cultivated soil. J. Agric. Food Chem. 51:7379-7384. Kuntz, R. L., P. G. Hartel, D. G. Godfrey, J. L. McDonald, K. W. Gates, and W. I. Segars. 2003. Targeted sampling protocol as prelude to bacterial source tracking with Enterococcus faecalis. J. Environ. Qual. 32:2311-2318. Lalande, T. L., H. D. Skipper, D. C. Wolf, C. M. Reynolds, D. L. Freedman, B. W. Pinkerton, P. G. Hartel, and L. W. Grimes. 2003. Phytoremediation of pyrene in a Cecil soil under field conditions. Int. J. Phytoremed. 5:1-12. Mesarch, M. B., C. H. Nakatsu, and L. Nies. 2004. Bench-scale and field-scale evaluation of catechol 2,3-dioxygenase specific primers for use in monitoring BTX bioremediation. Water Res. 38: 1281-1288. Myoda, S. P., C. A. Carson, J. J. Fuhrmann, B.?K. Hahm, P. G. Hartel, L. A. Johnson, R. L. Kuntz, C. H. Nakatsu, M. J. Sadowsky, M. Samadpour, and H. Yampara?Iquise. 2003. Comparison of genotypic-based microbial source tracking methods requiring a host origin database. J. Water Health 1:167-180. Park, J.-H., Y. Feng, P. Ji, T.C. Voice, and S.A. Boyd. 2003. Assessment of bioavailability of soil-sorbed atrazine. Appl. Environ. Microbiol. 69:3288-3298. Ritter K. J., E. Carruthers, C. A. Carson, R. D. Ellender, V. J. Harwood, K. Kingsley, C. Nakatsu, M. Sadowsky, B. Shear, B. West, J. E. Whitlock, B. A. Wiggins, J. D. Wilbur. 2003. Assessment of statistical methods used in microbial source tracking methods. J. Wat. Health 1: 209-223. Rumberger, A, Yao, S, Merwin, IA., Nelson, EB and Thies, JE. 2004. Rootstock genotype and orchard replant position rather than soil fumigation or compost amendment determine tree growth and rhizosphere bacterial community composition in an apple replant soil. Plant and Soil. In Press (Prepublication Date: 01/21/2004). Sylvia, D.M., A.K. Alagely, M.E. Kane, N.L. Philman. 2003. Compatible host/mycorrhizal fungus combinations for micropropagated sea oats. I. Field sampling and greenhouse evaluations. Mycorrhiza 13:177-183. Wu, G., Y. Feng, and S.A. Boyd. 2003. Characterization of bacteria capable of degrading soil-sorbed biphenyl. Bull. Environ. Contam. Toxicol. 71:768-775. Accepted Full Length Articles (Refereed Journals) Heinsch, F.A., J.L. Heilman, K.J. McInnes, D.R. Cobo, D.A. Zuberer and D.L. Roelke. 2003. Carbon dioxide exchange in a high marsh on the Texas Gulf Coast: effects of freshwater availability. Ag and Forest Meteor. In press. Keller, S.L, M.B. Jenkins, and W.C. Ghiorse. 2003. Simulating the effect of liquid CO2 on Cryptosporidium parvum oocysts in aquifer material. J. Environ. Engin. Accepted, 7 Nov. 2003. Rodgers, K., P. G. Hartel, R. L. Kuntz, and W. I. Segars. 2004. Presence of Enterococcus faecalis in human sewage, chicken litter, and wild birds. Water Res. (accepted). Stark, J.A., D.D. Bowman, M. Labare, E.A. Fogarty, A. Lucio-Forster, J. Barbi, M.B. Jenkins, M. Pavlo, and M.A. Butkus. 2003. Do iodine water purification tablets provide an effective barrier against Cryptosporidium parvum? Military Medicine. Accepted, 27 Oct. 2003. Submitted Full Length Articles (Refereed Journals) Hartel PG, S. Myoda, R. L. Kuntz, K. Rodgers, J. Entry, S. Ver Wey, E. Schröder, J. Calle, M. Lacourt, J. E. Thies, J. P. Reilly, and J. J. Fuhrmann. 2004. Geographic and temporal changes of Enterococcus faecalis ribotypes for bacterial source tracking. Submitted to J. Environ. Qual. Jones CM, and J. E. Thies. 2004. Laboratory evaluation of the entomopathogenic fungal species Metarhizium anisopliae and Beauvaria bassiana for use in controlling the alfalfa snout beetle, Otiorhynchus ligustici. Submitted to Biological Control. Meyers, S.K., S.P. Deng, N.T. Basta, W.W. Clarkson, and G. Wilber. 200_. Acetonitrile extractable and water leachable 2,4,6-Trinitrotoluene (TNT), Hexahydro-1,3,5-Trinitro-1,3,5-Triazine (RDX), and Octahydrol-1,3,5,7-Tetranitro-1,3,5,7-Tetrazocine (HMX) in soil and their impact on microbial community. (in review). Stromberger, M.E. Metal vs fire: a soil microbiology laboratory incubation study demonstrating microbial biomass and activities in disturbed forest soils. Submitted to J. Nat. Res. Life Sci. Ed. Vacca, D.J., M.C. Pellitteri, E. Padilla-Crespo, W.F. Bleam and W.J. Hickey. 2004. Isolation and molecular characterization of soil bacteria adapted to degrade humic acid-sorbed phenanthrene. Submitted to Appl. Environ. Microbiol. Yao S, I. A. Merwin, G. Bird, G. S.Abawi, and J. E. Thies. 2004. Orchard groundcover management practices that maintain soil cover stimulate soil microbial activity and alter microbial community composition. Submitted to Plant and Soil. In Print Abstracts: Ackerman, C. E., F. M. Wanjau, J. D. Wilbur, S. M. Brouder, R. W. Doerge, and C.H. Nakatsu. 2003. The effects of long term tillage and rotational treatments on the rhizosphere microbial community structure of soybean. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Baldwin, B., L. Nies, C. Nakatsu, and J. Simonds. 2003. Molecular genetic detection and enumeration of aromatic oxygenase genes at gasoline-contaminated sites. Abstr. Battelle Bioremed. Conf., Orlando, FL. Beasley, F. C., C. H. Nakatsu, N. Carmosini, A. Konopka. 2003. Genotypic and phenotypic characterization of Arthrobacter isolates from chromate contaminated soil. Annu. Meet. Abstr. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Carrero-Colon, M., C. Nakatsu, A. E. Konopka. 2003. Nutrient periodicity as a selective force in structuring microbial communities. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Devare, M, Londono, LM, Jones, CM and Thies, JE. 2003. Effects of rootworm-resistant transgenic corn and insecticide on soil microbial communities. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Entry, J. A., S. A. Ver Wey, P. G. Hartel, R. L. Kuntz, and K. Rodgers. 2003. Targeted sampling with Escherichia coli for bacterial source tracking in Idaho. Abstr. Annu. Meet. Am. Soc. Microbiol. p. 591. Feng, Y., D.W. Reeves, and P. Ji. 2003. Effect of crop rotation and tillage systems on soil microbial communities. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Feng, Y, P. G. Hartel, S. Deng, K. Rodgers, J. Fisher, and B. Liu. 2004. Survival of Enterococcus species and subspecies in sediment for bacterial source tracking. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C. Fisher, D.C., M.B. Jenkins, R.R. Lowrance, R.K. Hubbard, T.C., Strickland, G. Vellidis, and G.L. Newton. 2003. In vitro disappearances of E. coli and enterococci related to light, predation, and sedimentation. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Franzluebbers, A.J., M.B. Jenkins, D.A. Zuberer, N.S. Hill. 2003. Soil responses to tall fescue endophyte infection. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Hagedorn, C., A. H. Chapman, S. Herbein, M. Saluta, and P. McClellan. 2004. Microbial source tracking as a technology for identifying sources of fecal pollution in water. 4th National Conference on Science, Policy, and the Environment, National Council for Science and the Environment. Washington, D.C. Hagedorn, C. 2004. Bridging the gap between science and policy: Environmental Detection News. 4th National Conference on Science, Policy, and the Envrionment, National Council for Science and the Environment. Washington, D.C. Hahm, B.K., A. K.Bhunia, and C. H.Nakatsu. 2003. Application of AFLP for discriminating Escherichia coli isolated from livestock, wildlife and humans. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Hansom, J., M.-K. Lee, D.I. Doser, and Y. Feng. 2003. The hydrodynamics and related biogeochemical processes in the Permian Basin, western Texas. Geologic. Soc. Am. Annu. Meet. Abstr. 35(6):215. Hartel, P. G. 2003. Combining targeted sampling and Enterococcus faecalis isolation for bacterial source tracking of human fecal contamination. Abstr. Annu. Meet. Soc. Environ. Toxicol. Chem. p. 135-136. Hartel, P. G., S. Myoda, R. L. Kuntz, K. Rodgers, J. A. Entry, S. A. Ver Wey, E. C. Schröder, M. Lacourt, J. Calle, J. E. Thies, J. P. Reilly, and J. J. Fuhrmann. 2003. Geographic and temporal variability of Enterococcus faecalis ribotypes for bacterial source tracking. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C., p. 592. Hartel, P. G., K. Rodgers, S. N. J. Hemmings, J. L. McDonald, K. Gates, S. H. Jones, T. L. Bryant, B. O?Hara, E. Otero, and Y. Rivera?Torres. 2004. Desiccation as a mechanism of fecal enterococcal survival and regrowth for bacterial source tracking. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C. Hassall, A., and C. Hagedorn. 2004. Sources of fecal pollution in four mixed-use watersheds in Prince William County, Va. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Herbein, S., and C. Hagedorn. 2004. Fecal E. coli and Enterococcus isolated from two humans and three companion animals compared by pulsed-field gel electrophoresis. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Hickey, W.J., D.J. Vacca, M. Pellitterri and W.F. Bleam. 2003. Isolation and analysis of bacteria adapted to degrade humic-sorbed polynuclear aromatic hydrocarbons (PAH). Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Hristova, K. R., B. Inceoglu, C. Naktsu. K. M. Scow. 2003. Characterization of enzymes involved in MTBE biodegradation in strain PM1. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Hutchinson, J. L., K. S. McLean, Y. Feng, C. Burmester, and G. W. Lawrence. 2003. Accelerated breakdown of aldicarb in Alabama cotton field soils. Phytopath. 93:S38. Publication no. P-2003-0274-AMA. Jenkins, M.B. and A.J. Franzluebbers. 2003. Soil microbial community structure under endophyte-free and indophyte-infected tall fescue as determined by FISH analysis. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Jerke, K., C. H. Nakatsu, and A. Konopka. 2003. A physiological and genetic approach to studying lead resistance in Arthrobacter sp. VN23-1. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C. Jimenez-Esquilin, A.E., and M. E. Stromberger. 2003. Recovery of soil microorganisms following the Hayman wildfire. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Jimenez-Esquilin, A., and M. Stromberger. 2004. Structural characterization of the microbial community in a fire-affected forest soil. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C. Kerns, J.P.K., D.M. Vietor, R.H. White T.L. Provin and D.A. Zuberer. 2003. Physical and biological properties of soil amended with composted dairy manure during turf establishment. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Kourtev, P. S., N. Carmosini, F. Beasley, C. Nakatsu, A. E. Konopka. 2003. Heavy metals modulate microbial community responses to inputs of organic carbon. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C. Kuntz, R. L., P. G. Hartel, D. G. Godfrey, J. L. McDonald, K. W. Gates, and W. I. Segars. 2003. Targeted sampling protocol with Enterococcus faecalis for bacterial source tracking. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C., p. 573-574. McKinney, J., and C. Hagedorn. 2004. Identifying sources of fecal pollution in the Appotomattox River Watershed. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington D.C. Provin, T.L., R.H. White, D.A. Zuberer, J.C. Thomas and J.L. Pitt. 2003. Influence of compost amendments on sod establishment and soil physical, chemical and microbiological properties. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Reilly, JP, Thies, JE and Baeumner, A. 2003. Development of an optical strip-assay biosensor for Enterococcus faecalis. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, DC. Rodgers, K., P. G. Hartel, R. L. Kuntz, and W. I. Segars. 2003. Presence of Enterococcus faecalis in human sewage, chicken litter, and wild birds. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C., p. 592. Sangsupan, H.A., D.E. Radcliffe, M.B. Jenkins, P.G. Hartel, W.K. Vencill, and M.L. Cabrera. 2003. Transport characteristics of 17-beta estradiol and testosterone in no-till and conventionally tilled Georgia Piedmont soil. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Savin, M.C., S.E. Ziegler, G.J. Thoma, K.M. Greer , P.J. Tomlinson, and D.C. Wolf. 2003. Nematode population changes during phytoremediation of a crude oil-contaminated soil. p. 44. In 10th Annu. Internation. Petroleum Environ. Confer. Integrated Petroleum Environ. Consort., Tulsa, OK. Schröder, E. C., C. I. Castro, P. G. Hartel, R. L. Kuntz, and K. Rodgers. 2003. Targeted sampling with Escherichia coli for bacterial source tracking in Puerto Rico. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, D.C., p. 592. Sullivan, T.S, and M.E. Stromberger. 2003. Microbial community responses to biosolids in rangeland soils. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Tomlinson, P. J., K. R. Payne, K. R. Brye, and M. C. Savin. 2003. Microbial dynamics in long- term research plots receiving alum-treated and untreated poultry litter. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Wu, W, JE Thies, Q Ye, and H Min. 2003. Effect of Bt transgenic rice straw on the bacterial community diversity in a flooded soil. Annu. Meet. Abst. [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI. Wu, W, Q Ye, H Min, and JE Thies. 2003. Microbial enzyme activities and bacterial community composition in the rhizosphere of Bt-transgenic rice as compared to its non-transgenic cultivar. Abstr. Annu. Meet. Am. Soc. Microbiol., Washington, DC. Zhang, L. and Y. Feng. 2003. Sorption and desorption of carbaryl by soils. Annu. Meet. Abst. Southern Branch ASA [CD-ROM], Madison, WI. Zurdo-Piñeiro, J. L., Lorite, M. J., Brelles-Mariño, G, Schröder, E. C., Bedmar, E.J., Martinez-Molina, E., Mateos, P.F. and Velázquez, E. 2002. Identificación de rhizobia de crecimiento rápido que nodulan leguminosas en Puerto Rico. XXI Reunión Latinoamericana de Rhizobiología, Cocoyoc, México. p. 89. Accepted Abstracts Devare, M.H., L.M. Londono, and J. E. Thies. 2004. A comparative assessment of cry3Bb Bt corn and insecticide on microbial biomass, processes, and bacterial diversity in soil. ASA, Seattle, WA Jones, C.M. and J.E. Thies. 2004. Microbial community analysis in biocontrol research: Creation of a regional sequence database and refinement of T-RFLP for use with fungal populations. International Society for Microbial Ecology Meeting, Cancun, Mexico. Jones, C.M. and J.E. Thies. 2004. Soil bacterial and fungal community response to application of entomopathogenic fungi in the field. ASA, Seattle, WA Kantety, R., L.M. Londono, M. B. McBride, and J. E. Thies. 2004. Changes in microbial community structure in long-term sludge amended soils and naturally metalliferous soils. ASA, Seattle, WA Londoño, L.M., M. H. Devare, and J. E. Thies. 2004. Effects of Cry3Bb Bt corn and insecticide on diversity of fungi in soil. ASA, Seattle, WA Rumberger, A, J. E. Thies, E. B. Nelson, and I. A. Merwin. 2004. Apple replant disease and bacterial rhizosphere community composition. ASA, Seattle, WA Thies, JE, W. Wu, and M. H. Devare. 2004. Multi-year field trials validate transgenic Bt corn and Bt rice pose little risk to the osil microbial foodweb. International Society for Microbial Ecology Meeting, Cancun, Mexico. M.S. Theses McKinney, J. M. Identifying Sources of Fecal Pollution in the Appomattox River Watershed. M.S. Thesis, May, 2004. Porter, K. R., Identifying Sources of Fecal Pollution in Washington D.C. Waterways. M.S. Thesis, December, 2003. Non-Refereed Publications Hartel, P. G., E. A. Frick, A. L. Funk, J. L. Hill, J. D. Summer, and M. B. Gregory. 2004. Sharing of ribotype patterns of Escherichia coli isolates during baseflow and stormflow conditions. U. S. Geological Survey, Scientific Investigations Report 2004?5004. 10 p. Proceedings Chellemi, D. O., D.J. Mitchell, M.E. Kannwischer-Mitchell, E. Rosskopf and J.Graham. 2003 Soilborne disease and root colonizing fungi in vegetable crop systems, p. 334 in Proc. 8th Intern. Cong. Plant Pathology, Christchurch, New Zealand. Graham, J. H. M. S. Irey, J. B. Morton, L. Baucum, and D. Bright. 2003. Sugarcane yield decline and its association with early colonization rate by arbuscular mycorrhizal fungi (AMF). p. 265 in 4th Intern. Conf. on Mycorrhizas, Montreal, Canada. Graham, J. H., S. Ravnskov, J. Larsen. 2004. Testing models for mycorrhiza Pythium interactions in tomato roots. p. 37 in Proc. Development of Biocontrol Agents of Diseases for Commercial Applications in Food Production Systems. Sevilla, Spain. Hons, F.M., A.L. Wright, S.M. Kolodziej, V. A. Saladino, R.L. Lemon, M.L. McFarland and D.A. Zuberer. 2004. Rotation, tillage and nitrogen rate effects on cotton growth and yield. Proc. Of the 2004 Beltwide Cotton Conference, San Antonio, TX. Kolodziej, S.M., F.M Hons, Alan Wright and D.A, Zuberer. 2004. The effects of tillage and rotation on labile organic carbon and aggregation in a cotton cropping system. Soil Management and Plant Nutrition. Proc. Beltwide Cotton Conference, San Antonio, TX. Rodgers, K., P. G. Hartel, R. L. Kuntz, D. G. Godfrey, and W. I. Segars. 2003. Field-testing Enterococcus faecalis for bacterial source tracking. p. 837-841. In K. J. Hatcher (ed.) Proceedings of the 2003 Georgia Water Resources Conference, April 23-24, University of Georgia, Athens. Presentations Padilla-Crespo, E., D. Vacca and W. J. Hickey. 2003. Diversity of Burkholderia polycyclic aromatic hydrocarbons (PAH) degradation genes. NIEHS Superfund Sci. Mtg, Los Angeles, CA; 13-15 October 2003. Reilly, J.P., and Thies, J.E. 2003. Preliminary findings from ribotyping of enterococci isolated from Wappinger and Stony Creek catchments. Invited talk at the Dutchess Environment Council meeting: Emerging Water Contaminants. Sullivan, T., M. Stromberger, and R. Brobst. 2004. Microbial community responses to biosolids in rangeland soils. Sustainable Land Application Conference, Lake Buena Vista, FL, January 4-8. Torres-Albelo, J.J. and Schröder, Eduardo C. 2003. Arboles Fijadores de Nitrógeno. Pithecellobium arboreum (Subfalia Mimosoidea). Grupo Caribeño de Rhizobium, March, 2003. Book Chapters Hartel, P.G. 2004. Environmental factors affecting microbial activity. In D. Hillel, C. Rosenzweig, D. S. Powlson, K. M. Scow, M. J. Singer, D. L. Sparks, and J. Hatfield (ed.). Encyclopedia of soils in the environment. Elsevier, London (accepted). Hickey, W.J. 2004. Microbiology and biochemistry of xenobiotic compound degradation. In D.M. Sylvia et al. (eds.) Soil Microbiology: Environmental and Agricultural Perspectives, 2nd Ed. Oxford University Press, New York, NY. (In Press). Keeling, W. G., C. Hagedorn, and B. A. Wiggins. Concepts and applications of bacterial source tracking in water quality. In T. Younos, (ed.). TMDL: Approaches and Challenges, PennWell Press. Expected date of publication is June 2004. Stromberger, M.E. Fungal communities of agroecosystems. In J. Dighton et al. (ed.) The Fungal Community. Marcel Dekker, New York, NY (submitted). Books Sylvia, D. M., J. J. Fuhrmann, P. G. Hartel, and D. A. Zuberer (eds). 2004. Principles and applications of soil microbiology, 2nd edition. Prentice-Hall, Upper Saddle River, NJ. (accepted).

Impact Statements

NY BST efforts have brought to light that failing wastewater treatment facilities in the region studied are likely contributing to elevated levels of total coliforms and enterococci in the watershed.
Results from River Yagüez are almost completed, and there is indication that significant variation in microbial populations exists. These indicate that factors are affecting microbial biomass.
Results of the BST studies confirm that ribotyping of Enterococcus faecalis is a useful and highly discriminatory approach to BST, but that it must be combined with a targeted sampling protocol in order to overcome the high geographic and temporal variability in ribotypes.
Although differences in survival exist among fecal enterococcal species and subspecies, and differences exist between locations, none of the Enterococcus species or subspecies survived sufficiently well in moist sediment to serve as a reservoir of long-past human and nonhuman fecal contamination. Therefore, the American Public Health Association rule that fecal indicators not persist in the environment held.
However, results from desiccation experiments suggest that enterococcal survival and potential regrowth during conditions of desiccation may represent a source of relatively long-past human or nonhuman fecal contamination. Under these circumstances, the American Public Health Association rule that fecal indicators not persist in the environment fails. The implications of these results for testing fecal contamination of water are troubling.
The watershed project in Virginia will provide a real-world evaluation of three source tracking methods and two indicator organisms over time, and offers the opportunity to compare the size and origin of sediment populations as well as those in water samples.
The development of a method to detect optical brighteners will permit detection of fluorescent plumes from failing onsite systems or leaking sewer pipes, and should be adaptable as a instantaneous presence-absence test for human-derived pollution in recreational waters such as beaches.
The most important finding in our comparison of genotypic methods to determine a source of fecal contamination in the environment is the need to use the method most appropriate for the question being asked and is dependent on cost and level of specificity required.
The development of a species-specific primer that works in a biosensor to detect synthetic sequences of Enterococcus faecalis is a positive first step in the development of an inexpensive new technique to detect pathogens from the environment.
Results of the studies under objective 2 can guide development of bioremediation strategies that maximize the use of natural resources while minimize environmental risk.
We believe that this is the first report of a specific group of bacteria adapting to interact with humic acids in such a way that PAH sorbed by these environmental polymers is bioavailable.
These organisms may have value as bioremediation inoculants, and have an important role in the biodegradation processes affecting the fate of PAH in the environment.
Enumeration of aromatic oxygenase genes by real-time PCR combined with chemical analysis required in corrective action plans could be used to directly assess biodegradation of priority pollutants which can help provide more accurate assessment of biodegradation, facilitate assessment of the impact of remediation technologies on indigenous microbial populations, and enhance studies of contaminated site ecology.
Data from the phytoremediation experiment will help understand how establishment of grasses and management practices affect microbial dynamics over the long-term and thus allow us to gather insight and interpret the process and extent of ecosystem recovery during remediation.
Biodegradative strains isolated from RDX/TNT contaminated soils have been obtained, which may be used for environmental research and application.
Data can be used to provide guidelines for calculating nutrient amendments for bioremediation of nitroaromatic-contaminated soils.
Results improved knowledge of pollutant bioavailability and impact of nitroaromatic-contamination on indigenious microbial population and water quality.
The level of resistance measured in bacteria isolates appears to be a function of the antibiotic tested, management treatment, and time since application, but further research examining additional antibiotics and manure types is needed.
Preliminary results examining antibiotic resistance of bacteria in soils receiving animal wastes indicate that despite low rates of waste application, differences in percent resistance could be detected in bacterial isolates from soils where waste was applied compared to those that had never received waste.
Research results will contribute to developing a comprehensive means to identify soil biota that influence crop productivity under different management conditions which is critical to the long-range improvement of US agriculture because it addresses the contribution of the immense diversity of biota to both sustaining agricultural production and maintaining or improving soil and environmental quality.
We have established a comprehensive seasonal database describing the abundance of bacteria and fungi in highly managed turfgrass systems from sports fields with sand-based root zones to turfs managed as home lawns which provides a strong foundation upon which turf managers can make informed decisions regarding purchases of inputs for golf courses, sports fields, etc. All too often they are besieged with vendors selling dubious products with extravagant claims for turf management.
Microbiological data from the compost study are of value, along with the data describing the physical and chemical properties of the soils, in the decision-making process involving the disposition of large volumes of compost materials derived from the composting of dairy manure in North Central Texas.
Data from the cotton-management study indicate that there is a potential for increased carbon storage and less nitrogen is needed in soils managed in rotation and with reduced-tillage for cotton production
The consequence of AMF suppression in conventional vegetable production systems due to routine applications of biocides in terms of assessment of root health in pathogen-dominated vegetable soils in unknown. Preliminary indications are that low AMF incidence does not render roots more susceptible to pathogens, but may reduce plant tolerance to damage.
e conclude that AMF have an important role in the accumulation of As by Brake fern in As-contaminated soils.

Date of Annual Report: 10/03/2005

Report Information

Annual Meeting Dates: 06/11/2001 - 06/27/2005
Period the Report Covers: 10/01/2000 - 09/01/2005

Participants

Brief Summary of Minutes

Minutes Attachment:

Attachment

Accomplishments

Objective 1: to determine the geographic variability of E. coli ribotypes in the United States. We conducted collaborative experiments to determine the geographic variability of E. coli ribotypes in the United States. With the help of researchers from AL, DE, GA, IN, NY, PR, TN, TX, and USDA over the 5-year project, the objective was accomplished. We determined that there was good ribotype separation among host animal species at one location, and that the ability to match environmental isolates to a host origin database depended on a large number of environmental and host origin isolates that were not geographically separated. We extended the changes of ribotypes of E. coli with geography to time. Our results suggested that the majority of ribotypes were transient and unique to each sampling time; therefore, a large number of E. coli isolates per host were necessary to establish a host origin database that was independent of changes with time. We also extended this work to flow conditions, and established that ribotype changed with flow conditions. Finally, we established that ribotypes changed with an animals diet. All these results were discouraging because if ribotypes of E. coli changed with geography, time, flow rate, and diet, then a permanent host origin database large to encompass these patterns would be time-consuming and expensive to construct. Rather than continue with E. coli, we decided to test another species of fecal bacteria, Enterococcus faecalis, for changes in ribotype patterns with geography. We completed another collaborative manuscript this bacterium and the results were similar to that of E. coli. Again, these results are discouraging because a permanent host origin database large to encompass these patterns would be time-consuming and expensive to construct. Therefore, with two exceptions, we decided to move from library-dependent bacterial source tracking (BST) methods to library-independent methods. The first exception was a multi-year project to investigate at-risk or impaired watersheds in Virginia with two library-dependent BST methods, antibiotic resistance analysis and pulsed-field gel electrophoresis. The watersheds ranged from completely urban or suburban basins (all homes in the watershed served by sewers) to mixed-use basins dominated by agricultural activities (most homes served by onsite systems) to forested basins within the boundaries of a state park (no homes). Water and sediment samples were collected and analyzed for E. coli and enterococci. The goal of the project was to use the two library-dependent methods to identify some fecal sources that were apparent (should be present) and some sources that were not (should not be present, e.g., livestock sources should not be present in the completely suburban watershed). Based on the locations of the sample sites and their localized sources of contamination, two libraries, comprised of known source isolates collected from the county and the greater northern Virginia area, were used to analyze the patterns of isolates recovered from membrane-filtered stream water samples. The average rate of correct classification for the 1,490-isolate, four-way (human, domestic pets, wildlife, birds) urban library was 80.5%. The average rate of correct classification for the 2,012-isolate, four-way (human, livestock, wildlife, birds) rural library was 84.1%. To date, the major sources in the rural watersheds have been livestock and wildlife (with a minor human signature at certain locations), and the major sources in the urban areas have been birds and wildlife. After storm events, the livestock signatures were elevated in rural areas and wildlife signatures were elevated in urban areas. The second exception was to compare various library-dependent, genotypic BST methods for their strengths and weaknesses. All the methods correctly identified the dominant host source in the majority of samples, but false positive rates were as high as 57%. One S-297 researcher continued this research with another research team examining both genotypic and phenotypic methods. Their results were similar. Four projects with library-independent methods were conducted. The first project was to make BST quicker, more effective, and less expensive, and led to the development of targeted sampling as a prelude to BST. Targeted sampling is like the childrens game of hot and cold. The sampling is first divided into baseflow and stormflow conditions because runoff during stormflow typically increases fecal bacteria counts 10- to 100-fold. By combining local knowledge, sampling, and resampling of the water during one of these two flow conditions, it was possible to identify hotspots of fecal contamination quickly and easily. Targeted sampling also reduced the effects of bacterial changes with geography and time, and this reduction made existing BST methods better and less expensive by reducing the environmental complexity. At this point, targeted sampling has been conducted during both base- and storm-flow conditions in marine waters along the Georgia Coast, but has not yet been conducted in freshwater or in other states. The second project was to determine the limitations of fluorometry as a BST method. Fluorometry works by detecting optical brighteners from laundry detergents in environmental waters because these compounds fluoresce when exposed to UV light. Therefore, fluorometry is not a bacterial source tracking method per se, but a chemical source tracking method. Furthermore, because these fluorescent compounds are associated with human sewage, the method discriminates only between human and nonhuman sources of fecal contamination. Preliminary tests by researchers from GA and VA concluded that fluorometry was an acceptable, inexpensive method to detect human sewage in fresh and marine waters. The third project was to pursue enterococcal speciation as a simple method to discriminate between human and non-human fecal contamination. We determined that Ent. faecalis had a host range essentially restricted to humans and wild birds. Our results suggest that unless the fecal loading rate from migratory or resident wild birds is high, water samples collected during baseflow conditions with high numbers of Ent. faecalis may indicate human fecal contamination. The fourth project was a proof-in-concept project for a species-specific ribosomal DNA gene biosensor to detect Enterococcus faecalis rapidly in environmental samples (water and human feces). The biosensor used sulforhodamine B (SRB) dye-encapsulated liposome technology in a competitive assay to detect the presence of a synthetic 16S rRNA gene sequence specific to Ent. faecalis. Two different target analyte sequences were hybridized with liposomes that had reporter probes (ssDNA oligonucleotides) attached to them. The bioassay strip was sensitive when used with synthetic target sequences and could be run in under 20 minutes, but was not able to detect the target sequence amplified from environmental isolates or ATCC strains of the Ent. faecalis 16S rRNA gene. The positive signal generated with the control synthetic sequence suggests that perhaps secondary structures (e.g., hairpins) may inhibit the hybridization of the target analyte with the capture and reporter probes. Objective 2: To determine relationships among microbial taxonomic and functional diversity, contaminant bioavailability, and remediation rates for different organic-contaminated soils. A 5-year collaborative project was conducted to evaluate rhizosphere-enhanced bioremediation of organic contaminants. The participants included researchers from AL, AR, DE, FL, IN, NH, NC, OK, SC, WI, and Canada. We first evaluated the interactions of different plant species, soils, and nutrients to remediate soils contaminated with such organic contaminants as crude oil, nitroaromatic explosives, hexadecane, and pyrene. We began our evaluations by testing 21 warm- and cool-season grasses and legumes for their ability to germinate, survive, and grow in crude oil-contaminated soil. The most appropriate warm-season grasses were pearl millet (Pennisetum glaucum) and sudangrass (Sorghum sudanense), and the most appropriate cool-season grasses were ryegrass (Lolium multiflorum) and fescue (Festuca arundinacea). In soil amended with pyrene and sprigged with bermudagrass (Cynodon dactylon), the number of pyrene-degrading microbes was significantly higher than in soil not amended with pyrene. These results suggest that the presence of the contaminant and the plant increases the potential of the rhizosphere microbial community to degrade pyrene. In soil amended with pyrene containing no plants, whole soil fatty acid methyl ester (FAME) analysis indicated a shift in the composition of the soil microbial community in the pyrene-amended soil compared to the unamended soil. We then evaluated the influence of rhizosphere on soil remediation of spilled crude oil at a petroleum storage tank facility. The treatments were: 1) non-vegetated non-fertilized control, 2) fescue-ryegrass mixture plus fertilizer, and 3) bermudagrass-fescue mixture plus fertilizer. Vegetation was successfully established at the site, even though the soil had an initial total petroleum hydrocarbon concentration of 9,175 mg/kg. While alkylated two-ring naphthalenes degraded equally in all treatments, the larger three-ring alkylated phenanthrenes-anthracenes and dibenzothiophenes degraded more in the vegetated, fertilized plots than in the non-vegetated, non-fertilized plots. In this field study, the increased biodegradation of the more recalcitrant alkylated polycyclic aromatic hydrocarbon compounds in the crude oil-contaminated soil most likely occurred because of the increased rhizosphere soil volume (associated with increased root length) and because the added nutrients increased total bacterial, fungal, and polycyclic aromatic hydrocarbon (PAH) degrader numbers. In a separate study, pre-established bins of soil (0.9-m diameter) were amended with pyrene in a 10-month experiment under field conditions. After a 175-d lag period, the rate of pyrene loss followed first-order kinetics, with a rate constant significantly higher in nonvegetated than vegetated treatments. The delay of pyrene dissipation in the rhizosphere may be because of the presence of easily degradable organic material from the plant roots. The successful bioremediation of sites contaminated with aromatic compounds typically relies on the presence and stimulation of aromatic hydrocarbon-degrading bacteria from the indigenous microbial population. Although aerobic bioremediation is the common treatment, there is no direct biological assay to document the bioremediation during treatment. We developed a technique based on real time PCR amplification of aromatic oxygenase genes to detect and quantify a number of aromatic catabolic genes in environmental samples. Each primer set was specific for a family of oxygenase genes (e.g., toluene dioxygenase). Based on available published sequences, PCR primer sets were identified which targeted biphenyl dioxygenase, naphthalene dioxygenase, toluene dioxygenase, toluene/xylene monooxygenase, phenol monooxygenase, and ring hydroxylating-toluene monooxygenase genes. The primer sets and real-time PCR methods were then used to demonstrate the effectiveness of the approach, first in laboratory enrichment microcosms, and then at contaminated field sites. At the field sites, aromatic oxygenase genes were detected in groundwater monitoring wells with current or recent petroleum contamination, but not in wells with no history of contamination. Genes were no longer detected after contaminant concentrations reached zero. We determined that certain bacteria were uniquely adapted to interact with humic acids in a manner that allowed these organisms to access and degrade PAHs sorbed by humic acids. Although we isolated a wide variety of bacteria that degraded non-sorbed phenanthrene, only three strains of Burkholderia sp. and one strain of Delftia acidovorans degraded humic acid-sorbed phenanthene. We termed this phenotype characteristic competence. Competence was not a graded characteristic; bacteria either degraded humic acid-sorbed phenanthene or they did not. The characteristics of competent cells that support this phenotype, and the nature of the interaction occurring between humics and bacterial cells, are unknown. However, competence determinants were physically associated with the cells; diffusible agents (e.g., biosurfactants) did not have a significant role. As yet, the known physical differences between competent and closely related non-competent strains are phospholipid fatty acid content and extracellular matrix production. To the best of our knowledge, this is the first report of specific groups of bacteria adapting to interact with humic acids. We evaluated the microbial activity and community structure in soils contaminated with the nitroaromatic explosives 2,4,6-trinitrotoluene (TNT), hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), and octahydrol-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX). Our results suggest that natural attenuation of these compounds in highly contaminated soils may not be feasible. Because nonionic surfactants had little effect on bacterial growth, these surfactants may help overcome this natural attenuation. We studied the characteristics of isolates from two bacterial species, Pseudomonas putida and Rhodococcus erythropolis, to degrade soil-sorbed biphenyls. Both isolates showed strong tendencies to attach to soils. The pseudomonad was more motile and had a higher chemotactic response to biphenyl than the Rhodococcus isolate. It appears that attachment to soil, motility, and chemotaxis are important characteristics that influence the bacterial access to soil-sorbed biphenyl. Bacteria with higher chemotactic response and moderate cell surface hydrophobicity may access soil-sorbed biphenyl more efficiently than bacteria with a lower chemotactic response and less cell surface hydrophobicity. We determined the influence of soil properties on the degradation of 14C- and 13C-labelled glucose or hexadecane. Even though the parent compound was rapidly degraded (half-life <2 d), only 34% of the C was evolved as 14CO2 after 275 d, with the remaining C incorporated into microbial biomass and soil organic matter. Studies with 13C-labelled glucose show that the initial microbes that metabolized the C underwent degradation themselves, the C of which was then incorporated into phospholipid fatty acids of other members of the microbial community. There was sufficient 13C incorporated into phospholipid fatty acids that it was possible to assess what members were active in C degradation. Objective 3. To characterize taxonomic and functional diversity of bacteria and mycorrhizae in disturbed lands and urban landscapes. We conducted collaborative experiments to evaluate the effects of various disturbances or land-management systems on the microbial abundance and community structure and function of soil communities. Over the 5-year project, participants included researchers from AL, GA, IN, NC, NY, OR, SC, and TX. We divided land use into four management systems: pasture, cropping, turfgrass, and mycorrhizae. In pasture systems, we conducted both short-term growth studies and long-term field experiments with scientists from GA, TX, and USDA to determine changes in soil organic C and N during growth of tall fescue uninfected or infected with the fungus, Neotyphodium. We thought that plant metabolites produced in association with the endophytic fungus could alter soil microbial activity and how it processed C and N in soil. Tall fescue forage accumulation was greater with than without the fungus, confirming previous results that fungus infection also confers greater fitness to plants. Microbial communities under endophyte-infested and non-infested fescue were different in terms of C source utilization (BIOLOG); this difference suggested that the endophyte affected the metabolic capacities of the microbial populations in the soil. In cropping systems, we conducted three studies on the effects of continuous cotton management systems on soil biological properties. In the first study, phospholipid profiles suggested that microbial communities associated with conventional tillage and no tillage continuous cotton systems were dissimilar, and that the tillage effect varied with soil depth and time. The second study compared a standard cotton production system with an intensive cotton cropping system that maximized the production of crop residues and legume N inputs. Again, the phospholipid profiles showed that soil microbial communities under conventional tillage were different from those under the conservation tillage treatment, and that soil microbial communities shifted with soil depth. In the third study, we investigated several soil microbiological parameters in a cotton crop-management system: soil organic C, microbial biomass C, aggregate size classes, and water-extractable carbon (80°C for 16 h), and total glomalin in soils from plots under different tillage, fertility (0 and 90 kg N ha-1), and rotation schemes (conventional till continuous cotton, reduced-till cotton-corn rotation, and reduced-till continuous cotton). Soil from the reduced-till and cotton-corn rotation treatments contained the more organic matter compared with continuous cotton plots. No major differences were found in aggregate size classes among the treatments, except that the >2000-micrometer fraction increased in the cotton-corn rotation at the expense of the 53- to 250-micrometer fraction in the plots receiving 90 kg N ha-1. Soil microbial biomass carbon ranged from approximately 270 to 550 mg kg-1 across all treatments, including N application and depth. Hot-water extractable C ranged from approximately 261 to 445 mg kg-1 in all treatments including N and depth. Total glomalin ranged from approximately 2300 to 2800 mg kg-1 soil, and was well correlated with soil organic C (r2=0.87). Total glomalin C accounted for roughly 7 to 9% of the soil organic C. In a 3-year cropping system with plants other than cotton, we determined the effects of different agricultural management practices on soil and rhizosphere microbial communities of corn (Zea mays) and soybean (Glycine max). Corn and soybean plants grown under different tillage (no-till or plow) and rotation (monoculture or rotation) treatments were collected over different growth stages (V1, V2, V3, V6, V9, and maturity) during three growing seasons. When rhizospheres of corn and soybean plants were compared using PCR amplification of the small subunit rRNA gene and separated by denaturing gradient gel electrophoresis, the banding patterns of the microbial rhizosphere community grouped mainly according to agronomic treatment. This effect was more evident in the bacterial versus the eukaryotic communities, and in corn versus soybean rhizospheres. We identified the most significant bands in the corn rhizosphere that were consistently associated with improved plant growth for all three seasons. Nucleotide sequencing identified one of the four bands as Burkholderia species. We chose this species of bacteria for subsequent laboratory plant growth experiments and found significant differences in plant growth when the plants were inoculated with these isolates. We used a polyphasic approach (combining traditional and molecular methods) to assess changing soil and crop management strategies (or naturally occurring soil stresses) on soil microbial community abundance, activity, and diversity in the rhizosphere of transgenic (Bt) corn and rice. After 3 years of field trials, the transgenic plants had no adverse effects on the abundance, activity, or diversity of microbial communities compared with their non-transgenic isolines. Residues of two transgenic Bt corn varieties decomposed at the same rate as their non-transgenic counterparts and were colonized by similar microbial communities. Residues placed on the soil surface decomposed significantly slower than those buried at 5-cm depth, and were colonized by distinctly different bacterial and fungal communities. In the case of corn plant parts, cobs, stalks and leaves were colonized by distinctly different bacterial and fungal communities and decomposed at different rates, ranging from low to high, respectively. However, transgenic versus non-transgenic plants were not a significant factor. We conducted several long-term experiments with wheat to determine changes in bacterial populations with different tillage systems. Bacterial populations varied during the year, exhibiting a general decrease in the late fall and an increase in the early spring. Bacterial numbers increased in a no-till, summer-fallow system when compared to conventional tillage--contrary to some reports that suggest that microbial populations will decrease in these management systems. With respect to carbon dioxide flux, the flux in conventionally tilled fields was almost five times greater than no-till fields. After determining C and N mineralization rates of crop residue components, we developed a C sequestration model, CQESTR, to estimate residue loss and C sequestration, but the model was not sufficiently user friendly. Therefore, we developed a more user-friendly model by rewriting the user manual, adding a program tutorial, and inserting a Help function in the Microsoft Windows interface. In turfgrass systems, scientists from AL, NC, SC, and TX investigated various aspects of the microbiology of intensively managed turfgrass systems including golf courses, sports fields, and home lawns. Turfgrass systems differ from agricultural systems because, except for aeration, they are essentially zero-tillage systems maintained with plenty of water and nutrients. Therefore, they really represent highly productive, managed grasslands. We showed that sports turfs contained abundant microbial populations (bacteria and fungi) throughout the growing seasons. Further there was relatively little seasonal decline, and populations were relatively stable over periods of several years when turf maintenance practices were adequate. Furthermore, in terms of species composition, the populations were similar to those found in other agricultural systems. In mycorrhizal systems, four major studies were conducted. In the first study, a 5-year experiment was initiated to measure how alternative land management practices for fresh market tomato production affect soil health. The prevailing system uses raised beds and fumigation with methyl bromide for control of soil-borne nematodes, weeds, and root fungal pathogens. We tried five different management programs in replicated 1-acre plots over a 3-year period prior to a 2-year resumption of the prevailing system: 1) conventional tomato production using nematicide/fungicide and herbicides; 2) continuously maintained disk fallow, 3) weed fallow in which endemic plants colonized the plots; 4) bahiagrass conservation tillage where strips were tilled in the grass sod before tomato plantation, and 5) organic management consisting of cover crops of millet and Sunn hemp (a tropical legume) rotations and annual broadcast of broiler chicken litter mixed with urban plant debris. We monitored changes in soil fungal communities with length heterogeneity PCR and non-metric multivariate analysis. After three years, fungal communities in soil under continuous tomato production and continuous disk-fallow cultivation were similar. Communities in soil that were left undisturbed in a weed fallow system were similar to communities in a perennial pasture grass rotation. Communities within an organically managed system were unique. Following traditional tomato production in the fourth year, communities in the organic and pasture grass systems remained unique, whereas communities in the weed fallow were similar to communities under continuous tomato production or continuous cultivation. Fungal communities were dominated by a 341-bp rDNA amplicon fragment. Cloning and sequencing indicated the dominant fragment belong to Fusarium spp., which was further confirmed by the ITS 1 sequence from single spore isolates of F. oxysporum f. sp. lycopersici. The relative abundance of the 341-bp fragment was greatly decreased in organic and pasture grass systems that which also had a lower incidence of Fusarium wilt in the tomato crop. At the end of each crop season, tomato rhizosphere soil and fibrous roots of tomato and cover crops were collected. Roots were cleared, stained and examined for arbuscular mycorrhizae (AM) and other root endophyte colonization. Rhizosphere soil was assayed for mycorrhizal infection potential (MIP) based on maize seedling colonization after 35 days. Bahiagrass and weed fallow systems promoted the highest AM colonization in the tomato crop and produced moderate MIP levels and spore numbers in rhizosphere soils. Cover crops had moderate levels of AM colonization. The disk fallow, conventional and organic systems supported lower AM colonization of tomato roots, but systems that created the most disturbance (conventional and disk fallow), promoted the highest MIP and spore counts from trap cultures. Thus, managements potentially most disruptive to AM fungal communities produced the highest inoculum density, but the least infective inoculum with respect to the tomato crop grown under nutrient supply. Organic management with the highest P supply was least conducive for development of MIP or AM colonization of tomato. In contrast, the organic system supported one non-AM endophyte at high incidence (Microdochium bolleyi). In the other systems, a diversity of non-AM endophytes was observed, including a dark septate endophyte (Phialophora spp.). Considering the companion analysis of the bacterial and fungal diversity, communities were more similar and diverse in the systems that produced the least disturbance (bahiagrass sod and weed fallow). Thus, soil microbial activity and diversity in the agro-ecosystems studied were most affected by soil nutrient supply and disturbance. In the second major mycorrhizal study, we determined if sugarcane yield decline was linked with early and rapid colonization by AM or not. Fallow management (up to 1 year) of sugarcane soils followed by repeated tillage (to break up the root crown and to reduce weed cover before replanting) produced up to a 30% increase in biomass at the first cutting, but only a slight increase at the second cutting, and no increase at the third cutting. Treatments with soil biocides duplicated the fallowing effect. In most instances, no soil microorganisms deleterious to sugarcane roots were identified. Arbuscular mycorrhizae were trapped from successively planted sugarcane fields, and three Glomus isolates were selected to reconstitute a steamed local muck in glasshouse experiments. Muck treated or not treated with steam received a combined extract of the non-steamed field soil and pot cultures of each isolate to check for the deleterious effect of microorganisms that pass through 20-micrometer sieve openings. Roots in non-steamed soil were rapidly colonized in advance of shoot development. In reconstituted soils, rate of mycorrhizal development varied with the Glomus isolate. Shoot biomass gain was lowest in non-steamed soil, and highest in steamed soil, with no effect of the extract from the non-sterile field/pot culture soils. Total biomass gain was inversely related to colonization rate among soil and Glomus isolates. Reduction of biomass gain compared to the steamed soil treatment was best predicted by early root colonization for the three Glomus spp. and soil treatments at 2 weeks after shoot emergence, and to a lesser extent by later colonization at 4 and 6 weeks. In the third study, we evaluated arsenic uptake in the hyperaccumulating fern, Pteris vittata (Chinese brake fern), a fern that grows naturally in soils of the southern United States. A greenhouse experiment was conducted with and without P. vittata, where arbuscular mycorrhizae from an As-contaminated soil colonized the plants. Three levels of arsenic (0, 50, and 100 mg kg-1) and three levels of P (0, 25, and 50 mg kg-1) were used. Arbuscular mycorrhizae not only tolerated As amendment, but also their presence increased frond dry mass at the highest As application rate. The AM fungi increased As uptake across the range of P levels, while P uptake generally increased only when there was no As amendment. In the fourth study, we studied fungal and bacterial dynamics in relatively nutrient-rich and nutrient-poor wetland plant communities. The dominant wetland plant communities (Panicum, Cladium, Typha, Salix, mixed herbaceous, and slough area plants) were sampled seasonally from nutrient-rich and nutrient-poor areas. We measured ergosterol concentration, percentage root colonization by mycorrhizal fungi, and total bacterial counts in detrital and soil samples. Mycorrhizal fungi were not influenced by water level, and both mycorrhizal and total fungi had seasonal patterns that were influenced by the plant community. We continued these studies in greenhouse experiments. No arbuscular mycorrhizal fungal community had a consistent impact on plant growth and nutrition. For one wetland plant, flooding eliminated mycorrhizal fungal colonization, and in free-drained treatments, P amendment suppressed colonization. For the same wetland plant, some mycorrhizal communities affected shoot and root P concentrations, but there were no significant plant growth responses. For another wetland plant, the mycorrhizal association was suppressed, but not eliminated, by flooding and P amendment. Mycorrhizal colonization improved plant growth and P nutrition at lower P levels, but conferred no benefit or was detrimental at higher P levels. Objective #1 We established that the ribotypes of bacteria most widely used for bacterial source tracking, E. coli and the fecal enterococci, changed considerably with geography, time, and flow conditions. In addition, we showed that the ribotypes of E. coli changed with diet. These results were discouraging because they suggest that library-dependent BST methods will require a large host origin database (thousands of isolates) in order to encompass this genetic variability. Such a library will be time-consuming and expensive to construct. In the case were we did have libraries of sufficient size, our studies showed that library-dependent BST methods did work, and the information should provide city and county officials with information to improve impaired waters. We developed targeted sampling as a prelude to BST as a way to reduce the effects of bacterial changes with geography and time. Targeted sampling is like the childrens game of hot and cold, and by combining local knowledge, sampling, and resampling of the water during one of these two flow conditions, it was possible to identify hotspots of fecal contamination quickly and easily. By reducing the environmental complexity, we made existing BST methods quicker, more effective, and less expensive. We also extended BST research to fluorometry. This chemical BST method works by detecting optical brighteners from laundry detergents in environmental waters from malfunctioning septic drain fields and leaking sewer pipes. Because these fluorescent compounds are only associated with human sewage, the method discriminates between human and nonhuman sources of fecal contamination. Regulatory and municipal authorities in several states are currently evaluating fluorometry. The fluorescent signals appeared to be stable over seasons in different water conditions. Whenever fluorescent plumes were found, BST tests demonstrated a human signature in every case where it was performed. Therefore, our tests suggest that fluorometry was an acceptable, inexpensive method to detect human sewage in fresh and marine waters. We also tried enterococcal speciation as a simple method to discriminate between human and non-human fecal contamination, and confirmed that Ent. faecalis had a host range essentially restricted to humans and wild birds. Our results suggest that unless the fecal loading rate from migratory or resident wild birds is high, water samples collected during baseflow conditions with high numbers of Ent. faecalis indicate human fecal contamination. We started development of a species-specific primer that works in a biosensor to detect synthetic sequences of Ent. faecalis. This development is a positive first step in the development of an inexpensive new technique to detect pathogens from the environment. Objective #2 Although phytoremediation is a common and inexpensive form of environmental remediation, its efficacy is highly variable. This variability reduces our confidence in phytoremediation to remove toxic organic contaminants from the environment. Our goal was to define how biogeochemical factors and their interactions influence the phytoremediation of crude oil. We showed that adding fertilizer and establishing vegetation increased microbial populations, which, in turn, consistently reduced contaminant concentrations of crude oil. Our results suggest that agronomic practices are important to consider when developing systems to phytoremediate crude oil-contaminated sites. By assessing the degradation potential of the indigenous microbial community, determining degrader numbers, and detailing their molecular makeup, we identified important microbial information necessary to assess remediation technologies. In this manner, we enhanced studies of contaminated site ecology, and ultimately established a coherent management strategy for cleanup of crude oil-contaminated soils. For bioremediation as humic acid-sorbed PAHs, our studies show that some organisms may be uniquely suited to overcome many of the bioavailability limitations that hinder biodegradation of PAHs in soil. More broadly, because humic acids are the most pervasive organic molecules to which bacteria are exposed, niches occupied by bacteria adapted to interact with humic acids could have far-reaching implications. Objective #3 In pasture systems, agricultural management practices (e.g., crop rotations, reduced tillage, organic matter amendments, or introduction of toxins) may alter the nature of the soil environment and influence the composition and activity of the microbial community. Our studies suggest that it may be possible to manage naturally occurring plant metabolites to increase agronomic productivity or to sustain environmental quality. In the short term, our investigations with toxic alkaloids in tall fescue indicated soil responses to these alkaloids were relatively minor, but statistically significant. These compounds also increased tall fescue fitness. In the long term, soil biochemical responses to toxic alkaloids led to greater soil organic C sequestration, thereby mitigating greenhouse gas emissions. In cropping systems, our cotton management studies indicated that there is a potential for increased C storage in soils managed with reduced tillage. Less N was needed for cotton production in rotation and reduced till systems than in conventionally tilled systems. Changes in tillage practices not only increased microbial biomass C, but also shifted microbial community structure. However, how these changes affect the long-term sustainability of cotton cropping systems is still unclear. Continued use of conventional farming systems (i.e., plowing, rod-weeding, and summer fallow) were detrimental to soil quality and sustainable crop production because these systems reduce organic matter. Our data should encourage producers to convert from conventional tillage to conservation tillage (e.g., direct seed), thus reducing erosion and improving soil health by increasing the soil organic matter.

Publications

PUBLICATIONS (* denotes collaboration between two or more S-297 members) Books *Edge, T., J. Griffith, J. Hansel, V. J. Harwood, M. Jenkins, A. Layton, M. Molina, C. Nakatsu, R. Oshiro, M. Sadowsky, J. Santo Domingo, O. Shanks, G. Stelma, J. Stewart, D. Stoeckel, B. Wiggins, and J. Wilbur. 2005. Microbial source tracking guide document. U. S. Environmental Protection Agency, Office of Research and Development, EPA/600-R-05-064. *Sylvia, D. M., J. J. Fuhrmann, P. G. Hartel, and D. A. Zuberer. 2005. Principles and applications of soil microbiology, 2nd edition. Prentice-Hall, Upper Saddle River, NJ. *Sylvia, D.M., P.G. Hartel, J.J. Fuhrmann, and D. Zuberer (ed.) 2005. Instructors manual: Principles and applications of soil microbiology, 2nd edition. Prentice Hall, Upper Saddle River, NJ. Book Chapters Chaney, R. L., Y.M. Li, J. S. Angle, A. J. M. Baker, R. D. Reeves, S. L. Brown, F. A. Homer, M. Malik, and M. Chin. 2000. Improving metal hyperaccumulator wild plants to develop commercial phytoextraction systems: Approaches and progress. p. 131- 160. In N. Terry and G.S. Banuelos (ed.) Phytoremediation of contaminated soil and water. CRC Press, Boca Raton, FL. Chaney, R. L., J. A. Ryan, U. Kukier, S. L. Brown, G. Siebielec, M. Malik, and J. S. Angle. 2001. Heavy metal aspects of compost use. p. 323-360. In P. J. Stofella and B. A. Kahn (ed.) Compost utilization in horticultural cropping systems. CRC Press, Boca Raton, FL. Entry, J. A., P. A. Rygiewicz, L. S. Watrud, and P. K. Donnelly. 2002. Response of arbuscular mycorrhizae to adverse soil conditions. p. 135-158. In K. Sharma (ed.) Arbuscular mycorrhizae: Interactions in plants, rhizosphere and soils. Oxford Press, NY. Feng, Y. 2004. Microbial and photolytic degradation of 3,5,6-trichloro-2-pyridinol. p. 15-24. In J. J. Gan et al. (ed.) Deactivation (neutralization or detoxification) and safe disposal of germicides and pesticides. American Chemical Society, Washington, DC. *Fuhrmann, J. J. 2005. Microbial metabolism. p. 54-84. In D.M. Sylvia et al. (ed.) Principles and applications of soil microbiology. 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. *Germida, J. J. 2005. Transformations of sulfur. p. 433-462. In D.M. Sylvia et al. (ed.) Principles and applications of soil microbiology. 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Graham, J. H. 2000. Assessing costs of arbuscular mycorrhizal symbiosis in agroecosystems. p. 127-140. In G. K. Podila and D. D. Douds, Jr. (ed.) Current advances in mycorrhizal research. APS Press, St. Paul, MN. *Graham, J. H. 2005. Biological control of soilborne plant pathogens and nematodes. p. 562-586. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Graham, J. H., and R. M. Miller. 2005. Mycorrhizas: Gene to function. In H. Lambers and T. D. Colmer (ed.) Root ecophysiology: From gene to function. Kluwer Academic Publisher, Dordrecht, Netherlands (in press). Hartel, P.G. 2004. Environmental factors affecting microbial activity. p. 448-455. In D. Hillel, C. Rosenzweig, D. S. Powlson, K. M. Scow, M. J. Singer, D. L. Sparks, and J. Hatfield (ed.). Encyclopedia of soils in the environment. Elsevier, London. *Hartel, P. G. 2005. The soil habitat. p. 26-53. In D.M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Hickey, W. J. 2005. Microbiology and biochemistry of xenobiotic compound degradation. p. 510-535. In D.M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Jack, A. L., and J. E. Thies. 2005. Compost and vermicompost as amendments promoting soil health. In Uphoff et al. (ed.) Biological approaches to sustainable soil systems. CRC Press. (in press). Jarstfer, A.G. and D.M. Sylvia. 2001. Isolation, culture and detection of arbuscular mycorrhizal fungi. p. 535-542. In C.J. Hurst et al. (ed.) Manual of environmental microbiology, 2nd ed. American Society of Microbiology, Washington, D.C. Keeling, W. G., C. Hagedorn, B. A. Wiggins, and K. R. Porter. 2005. Bacterial source tracking: Concept and application to the TMDL. p. 207-246. In T. Younos (ed.) TMDL: Approaches and challenges. PennWell Books, Tulsa, OK. *Morton, J. B. 2005. Fungi. p. 141-161. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Morton, J. B., R. E. Koske, S. L. Stuermer, and S. P. Bentivenga. 2004. Mutualistic arbuscular endomycorrhizal fungi. p. 317-336. In: G. M. Mueller, G. F. Bills, and M. S. Foster (ed.) Biodiversity of fungi: inventory and monitoring methods. Smithsonian Institution Press, Washington, DC. *Mullen, M.D. 2005. Phosphorus and other elements. p.463-488. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. Pearson-Prentice Hall Publishers, Upper Saddle River, NJ. *Nakatsu, C. H. 2005. Fundamentals of microbial genetics. p. 85-98. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Nakatsu, C. H. 2004. Microbial community analysis. p. 455-463. In: D. Hillel, C. Rosenzweig, D. Powlson, K. Scow, M. Singer and D. Sparks (ed.) Encyclopedia of soils in the environment. Elsevier, Oxford. Nakatsu, C. H. and L. J. Forney. 2004. Parameters of nucleic acid hybridization experiments. In: G. A. Kowalchuk, F.J. de Bruijn, I. M. Head, A.D.L. Akkermans, and J.D. van Elsas (ed.) Molecular microbial ecology manual, 2nd ed. Springer Publishers, Netherlands. *Reynolds, C. M., and H. D. Skipper. 2005. Bioremediation of contaminated soils. p. 536-561. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Schnabel, R. R., A. J. Franzluebbers, W. L. Stout, M. A. Sanderson, and J. A. Stuedemann. 2001. The effects of pasture management practices. p. 291-322. In R. F. Follett et al. (ed.) The potential of U. S. grazing lands to sequester carbon and mitigate the greenhouse effect. CRC Press, Boca Raton, FL. Schröder, E. C. 2001. Importance of symbiotic nitrogen fixation in tropical forage legume production. p. 251-268. In A. Sotomayor-Ríos and W. D. Pitman (ed.) Tropical forage plants: Development and use. CRC Press, Boca Raton. FL. Sojka, R. E., D. L. Bjorneberg, and J. A. Entry. 2002. Irrigation, a historical perspective. p. 745-749. In R. Lal (ed). Encyclopedia of Soil Science. Marcel Dekker, New York. Sojka, R. E., W. J. Orts, and J. A. Entry. 2004. Soil physics and hydrology: Conditioners. P. 301-306. In: D. Hillel, C. Rosenzweig, D. Powlson, K. Scow, M. Singer and D. Sparks (ed.) Encyclopedia of soils in the environment. Elsevier, Oxford. Stromberger, M. E. 2005. Fungal communities of agroecosystems. p. 813-832. In J. Dighton et al. (ed.) The fungal community. CRC Press, Boca Raton, FL. *Sylvia, D. M. 2005. Mycorrhizal symbioses. p. 263-282. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. Pearson-Prentice Hall Publishers, Upper Saddle River, NJ. Thies, J. E., and J. M. Grossman. 2005. Soil ecology and the soil habitat. In Uphoff et al. (ed.) Biological approaches to sustainable soil systems. CRC Press, Boca Raton, FL (in press). *Wolf, D. C., and G. H. Wagner. 2005. Carbon transformations and soil organic matter formation. p. 285-332. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology. 2nd ed. Pearson-Prentice Hall publishers, Upper Saddle River, NJ. *Wollum, A. G., and D. A. Zuberer. 2005. Introduction and historical perspective. p. 3-25. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology. 2nd ed. Pearson-Prentice Hall publishers, Upper Saddle River, NJ. *Zuberer, D. A. 2005. Biological dinitrogen fixation: Introduction and nonsymbiotic. p. 373-404. In D. M. Sylvia et al. (ed.) Principles and applications of soil microbiology, 2nd ed. PearsonPrentice Hall Publishers, Upper Saddle River, NJ. Journal Articles Alagely, A. K., D. M. Sylvia, and L. Ma. 2005. Mycorrhizae increase arsenic uptake by a hyperaccumulating fern. J. Environ. Qual. (in press). Allen, L. H. Jr., S. L. Albrecht, W. Colón, S. A. Covell, J. T. Baker, D. Pan, and K. J. Boote. 2003. Methane emissions of rice increased by elevated carbon dioxide and temperature. J. Environ. Qual. 32:1978-1991. Angle, J. S., R. L. Chaney, A. J. M. Baker, Y.-M. Li, R. Reeves, V. Volk, R. Roseberg, E. Brewer, S. Burke, and J. P. Nelkin. 2001. Developing commercial phytoextraction technologies: Practical considerations. S. Afr. J. Sci. 97:619-623. Augé, R. M., J. L. Moore, K. Cho, J. C. Stutz, D. M. Sylvia, A. K. Al-Agely, and A. M. Saxton. 2003. Relating drought resistance of Phaseolus vulgaris to soil and root colonization by mycorrhizal hyphae. J. Plant Physiol. 160:1147-1156. Augé, R. M., J. L. Moore, D. M. Sylvia, and K. Cho. 2004. Mycorrhizal promotion of host stomatal conductance in relation to irradiance and temperature. Mycorrhiza 14:85-92. Baldwin, B., C. H. Nakatsu, and L. Nies. 2003. Detection and enumeration of aromatic oxygenase genes by multiplex and real-time PCR. Appl. Environ. Microbiol. 69:3350-3358. Bever, J. D., P. A. Schultz, A. Pringle, and J. B. Morton. 2001. Arbuscular mycorrhizal fungi: More diverse than meets the eye and the ecological tale of why. BioScience 51:923-931. Bigelow, C. A., D. C. Bowman, and A. G. Wollum. 2002. Characterization of soil microbial population dynamics in newly constructed sand-based rootzones. Crop Sci. 42:1611-1614. Booth, A. M., A. K. Graves, C. Hagedorn, S. C. Hagedorn, and K. H. Mentz. 2003. Sources of fecal pollution in Virginias Blackwater River. J. Environ. Eng. 129:547-552. Bray, S. R., K. Kitajima, and D. M. Sylvia. 2003. Mycorrhizae differentially alter growth, physiology, and competitive ability of an invasive shrub. Functional Ecol. 13:565-574. Bressan, W., C. H. S. de Carvalho, and D. M. Sylvia. 2000. Inoculation of somatic embryos of sweet potato with an arbuscular mycorrhizal fungus improves embryo survival and plantlet formation. Can. J. Microbiol. 46:741-743. Burke, S., J. S. Angle, R. L. Chaney, and S. Cunningham. 2000. Arbuscular Mycorrhizae effects on heavy metal uptake by corn. Int. J. Phytoremed. 2:23-29. Chang, Y-J., A. K. M. Anwar Hussain, J. R. Stephens, M. D. Mullen, D. C. White, and A.D. Peacock. 2001. Impact of herbicides on the abundance and structure of indigenous subgroup ammonia oxidizer communities in soil microcosms. Environ. Toxicol. Chem. 20:2462-2468. Coffin, R. B., P. H. Miyares, C. A. Kelly, L. A. Cifuentes, and C. M. Reynolds. 2001. 13C and 15N isotope analysis of TNT: Two dimensional source identification. Environ. Tox. Chem. 20:2676-2680. Curtis, P., C. H. Nakatsu, and A. Konopka. 2002. Aciduric proteobacteria isolated from pH 2.9 soil. Arch. Microbiol. 178:65-70. Darnault, C. J. G., P. Garnier, Y.-J. Kim, K. Oveson, T. S. Steenhuis, J. Y. Parlange, M. B. Jenkins, W. C. Ghiorse, and P. C. Baveye. 2003. Transport of Cryptosporidium parvum oocysts in the subsurface environment. Water Environ. Res. 75:113-120. Darnault, C. J. G., T. S. Steenhuis, P. Garnier, Y.-J. Kim, M. B. Jenkins, W. C. Ghiorse, P. C. Baveye, and J.-Y. Parlange. 2004. Preferential flow and transport of Cryptosporidium parvum oocysts through the vadose zone: Experiments and modeling. Vadose Zone J. 3:262-270. Davis, K. C., C. H. Nakatsu, R. Turco, S. Weagant, and A. K. Bhunia. 2003. Analysis of environmental Escherichia coli isolates for virulence genes using the TaqMan PCR system. J. Appl. Microbiol. 95:612-620. Delorme, T., J. S. Angle, F. Coale, and R. Chaney. 2001. Phosphorus accumulation by select plant species. Inter. J. Phytoremed. 2:125-131. Devare, M., C. M. Jones, and J. E. Thies. 2004. Effects of CRW transgenic corn and tefluthrin on the soil microbial community: Biomass, activity, and diversity. J. Environ Qual. 33:837-843. Elliott, M. L., E. A. Guertal, E. A. Des Jardin, and H. D. Skipper. 2003. Effect of nitrogen rate and root-zone mix on rhizosphere bacterial populations and root mass in creeping bentgrass putting greens. Biol. Fertil. Soils 37:348-354. Elliott, M. L., E. A. Guertal, and H. D. Skipper. 2004. Rhizosphere bacterial population flux in golf course putting greens in the southeastern United States. HortScience 39:1754-1758. Entry, J. A., and N. Farmer. 2001. Movement of coliform bacteria and nutrients in groundwater flowing through basalt and sand aquifers. J. Environ. Qual. 30:1533-1539. *Entry, J. A., J. J. Fuhrmann, R. E. Sojka, and G. E. Shewmaker. 2004. Influence of irrigated agriculture on soil carbon and microbial community structure. Environ. Manage. 33:S363-S373. *Entry, J. A., R. K. Hubbard, J. E. Thies, and J. J. Fuhrmann. 2000. The influence of vegetation in riparian filterstrips on coliform bacteria. I. Movement and survival in water. J. Environ. Qual. 29:1206-1214. *Entry, J. A., R. K. Hubbard, J. E. Thies, and J. J. Fuhrmann. 2000. The influence of vegetation in riparian filterstrips on coliform bacteria. II. Survival in soils. J. Environ. Qual. 29:1215-1224. *Entry, J. A., I. Phillips, H. Stratton, and R. E. Sojka. 2001. Efficacy of polyacrylamide+Al(SO4)3 and polyacrylamide+CaO to filter microorganisms and nutrients from animal wastewater. Environ. Pollut. 121:453-462. Entry, J. A., P. A. Rygiewicz, L. S. Watrud, and, P. K. Donnelly. 2001. The influence of adverse soil conditions on formation and function of arbuscular mycorrhizae. Adv. Environ. Res. 7:123-138. Entry, J. A. and R. E. Sojka. 2003. The efficacy of polyacrylamide to reduce nutrient movement from an irrigated field. Trans. ASAE. 46:75-83. Entry, J. A., R. E. Sojka, and G. Shewmaker. 2003. Management of irrigated agriculture to increase carbon storage in soils. Environ. Management 33:S309-S317. Entry, J. A., R. E. Sojka, M. E. Watwood, and C. Ross. 2002. Environmental and agricultural applications of polyacrylamide preparations. Environ. Pollut. 120:191-200. Entry, J. A., C. A. Strausbaugh, and R. E. Sojka. 2001. Wood chip - polyacrylamide cores as a carrier for biocontrol bacteria suppresses Verticllium wilt on potato. Biocontrol Sci. Technol. 10:677-686. Fang, C., M. Radosevich, and J. J. Fuhrmann. 2001. Characterization of rhizosphere microbial community structure in five similar grass species using FAME and BIOLOG Analyses. Soil Biol. Biochem. 33:679-682. Fang, C., M. Radosevich, and J. J. Fuhrmann. 2001. Atrazine and phenanthrene degradation in grass rhizosphere soil. Soil Biol. Biochem. 33:671-678. Feng, Y., A. C. Motta, D. W. Reeves, C. H. Burmester, E. van Santen, and J. A. Osborne. 2003. Soil microbial communities under conventional-till and no-till continuous cotton systems. Soil Biol. Biochem. 35:1693-1703. Feng, Y., D. M. Stoeckel, E. van Santen, and R. H. Walker. 2002. Effects of subsurface aeration and Trinexapac-ethyl application on soil microbial communities in a creeping bentgrass putting green. Biol. Fertil. Soils. 36:456-460. *Franke-Snyder, M., D. D. Douds, L. Galvez, J. G. Phillips, P. Wagoner, L. Drinkwater, and J. B. Morton. 2001. Diversity of communities of arbuscular mycorrhizal (AM) fungi present in conventional versus low-input agricultural sites in eastern Pennsylvania, USA. Appl. Soil Ecol. 16:35-48. Franzluebbers, A. J., and N. S. Hill. 2005. Soil carbon, nitrogen, and ergot alkaloids with short- and long-term exposure to endophyte-free and -infected tall fescue. Soil Sci. Soc. Am. J. 69:404-412. Franzluebbers, A. J., and J. A. Stuedemann. 2005. Soil carbon and nitrogen pools in response to tall fescue endophyte infection, fertilization, and cultivar. Soil Sci. Soc. Am. J. 69:396-403. Fuentes, J. P., D. F. Bezdicek, M. Flury, S. L. Albrecht, and J. L. Smith. 2005. Microbial activity affected by lime in a long term no-till soil. Soil Tillage Res. (in press). *Gagliardi, J. V., J. S. Angle, J. J. Germida, R. C. Wyndham, C. P. Chanway, R. J. Watson, C. Greer, T. McIntyre, H. H. Yu, M. A. Levin, E. Russek-Choen, S. Rosolen, J. Nairn, A. Seib, T. Martin-Heller, and G. Wisse. 2001. Intact soil-core microcosms for pre-release testing of introduced microbes: Comparison with multi-site field releases in diverse soils and climates. Can. J. Microbiol. 47:237-252 Gagliardi, J. V., J. S. Buyer, J. S. Angle, and E. Russek-Cohen. 2001. Structural and functional analysis of whole-soil microbial communities for risk and efficacy testing following microbial inoculation of wheat roots in diverse soils. Soil Biol. Biochem. 33:25-40. *Gentry, T. 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Carbon source utilization profiles as a method to identify sources of fecal pollution in water. J. Appl. Microbiol. 94:792-799. Hahm, B. K., Y. Maldonado, E. Schreiber, A. K. Bhunia, and C. H. Nakatsu. 2003. Subtyping of clinical and environmental isolates of Escherichia coli by multiplex PCR, AFLP, rep-PCR, PFGE and ribotyping. J. Microbiol. Methods 53:387-399. *Haney, R. L., A. J. Franzluebbers, F. M. Hons, and D. A. Zuberer. 2001. Molar concentration of K2SO4 and soil pH affect estimation of extractable C with chloroform fumigation-extraction. Soil Biol. Biochem. 33:1501-1507. Haney, R. L., S. A. Senseman, F. M. Hons, and D. A. Zuberer. 2000. Effect of glyphosate on soil microbial activity and biomass. Weed Sci. 48:8993. Hartel, P.G., K. Gates, K. Payne, J. McDonald, K. Rodgers, S. Hemmings, J. Fisher, and L. Gentit. 2005. Targeted sampling to determine sources of fecal contamination. Stormwater 6:46-53. Hartel, P. G., E. A. Frick, A. L. Funk, J. L. Hill, J. D. Summer, and M. B. 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