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Microbiological & Chemical Exposure Assessment Research Division Publications: 2007
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This page lists publication titles, citations and abstracts produced by NERL's Microbiological & Chemical Exposure Assessment Research Division for the year 2007, organized by Publication Type. Your search has returned
83 Matching Entries.
See also Microbiological & Chemical Exposure Assessment Research Division citations with abstracts:
1999,
2000,
2001,
2002,
2003,
2004,
2005,
2006,
2007,
2008
Technical Information Manager: Angela Moore - (513) 569-7341 or moore.angela@epa.gov
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Detection of Protozoan Parasites in Source and Finished Water 3rd Edition Asm's Methods in Environmental Microbiology |
05/01/2007
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SCHAEFER, F. W. Detection of Protozoan Parasites in Source and Finished Water 3rd Edition Asm's Methods in Environmental Microbiology. 3rd.Chapter 21, Ronald L. Crawford, Jay L. Garland, David A. Lipson, Aaron L. Mills, Linda D. Stetzenbach (ed.), Methods in Environmental Microbiology. American Society for Microbiology, Washington, DC, 265-279, (2007).
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Protozoans are eukaryotic organisms which can live either a free-living or parasitic existence. Some free-living forms, under the right conditions, can become opportunistic parasites. Enteric pathogenic protozoans, like Giardia and Cryptosporidium, which are now known to be transmitted by water have been responsible for numerous waterborne outbreaks of gastroenteritis. The primary means for detection, since density levels in water are low, involves processing a large volume of water by filtration, extracting the particulates from the filter, concentrating the organisms from the particulates, and assaying for the pathogens. The most widely used method for detecting protozoans has been the indirect immunofluorescent assay. While Method 1623 has improved upon the utility of the immunofluorescent assays for Giardia and Cryptosporidium, the procedure is still labor intensive and highly dependent on the skill of the microscopist. Even with the improvements to date, this technique is known to have a number of deficiencies including false positives, inability to determine the viability and species of the detected organisms, and low average percent recovery of cysts and oocysts. Various attempts have been made to improve the immunofluorescent detection method. Rather than sampling by filtration and buoyant density centrifugation to purify the organisms, carbonate flocculation is reported to improve recoveries. PCR, cell culture, FACS, and solid phase cytometry are currently being evaluated as alternate test procedures. As each of these approaches is relatively new and much more research is needed, it remains to be seen whether they will be equal to or better than the current fluorescent assay procedure.
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| BOOK CHAPTER |
Genetic-Based Analytical Methods for Bacteria and Fungi |
03/01/2007
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HAUGLAND, R. A. AND S. J. VESPER. Genetic-Based Analytical Methods for Bacteria and Fungi. , Chapter 7, C.S. Yang and P. Heinsohn (ed.), Sampling and Analysis of Indoor Micoorganisms. John Wiley & Sons Incorporated, New York, NY, 133-152, (2007).
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In the past two decades, advances in high-throughput sequencing technologies have lead to a veritable explosion in the generation of nucleic acid sequence information (1). While these advances are illustrated most prominently by the successful sequencing of the human genome, they have also factored heavily in our current knowledge of nucleic acid sequences from a variety of microorganisms. Concurrent with this growth in sequencing activity has been the development of a variety of techniques for the amplification and manipulation of nucleic acids. The combination of these technologies is currently supporting a significant shift away from the use of traditional culture-based analyses for the detection and characterization of microorganisms and towards the use of new analytical methods based on genetic composition, i.e. nucleic acids, (45, 113). This shift is evident in a number of fields including the microbiological analyses of indoor environments.
Genetic analysis methods for bacterial and fungal microorganisms are presently becoming increasingly more widespread in their applications, not only in research settings, but also in clinical and environmental testing laboratories. Advantages of these methods over culture and phenotypic analyses can include higher speed, sensitivity and accuracy in the detection and identification of microorganisms as well as better resolution between similar organisms and an ability to detect non-cultivatable or fastidious organisms. Through their potential for automation, these methods also offer the possibility for less reliance on analyst training and decreased labor investments. As will be discussed in more detail below, current limitations of many of these genetic analysis methods can include on-going uncertainty of the extent to which available sequence information circumscribes the genetic variability of different target microbial groups, technical and quality control issues and higher costs related to expenditures for instruments and reagents. It is reasonable to expect, however, that each of these limitations will decrease in the future making genetic microbial testing methods an increasingly attractive option for many clinical and environmental applications.
The first section of this chapter provides an overview (with references for additional reading) of currently available genetic based analytical techniques that may be useful for investigations of bacterial and fungal microorganisms in the indoor environment. These techniques are grouped into four general categories: 1) in vitro nucleic acid amplification; 2) hybridization probes; 3) nucleic acid sequencing; and 4) microbial strain typing. The second section provides example applications of techniques within each of these categories for the study of indoor microbiology. The third section provides a synopsis of quality assurance issues and presently accepted quality control measures for laboratories performing genetic analysis methods, focusing primarily on methods involving nucleic acid amplification techniques. Other current technical limitations of these methods and their outlook for the future are also discussed.
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| BOOK CHAPTER |
The Lognormal Distribution and Use of the Geometric Mean and the Arithmetic Mean in Recreational Water Quality Measurement |
11/01/2007
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WYMER, L. J. AND T. J. WADE. The Lognormal Distribution and Use of the Geometric Mean and the Arithmetic Mean in Recreational Water Quality Measurement. , Chapter 6, Larry Wymer (ed.), Statistical Framework for Recreational Water Quality Criteria and Monitoring. John Wiley and Sons, LTD, , Uk, 91-112, (2007).
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Since 1968 United States recreational water quality criteria have set a limit on the geometric mean for fecal indicator bacteria from a number water samples taken over a period of time (National Technical Advisory Committee, 1968; U.S. Environmental Protection Agency, 1976 and 1986). On the other hand, for purposes of determining limits on effluents, including sewage, discharged into surface waters, the U.S. EPA specifies that calculations for all limitations which require averaging of measurements shall utilize an arithmetic mean unless otherwise specified by the Director in the permit (U.S. EPA, 1980, 2003). These limits, a geometric mean criterion for beaches and arithmetic mean for discharges, both pertain to provisions of the Clean Water Act of 1977 (CWA) as amended by the Beaches Environmental Assessment and Coastal Health (BEACH) Act of 2000. In addition to this disagreement between types of means that are used in beach monitoring and those used in limiting discharges, a trio of paper published in the late 1990's evaluated uses of the geometric mean and reached conclusions such as the use of this statistic is inappropriate for characterizing risk (Haas, 1996) and geometric means should be phased out as regulatory criteria as soon as it is practical (Parkhurst, 1998a). Statements such as these serve to further create doubt about the appropriateness of geometric means in the minds of federal and state regulators and stakeholders.
This chapter examines criticisms of the use of the geometric mean in risk assessment and environmental monitoring and evaluates its relevance to risk-based recreational water monitoring. Reasons for using the geometric mean (or rather the mean of the logarithms of the indicator densities, as we shall see) in modeling risk attributable to swimming in contaminated waters are explored and alternative models examined.
In the course of this discussion, we will refer to properties of the normal and lognormal probability distributions. For reference, a comparison of some characteristics of normal and lognormal distributions is presented in Table 1. The interested reader can find more detailed information and discussions in Crow and Shimizu (1988), Aitchison and Brown (1969) or Johnson and Kotz (1970) among other works. Estimation of lognormal parameters specifically in the context of environmental monitoring is discussed in Gilbert (1987).
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| BOOK CHAPTER |
The Empact Beaches: A Case Study in Recreational Water Sampling |
11/01/2007
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WYMER, L. J. The Empact Beaches: A Case Study in Recreational Water Sampling. , Chapter 7, Larry Wymer (ed.), Statistical Framework for Recreational Water Quality Criteria and Monitoring. John Wiley and Sons, LTD, , Uk, 113-134, (2007).
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Various chapters describe sample and experimental design, use of a geometric mean or an arithmetic mean, modeling and forecasting, and risk assessment in relation to monitoring recreational waters for fecal indicators. All of these aspects of monitoring are dependent on the spatial and temporal distribution of fecal indicator bacteria in the water. Knowledge of the distribution of indicator bacteria in water is particularly important in sampling design for monitoring. The U.S. Environmental Protection Agency (EPA) conducted intensive water sampling studies during the summer of 2000 at five beaches in order to characterize temporal and spatial distribution of fecal indicator bacteria within the bathing areas of these beaches (Wymer et al., 2005, hereafter referred to as the EMPACT report). Study beaches encompassed a variety of environments (Table 1).
During the months of July and August in 2000, water samples were collected at least twice daily from each of nine "fixed" locations in the water, as determined by a transect and zone (Figure 1). A transect consisted of an imaginary line through a fixed point on the beach perpendicular to the shoreline. A zone (or "depth zone") was defined as a contour line of equal water depth. As illustrated in Figure 1, each sampling location was defined by the intersection of transect and zone on a grid comprising three transects and three zones projected on the water's surface. A random point along the shoreline within the recognized beach area was selected to define the first transect (the leftmost transect in Figure 1). The middle transect was, then, determined as the parallel to the first transect at a distance of 20 meters, and the remaining transect, as the parallel to the middle transect at an additional distance of 20 meters, or 40 meters from the first transect.
Locations at which the water attained a constant depth of 0.15, 0.5, and 1.3 meters (1.0 meters at Belle Isle, since buoys demarcating the swimming area were located at approximately this depth), corresponding to "ankle deep", "knee deep," and "chest deep" water, were selected as sampling zones. Because three of the beaches were affected by ocean tides, the actual geographic locations of these zones varied according to the tide stage. Hence, sampling locations at these beaches were fixed only in the sense that they correspond to locations at which the water depth was constant. Note the use of the term "zone" rather than "depth" in referring to areas of different water depth. This is to avoid confusion between water depth, which defines the sampling locations in Figure 1, and sampling depth, the depth below the surface of the water from which the samples were taken. In knee deep and chest water, the sampling depth was nearly always 0.3 meters from the surface, which is approximately "face deep" for a swimmer. A relatively small number of samples were taken from other sampling depths as part of a parallel study. Zones may be considered as roughly corresponding to different activities among bathers, mainly wading and infant or toddler bathing (ankle deep water), play (knee deep), and swimming or diving (chest deep).
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| BOOK CHAPTER |
The Evolution of Water Quality in the United States 1922-2003 |
11/01/2007
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DUFOUR, A. P. AND S. SCHAUB. The Evolution of Water Quality in the United States 1922-2003. , Chapter 1, L. J. Wymer (ed.), Statistical Framework for Recreational Water Quality Criteria and Monitoring. John Wiley and Sons, LTD, , Uk, 1-12, (2007).
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The microbiological quality of recreational waters was first discussed in the United States as early as 1922 by the American Public Health Association's Committee on Bathing Beaches (APHA,1922) . The Committee surveyed 2000 physicians and state health officials inquiring about the prevalence of infections associated with bathing places. The Committee Report in 1924 (APHA,1924) reviewed the survey results and concluded that there was not enough evidence to develop bathing water standards for natural waters. In June, 1933 the Joint Committee on Bathing Places was formed and in their first report noted that because of the great lack of epidemiological information no bacterial standards were adopted (APHA, 1933). They also stated that the Committee did not want to propose arbitrary standards or measures that might promote public hysteria about the dangers of outdoor bathing places. The Committee, in 1936, was still not convinced that bathing places were a major health problem and re-stated their position on developing bacterial standards for bathing places. The reluctance to propose bacterial standards for outdoor bathing places was again evident in 1936, 1940 and 1955 (APHA, 1936, APHA, 1940, APHA, 1957). The Committee did attempt to find evidence of the risk of illness from bathing waters prior to their reports in 1940 and 1955, but they found no compelling evidence. They stated that very little reliable data were available to implicate bathing places in the spread of disease (APHA,1957).
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| JOURNAL |
Changes in Mouse Cirulating Leukocyte Numbers in C57bl/6 Mice Immunosuppressed for Cryptosporidium Parvum Oocyst Production |
01/01/2007
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Miller, T. A. AND F. W. Schaefer III. Changes in Mouse Cirulating Leukocyte Numbers in C57bl/6 Mice Immunosuppressed for Cryptosporidium Parvum Oocyst Production. Veterinary Parasitology. Elsevier, Shannon, Ireland, 143:99-105, (2007).
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The Iowa strain of Cryptosporidium parvum will not propagate in immunocompetent mice, but will successfully infect genetically immunocompromised Nude or SCID mice as well as immunocompetent mice which have been immunosuppressed with glucocorticoids. Using dexamethasone - tetracycline is one method for immunosuppressing mice for the production of C. parvum oocysts. However, dexamethasone induced immunosuppression is variable, because it is dependent on the total daily water consumption of each individual mouse. In an attempt to more accurately characterize the immunocompromised state for future studies on the infectivity of Cryptosporidium in immunocompromised mice, the changes in circulating leukocytes and other immune system associated organs before, during and after dexamethasone suppression were analyzed. The dexamethasone induced immunocompromised state was associated with greater than 90 percent sustained drop in the circulating T-lymphocyte count, a greater than 700 percent increase in circulating mature segmented neutrophils and a severe depletion of circulating monocytes. The thymus and spleen decreased in size by over 80 percent. Oocyst shedding in suppressed mice started within four days of oocyst inoculation and persisted for six days after dexamethasone withdrawal. Circulating neutrophils rose dramatically by 714 percent while on dexamethasone; seven days after dexamethasone withdrawal circulating neutrophils still were 549 percent higher than normal. Circulating CD3 and CD4 lymphocytes remained 85 to 90 percent below normal while on dexamethasone and for seven days after discontinuing dexamethasone. CD8 lymphocyte numbers initially decreased by 90 percent, but rose even while on dexamethasone and even with severe thymic involution. At day seven post dexamethasone treatment, the spleen was 119 mm3, approximating normal. After fourteen days of dexamethasone withdrawal, the CD8 counts were only 1.6 percent below normal while the CD3 and CD4 counts were still 66 percent below normal. The thymus now was about three quarters of its normal size. The rise in circulating CD8 lymphocytes when oocyst production stopped suggests that CD8 positive lymphocytes may play a significant role in vivo in clearing the parasite.
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| JOURNAL |
A Critical Evaluation of a Flow Cytometer Used for Detecting Enterococci in Recreational Waters |
03/31/2007
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STANG, D., K. P. BRENNER, AND M. R. RODGERS. A Critical Evaluation of a Flow Cytometer Used for Detecting Enterococci in Recreational Waters. JOURNAL OF WATER AND HEALTH. IWA Publishing, London, Uk, 5(2):295-305, (2007).
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The current U. S. Environmental Protection Agency-approved method for enterococci (Method 1600) in recreational water is a membrane filter (MF) method that takes 24 hours to obtain results. If the recreational water is not in compliance with the standard, the risk of exposure to enteric pathogens may occur before the water is identified as hazardous. Because flow cytometry combined with specific fluorescent antibodies has the potential to be used as a rapid detection method for microorganisms, this technology was evaluated as a rapid, same-day method to detect enterococci in bathing beach waters. The flow cytometer chosen for this study was a laser microbial detection system designed to detect labeled antibodies. A comparison of MF counts with flow cytometry counts of enterococci in phosphate buffer and sterile-filtered recreational water showed good agreement between the two methods. However, when flow cytometry was used, the counts were several orders of magnitude higher than the MF counts with no correlation to Enterococcus spike concentrations. The unspiked sample controls frequently had higher counts than the samples spiked with enterococci. Particles within the spiked water samples were probably counted as target cells by the flow cytometer because of autofluorescence or non-specific adsorption of antibody and carryover to subsequent samples. For these reasons, this technology may not be suitable for enterococci detection in recreational waters. Improvements in research and instrument design that will eliminate high background and carryover may make this a viable technology in the future.
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| JOURNAL |
Relative Moldiness Index© as Predictor of Childhood Respiratory Illness |
01/01/2007
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VESPER, S. J., C. MCKINSTRY, R. A. HAUGLAND, Y. IOSSIFOVA, G. LEMASTERS, L. LEVIN, G. K. HERSHEY, M. VILLAREAL, D. I. BERNSTEIN, J. LOCKEY, AND T. REPONEN. Relative Moldiness Index© as Predictor of Childhood Respiratory Illness. Journal of Exposure Science and Environmental Epidemiology . Nature Publishing Group, London, Uk, 17(1):88-94, (2007).
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The results of a traditional visual mold inspection were compared to a mold evaluation based on the Relative Moldiness Index (RMI). The RMI is calculated from mold specific quantitative PCR (MSQPCR) measurements of the concentation of 36 species of molds in floor dust samples. These two prospective mold evaluations were used to classify the mold condition in 271 homes of infants. Later, the development of respiratory illness was measured in the infants living in these homes and the predictive value of each classification system evaluated. The binary classification of homes as either moldy or non-moldy by on-site vidual home inspection was not predictive of the development of respiratory illness (wheeze and/or rhinitis) (p=0.27). Conversely, a method developed and validated in this paper using the RMI index fit to a logistic function, can be used to predict the occurrence of illness in homes and allows stakeholders the choice among various levels of risk.
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| JOURNAL |
Dimethylthioarsinic Anhydride: A Standard for Arsenic Speciation |
01/30/2007
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FRICKE, M., M. ZELLER, W. R. CULLEN, M. R. WITKOWSKI, AND J. T. CREED. Dimethylthioarsinic Anhydride: A Standard for Arsenic Speciation. ANALYTICA CHIMICA ACTA. Elsevier Science Ltd, New York, NY, 583(1):78-83, (2007).
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Dimethylthioarsinic acid (DMTAV) has recently been identified in biological, dietary and environmental matrices. The relevance of this compound to the toxicity of arsenic in humans is unknown and further exposure assessment and metabolic studies are difficult to conduct because of the unavailability of a well characterized standard. The synthesis of DMTAV was accomplished by the reaction of dimethylarsinic acid (DMAV) with hydrogen sulfide. The initial reaction product produced is DMTAV but multiple products over the course of the reaction are also observed. Therefore, a chromatographic separation was developed to monitor the reaction progress via LC ICP MS. In this synthesis, conversion of DMAV to DMTAV was not taken to completion to avoid the production of side products. The product was isolated from the starting material by standard organic techniques. Single crystal diffraction demonstrated that solid DMTAV is present in the form of the oxygen-bridged dimethylthioarsinic anhydride. Dissolution of the anhydride in water produces the acid form of DMTAV and the aqueous phase DMTAV provided a characteristic molecular ion of m/z 155 by LC ESI-MS. The synthesis and isolation of dimethylthioarsinic anhydride provides a stable crystalline standard suitable for identification, toxicological study and exposure assessment of dimethylthioarsinic acid.
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| JOURNAL |
Differentiation of Aeromonas Isolates Obtained from Drinking Water Distribution System Using Matrix-Assisted Laser Description/Ionization-Mass Spectrometry (Maldi-MS) |
02/02/2007
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DONOHUE, M. J., J. BEST, W. SMALLWOOD, M. KOSTICH, M. R. RODGERS, AND J. A. SHOEMAKER. Differentiation of Aeromonas Isolates Obtained from Drinking Water Distribution System Using Matrix-Assisted Laser Description/Ionization-Mass Spectrometry (Maldi-MS). ANALYTICAL CHEMISTRY. American Chemical Society, Washington, DC, 79(5):1939-1946, (2007).
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The genus Aeromonas is one of several medically significant genera that have gained prominence due to their evolving taxonomy and controversial role in human diseases. In this study, matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) was used to analyze the whole cells of both reference strains and unknown Aeromonas isolates obtained from water distribution systems. A library of over 45 unizue m/z signatures was created from 40 strains that are representative of the seventeen recognized species of Aeromonas, as well as three reference strains from genus Vibrio and two reference strains from Plesiomonas shigelloides. The library was used to help speciate 52 isolates of Aeromonas. The environmental isolates were broken up into two blind studies. Group 1 contained isolates that had a recognizable phenotypic profile and Group 2 contained isolates that had an atypical phenotypic profile. MALDI-MS analysis of the water isolates in Group 1 matched the phenotypic identification in all cases. In Group 2, the MALDI-MS based determination confirmed the identity of 18 of the 27 isolates. These results demonstrate the MALDI-MS analysis can rapidly and accurately classify species of the genus Aeromonas, making it a powerful tool especially suited for environmental monitoring and detection of microbial hazards in drinking water.
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| JOURNAL |
Characterization of a Cryptosporidium Muris Infection and Reinfection in Cf-1 Mice |
03/31/2007
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MILLER, T. A. Characterization of a Cryptosporidium Muris Infection and Reinfection in Cf-1 Mice. Veterinary Parasitology. Elsevier, Shannon, Ireland, 144(3-4):208-221, (2007).
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To establish control values for circulating cells and immune associated organs over the course of a self-limiting Cryptosporidium muris infection and rechallenge infection, mice were evaluated at intervals starting before oral inoculation and ending after oocyst shedding had ceased. These values were used in other experiments to evaluate changes in these parameters induced by a single dose glucocorticoid immunosuppression model and in other immunosuppression studies.
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| JOURNAL |
Blind Trials Evaluating in Vitro Infectivity of Cryptosporidium Parvum Oocysts Using Cell Culture Immunofluorescence |
05/01/2007
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BUKHARI, Z., D. M. HOLT, M. W. WARE, AND F. W. SCHAEFER. Blind Trials Evaluating in Vitro Infectivity of Cryptosporidium Parvum Oocysts Using Cell Culture Immunofluorescence. CANADIAN JOURNAL OF MICROBIOLOGY. NRC Research Press, Ottawa, Canada, 53(5):656-663, (2007).
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An optimized cell culture-immunofluorescence (IFA) procedure, using the HCT-8 cell line, was evaluated in 'blind' trials to determine the sensitivity and reproducibility for measuring infectivity of flow cytometry prepared inocula of C. parvum oocysts. In separate trials, suspensions consisting of between 0-100% viable oocysts were prepared at the U.S. EPA, shipped to the American Water (AW) laboratory and analyzed 'blindly' by cell culture-IFA. Data indicated the control (100% live) oocyst suspensions yielded statistically similar results to a cell culture dose response curve data developed previously at AW. For test samples containing oocyst suspensions of unknown infectivity, cell culture-IFA analyses indicated a high degree of correlation (r2= 0.89; n=26) with the values expected by U.S. EPA. Cell culture infectivity correlates well with neonatal mouse infectivity assays and these 'blind' validation trials provide credibility for the cell culture-IFA procedure as a cost-effective and expedient alternative to mouse infectivity assays for determining in vitro infectivity of C. parvum oocysts.
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| JOURNAL |
Quantitative Pcr Analysis of Molds in the Dust from Homes of Asthmatic Children in North Carolina |
08/03/2007
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VESPER, S. J., C. MCKINSTRY, P. ASHLEY, R. A. HAUGLAND, K. YEATTS, K. D. BRADHAM, AND E. R. SVENDSEN. Quantitative Pcr Analysis of Molds in the Dust from Homes of Asthmatic Children in North Carolina. JOURNAL OF ENVIRONMENTAL MONITORING. Royal Society of Chemistry, Cambridge, Uk, 9(8):826-830, (2007).
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The vacuum bag (VB) dust was analyzed by mold specific quantitative PCR. These results were compared to the analysis survey calculated for each of the homes. The mean and standard deviation (SD) of the ERMI values in the homes of the NC asthmatic children was 16.4 (6.77), compared to the HUD survey p = 0.003 in the NC asthmatic children's homes. The molds Chaetomium globosum, Aspergillus fumigatus, and Eurotium in the NC homes of asthmatics making the ERMI values significantly higher. Dust analysis may be useful method for estimating the mold burden in a home.
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| JOURNAL |
Characterization of Aeromonas Virulence Using An Immunocompromised Mouse Model |
03/01/2007
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LYE, D. J., M. R. RODGERS, G. N. STELMA, S. J. VESPER, AND S. L. HAYES. Characterization of Aeromonas Virulence Using An Immunocompromised Mouse Model. CURRENT MICROBIOLOGY. Springer, New York, NY, 54(3):195-198, (2007).
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An immunocompromised mouse model was used to characterize Aeromonas strains for their ability to cause opportunistic, extraintestinal infections. A total of 34 isolates of Aeromonas (A. hydrophila [n = 12]), A. veronii biotype sobria [n = 7], A. caviae [n = 4], A. enchelia [n = 4], A. allosaccharophila [n = 2], A. salmonicida (n = 4), and A. bestiarum [n = 1]) were introduced by intraperitoneal injection into immunocompetent or chemically compromised (using cyclophosphamide) mice. The ability of each isolate to persist in the liver and spleen tissue was monitored at 24 hours after exposure.Amajority of A. hydrophila and A veronii v. sobria strains, but none of the isolates of other Aeromonas species, were capable of persistent colonization (<300 cells/mg spleen and liver tissue at 24 hours). The presence or absence of several putative virulence factors (cytotoxicity to HEp-2, lipase activity, elastase activity, and hemolysis) were determined for each isolate using in vitro tests. There were no correlations between the presence or absence of biochemical test results for putative virulence factors and persistence of the isolate in spleen and liver tissue at 24 hours.
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| JOURNAL |
Comparison of Mold Concentrations in Indoor and Outdoor Air Sampled Simultaneously and Then Quantified By Msqpcr |
08/15/2007
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MEKLIN, T., T. REPONEN, C. MCKINSTRY, S. CHO, S. A. GRINSHPUN, A. NEVALAINEN, A. VEPSALAINEN, R. A. HAUGLAND, G. LEMASTERS, AND S. J. VESPER. Comparison of Mold Concentrations in Indoor and Outdoor Air Sampled Simultaneously and Then Quantified By Msqpcr. SCIENCE OF THE TOTAL ENVIRONMENT. Elsevier Science Ltd, New York, NY, 382(1):130-134, (2007).
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Mold specific quantitative PCR (MSQPCR) was used to measure the concentrations of the 36 mold species in indoor and outdoor air samples that were taken simultaneously for 48 hours in and around 17 homes in Cincinnati, Ohio. The total spore concentrations of 353 per m3 of indoor air and 827 per m3 of outdoor air samples were significantly different (p < 0.05). However, only the concentrations of Aspergillus penicillioides, Cladosporium cladosporioides types 1 and 2 and C. herbarum were correlated in indoor and outdoor air samples (p value < 0.05 and sufficient data for estimate and absolute value rho estimate > 0.5). These results suggest that interpretation of the meaning of short-term (< 48 h) mold measurements in indoor and outdoor air samples must be made with caution.
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| JOURNAL |
Photocatalytic Tio2 Films and Membranes for the Development of Efficient Wastewater Treatment and Reuse Systems |
01/05/2007
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CHOI, H., M. G. ANTONIOU, A. A. DELACRUZ, E. STATHATOS, AND D. D. DIONYSIOU. Photocatalytic Tio2 Films and Membranes for the Development of Efficient Wastewater Treatment and Reuse Systems. DESALINATION. Elsevier Science Ltd, New York, NY, 202(1-3):199-206, (2006).
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In order to develop efficient photocatalytic TiO2 films and membranes for application in water and wastewater treatment and reuse systems, there is a great need to tailor-design the structural properties of TiO2 material and enhance its photocatalytic activity. Through a simple sol-gel route, employing self-assembled surfactant molecules as pore directing agents along with acetic acid-based sol-gel route, we have fabricated nanostructured crystalline TiO2 thin films and TiO2/Al2O3 composite membranes with simultaneous photocatalytic, disinfection, separation, and anti-biofouling properties. The highly porous TiO2 material exhibited high specific surface area and porosity,narrow pore size distribution, homogeneity without cracks and pinholes, active anatase crystal phase, and small crystallite size. These TiO2 materials were highly efficient in the decomposition of methylene blue dye and creatinine, destruction of biological toxins (microcystin-LR), and inactivation of pathogenic microorganisms (Escherichia coli). Moreover, the photocatalytic TiO2 membranes exhibited not only high water permeability and sharp polyethylene glycol retention but also less adsorption fouling tendency. Here, we report results on the synthesis, characterization, and environmental application and implication of photocatalytic TiO2 films and membranes.
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| JOURNAL |
Speciation of Organotins in Poly Vinyl Pipe Via X-Ray Absorption Spectroscopy and in Leachates By Elthylation/Derivitization |
02/01/2007
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IMPELLITTERI, C., O. M. EVANS, AND B. RAVEL. Speciation of Organotins in Poly Vinyl Pipe Via X-Ray Absorption Spectroscopy and in Leachates By Elthylation/Derivitization. JOURNAL OF ENVIRONMENTAL MONITORING. Royal Society of Chemistry, Cambridge, Uk, 9(4):358-365, (2007).
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Three different polyvinyl chloride (PVC) pipe types were subjected to de-ionized water exposures over the course of at least 180 days. Water exposed to the pipe was analyzed for organotin speciation and concentration. Organotin concentrations were the highest during the first 1-5 days. The species and concentrations of organotins leached varied by pipe type. Data were normalized by surface area in order to compare laboratory results with results from a residential pipe system. For one pipe type, the lowest non-zero concentrations from the laboratory tests overestimated organotin concentrations in solution when compared with water samples from the same pipe type in a residence. For organotin exposure estimates, a range of 0.1 ng mµ 2 to 10 ng mµ 2 could be used for mature pipes (e.g. in use for 1 year). These estimates should be refined with more field study, however, due to the high variation in organotin species and concentrations leached as a function of pipe type, accuracy within an order of magnitude may be optimal as, in many instances, the type of pipe installed or buried may be unknown. X-ray absorption spectroscopy (XAS) was used to identify organic and inorganic tin species in reference materials and the PVC samples. Monobutyl tin was identified as the primary organotin species in the pipes. Results from the XAS analyses also indicate that the technique shows promise for distinguishing between inorganic tin and organotins. Furthermore, organotins may be distinguished between mono-, di-, and tri-ligand species using XAS.
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| JOURNAL |
Opportunistic Aspergillus Pathogens Measured in Home and Hospital Tap Water By Mold Specific Quantitative Pcr (Msqpcr) |
08/07/2007
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VESPER, S. J., M. E. ROGERS, A. N. NEELY, AND R. A. HAUGLAND. Opportunistic Aspergillus Pathogens Measured in Home and Hospital Tap Water By Mold Specific Quantitative Pcr (Msqpcr). Charles P. Gerber (ed.), JOURNAL OF WATER AND HEALTH. IWA Publishing, London, Uk, 5(3):427-431, (2007).
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| Abstract:
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Opportunistic fungal pathogens are a concern because of the increasing number of immunocompromised patients. The goal of this research was to test a simple extraction method and rapid quantitative PCR (QPCR) measurement of the occurrence of potential pathogens, Aspergillus fumigatus, A. flavus, A. terreus and A. niger, in home tap water and a hospital water supply.
Water samples were taken from the kitchen tap in homes of 60 patients who were diagnosed with legionellosis. Water samples were also taken from three locations in a hospital that generated all of its hot water by flash heating. Opportunistic infectious agents Aspergillus fumigatus, A. flavus, A. terreus and A. niger were measured using QPCR. Aspergillus terreus DNA was found in 16.7% and A. fumigatus DNA in 1.7% of the samples taken from the kitchen tap. None of the Aspergillus species were found in any of the hospital water samples.
The development of a simple DNA extraction method along with QPCR analysis is suitable for rapid screening of tap water for opportunistic fungal pathogens. This simple method can be used to obtain pathogen occurrence results in about 3 hours; instead of waiting days to weeks for culture data. Obtaining pathogen occurrence data in a timely manner, could promote the elimination of the pathogens from the water supply of immunocompromised patients.
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| JOURNAL |
Amplified Fragment Length Polymorphism Analysis of Mycobacterium Avium Complex Isolates Recovered from Southern California |
09/01/2007
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PFALLER, S. L., T. C. COVERT, T. A. ARONSON, AND A. H. HOLTZMAN. Amplified Fragment Length Polymorphism Analysis of Mycobacterium Avium Complex Isolates Recovered from Southern California. Journal of Medical Microbiology. Society for General Microbiology, 56(9):1152-1160, (2007).
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Fine-scale genotyping methods are necessary in order to identify possible sources of human exposure to opportunistic pathogens belonging to the Mycobacterium avium complex (MAC). In this study, amplified fragment length polymorphism (AFLP) analysis was evaluated for fingerprinting 159 patient and environmental MAC isolates from southern California. AFLP analysis accurately identified strains belonging to M. avium and Mycobacterium intracellulare and differentiated between strains within each species. The method was also able to differentiate strains that were presumed to be genetically identical in two previous studies using large RFLP analysis with PFGE, or PCR-amplification of DNA segments located between insertion sequences IS1245 and IS1311. For M. avium, drinking-water isolates clustered more closely with each other than with patient or food isolates. Patient isolates were more genetically diverse. None of the environmental isolates shared identical AFLP patterns with patient isolates for either species. There were, however, environmental isolates that shared identical patterns, and patient isolates that shared identical patterns. A subset of the isolates, which are referred to as MX isolates due to their ambiguous identification with the Gen-Probe system, produced AFLP patterns similar to those obtained from M. intracellulare isolates. Sequence analysis of 16S rDNA obtained from the MX isolates suggests that they are strains of M. intracellulare that were not correctly identified by the M. intracellulare AccuProbe from Gen-Probe.
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| JOURNAL |
Development of An Internal Control for Evaluation and Standardization of a Quantitative Pcr Assay for Detection of Helicobacter Pylori in Drinking Water |
11/01/2007
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SEN, K., N. A. Schable, AND D. J. LYE. Development of An Internal Control for Evaluation and Standardization of a Quantitative Pcr Assay for Detection of Helicobacter Pylori in Drinking Water. APPLIED AND ENVIRONMENTAL MICROBIOLOGY. American Society for Microbiology, Washington, DC, 73(22):7380-7387, (2007).
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Due to metabolic and morphological changes that can prevent Helicobacter pylori cells in water from growing on conventional media, an H. pylori-specific TaqMan quantitative PCR (qPCR) assay was developed that uses a 6-carboxyfluorescein-labeled probe (A. E. McDaniels, L. Wymer, C. Rankin, and R. Haugland, Water Res. 39:4808-4816, 2005). However, proper internal controls are needed to provide an accurate estimate of low numbers of H. pylori in drinking water. In this study, the 135-bp amplicon described by McDaniels et al. was modified at the probe binding region, using PCR mutagenesis. The fragment was incorporated into a single-copy plasmid to serve as a PCR-positive control and cloned into Escherichia coli to serve as a matrix spike. It was shown to have a detection limit of five copies, using a VIC dye-labeled probe. A DNA extraction kit was optimized that allowed sampling of an entire liter of water. Water samples spiked with the recombinant E. coli cells were shown to behave like H. pylori cells in the qPCR assay. The recombinant E. coli cells were optimized to be used at 10 cells/liter of water, where they were shown not to compete with 5 to 3,000 cells of H. pylori in a duplex qPCR assay. Four treated drinking water samples spiked with H. pylori (100 cells) demonstrated similar cycle threshold values if the chlorine disinfectant was first neutralized by sodium thiosulfate.
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| JOURNAL |
Identification of a New Hemolysin from Diarrheal Isolate Ssu of Aeromonas Hydrophilia |
10/01/2007
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Erova, T. E., J. Sha, A. J. Horneman, B. K. Khajanchi, A. A. Fadl, AND A. Chopra. Identification of a New Hemolysin from Diarrheal Isolate Ssu of Aeromonas Hydrophilia. FEMS MICROBIOLOGY LETTERS. Elsevier Science Ltd, New York, NY, 275(2):301-311, (2007).
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A clinical strain SSU of Aeromonas hydrophila produces a potent cytotoxic enterotoxin (Act) with cytotoxic, enterotoxic, and hemolytic activities. A new gene, which encoded a hemolysin of 439-amino acid residues with a molecular mass of 49 kDa, was identified. This hemolysin (HlyA) was detected based on the observation that the act gene minus mutant of A. hydrophila SSU still had residual hemolytic activity. The new hemolysin gene (hlyA) was cloned, sequenced, and overexpressed in Escherichia coli. The hlyA gene exhibited 96% identity with its homolog found in a recently annotated genome sequence of an environmental isolate, namely the type strain ATCC 7966 of A. hydrophila subspecies hydrophila. The hlyA gene did not exhibit any homology with other known hemolysins and aerolysin genes detected in Aeromonas isolates. However, this hemolysin exhibited significant homology with hemolysin of Vibrio vulnificus as well as with the cystathionine β synthase domain protein of Shewanella oneidensis. The HlyA protein was activated only after treatment with trypsin and the resulting hemolytic activity was not neutralizable with antibodies to Act. The presence of the hlyA gene in clinical and water Aeromonas isolates was investigated and DNA fingerprint analysis was performed to demonstrate its possible role in Aeromonas virulence.
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| JOURNAL |
Further Characterization of a Type III Secretion System (T3ss) and of a New Effector Protein from a Clinical Isolate of Aeromonas Hydrophila Part I |
10/01/2007
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Sha, J., S. F. Wang, J. C. Sierra, A. A. Fadl, T. E. Erova, S. M. Foltz, B. K. Khajanchi, A. Silver, J. Graf, C. H. Schein, AND A. K. Chopra. Further Characterization of a Type III Secretion System (T3ss) and of a New Effector Protein from a Clinical Isolate of Aeromonas Hydrophila Part I. Microbial Pathogenesis. Elsevier BV, AMSTERDAM, Netherlands, 43(4):127-146, (2007).
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A type III secretion system (T3SS)-associated cytotoxin, AexT, with ADP-ribosyltransferase activity and homology to Pseudomonas aeruginosa bifuncational toxins ExoT/S, was recently identified from a fish pathogen Aeromonas salmonicida. In this study, we reported the molecular characterization of an aexT-like toxin gene (designated as aexU) from a diarrheal isolate SSU of A. hydrophila. The aexU gene was 1539 bp in length and encoded a protein of 512 amino acid (aa) residues. The NH2-terminus of AexU (aa residues 1–231) exhibited a 67% homology with the NH2-terminus of AexT from A. salmonicida. Importantly, its COOH-terminus (aa residues 232–512) had no homology with any known functional proteins in the database; however, the full-length AexU retained ADP-ribosyltransferase activity. The expression and subsequent secretion of AexU was T3SS dependent, as inactivation of the ascV gene that codes for an inner-membrane component of the T3SS channel from the wild-type (WT) bacterium, blocked translocation of AexU in HT-29 human colonic epithelial cells. We provided evidence that inactivation of acrV and axsE genes (homologs of lcrV and exsE in Yersinia species and P. aeruginosa, respectively) from A. hydrophila SSU, altered expression and/or secretion of AexU. We deleted an aexU gene from the WT, as well as from the ΔaopB mutant, of A. hydrophila, generating a single knockout (ΔaexU) and a double knockout mutant, ΔaopB/ΔaexU. Increased phagocytosis was observed in RAW264.7 murine macrophages infected with the ΔaopB/ΔaexU mutant, as compared to macrophages when infected with the parental ΔaopB strain. Further, mice infected with the ΔaexU mutant had a 60% survival rate, compared to animals infected with the WT or the ΔaexU-complemented strain that caused 90–100% of the animals to die at a 2–3 LD50s dose. Immunization of mice with the recombinant AexU protected them from subsequent lethal challenge dose by the WT bacterium. Finally, we detected specific anti-AexU antibodies in the sera of mice that survived challenge by the WT bacterium, which may indicate that AexU plays an important role in the pathogenesis of Aeromonas infections.
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| PRESENTATION |
Sulfate Radical-Based Advanced Oxidation Processes |
03/25/2007
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DIONYSIOU, D. D., A. RASTOGI, M. G. ANTONIOU, G. P. ANIPSITAKIS, S. R. AL-ABED, A. A. DELACRUZ, AND J. A. SHOEMAKER. Sulfate Radical-Based Advanced Oxidation Processes. Presented at American Chemical Society National Meeting, Chicago, IL, March 25 - 29, 2007.
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This will be presented at the American Chemical Society National Meeting, Chicago, IL, March 25-29, 2007
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| PRESENTATION |
Ultraviolet-and Solar Light-Activated Nanostructured Tio2 Photocatalysts: Application in the Destruction of Cyanotoxins, a Group of Cyanotoxins Emerging Drinking Water Contaminants |
03/29/2007
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DIONYSIOU, D. D., M. G. ANTONIOU, H. CHOI, A. A. DELACRUZ, AND J. A. SHOEMAKER. Ultraviolet-and Solar Light-Activated Nanostructured Tio2 Photocatalysts: Application in the Destruction of Cyanotoxins, a Group of Cyanotoxins Emerging Drinking Water Contaminants. Presented at American Chemical Society National Meeting, Chicago, IL, March 25 - 29, 2007.
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To be presented at the American Chemical Society National Meeting in Chicago, IL, March 25-29, 2007
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| PRESENTATION |
Water Microbiology Methods: Membrane Filter Techniques |
01/09/2007
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BRENNER, K. P. Water Microbiology Methods: Membrane Filter Techniques. Presented at UPRM Water Microbiology Methods Course, Mayaguez, PR, January 09 - 12, 2007.
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To be presented at the UPRM Water Microbiology Methods Course, Mayaguez, Puerto Rico, January 9-12, 2007
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| PRESENTATION |
Collection of Water Samples |
01/09/2007
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BRENNER, K. P. Collection of Water Samples. Presented at UPRM Water Microbiology Methods Course, Mayaguez, PR, January 09 - 12, 2007.
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| Abstract:
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Powerpoint presentation to be presented at the UPRM Water Microbiology Methods Course, Mayaguez, Puerto Rico, January 9-12, 2007
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| PRESENTATION |
Recreational Water Quality and Swimming Associated Health Effects 7th Annual Water Monitoring Conference |
02/01/2007
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DUFOUR, A. P. Recreational Water Quality and Swimming Associated Health Effects 7th Annual Water Monitoring Conference. Presented at 7th Annual Water Monitoring Conference, Ames, IA, February 01 - 02, 2007.
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To be presented at the 7th Annual Water Monitoring Conference, Ames, Iowa, February 1-2, 2007
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| PRESENTATION |
Use of Data to Inform Characterization and Management in Addressing Biofilm Problems |
01/31/2007
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ASHBOLT, N. Use of Data to Inform Characterization and Management in Addressing Biofilm Problems. Presented at TCR Stakeholder Workshop, Washington, DC, January 31 - February 01, 2007.
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To be presented at the TCR Stakeholder Workshop, Washington, DC, January 31-February 1, 2007
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| PRESENTATION |
A Study of the Interconversion of Methylated Arsenic Oxides to Methylated Arsenic Sulfides in Solutions Containing Free Sulfide |
02/18/2007
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CONKLIN, S., P. A. CREED, AND J. T. CREED. A Study of the Interconversion of Methylated Arsenic Oxides to Methylated Arsenic Sulfides in Solutions Containing Free Sulfide. Presented at 2007 European Winter Conference on Plasma Spectrochemistry, Taormina, ITALY, February 18 - 23, 2007.
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Evidence suggests that thiolated arsenicals are urinary metabolites in both humans and rats. These thiolated species may be formed in the digestive system or as metabolites within the body. The role they may play in the overall toxicity of arsenic is an active area of research. This research effort would benefit from an improved understanding of how the oxide and thiolated forms of arsenic can interconvert based on matrix constituents.
Data will be presented demonstrating that trimethylarsine oxide (TMAO), dimethlyarsinic acid (DMA) and monomethylarsonic acid (MMA) all produce thiolated analogs in the presence of solution phase sulfide. This conversion is shown to be pH sensitive and the conversion rate is shown to increase in the following order: MMA
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| PRESENTATION |
Global Warming and Trans-Boundary Movement of Waterborne Microbial Pathogens |
02/26/2007
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ASHBOLT, N. Global Warming and Trans-Boundary Movement of Waterborne Microbial Pathogens. Presented at International Symposium on Dialogue between Social and Natural Sciences, Honolulu, HI, February 26 - 28, 2007.
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Subtle increases in temperatures can have profound impacts on the prevalence of various waterborne microbial pathogens. Such impacts may be seen in three major areas: 1) fecally-contaminated drinking waters; 2) fresh produce that has been irrigated or processed with contaminated water; and 3) seafood where pathogens and microbial toxins are present. Each of these areas is influenced by rainfall events, which have a fundamental influence on the fate and transport of pathogens.
Temperature alone can also impact, as seen with the epidemic strains of cholera (disease from certain Vibrio cholerae bacteria). In coastal environments, cholera outbreaks associate with particular phytoplankton blooms in nutrient-enriched coastal waters; with blooms varying due to changing precipitation regimes and El Niño. A further ecological interaction is seen by bacteriophage mediated cholera toxin genes inserted into non-toxic V. cholerae strains.
For microorganisms, boundaries occur at the tens of micron scale, with most terrestrial microbes living in slime on surfaces (biofilm). As ambient water temperatures increase, there is a clear increase in the growth of biofilms and their associated pathogens, many of which appear to be amoeba-associated. Most opportunistic bacterial pathogens (such as strains of Aeromonas, Legionella, Mycobacterium) and even frank, fecally-derived pathogens (e.g., Campylobacter jejuni, Cryptosporidium parvum, and enteric viruses) have been observed to accumulate within biofilm amoeba. With rising temperatures and increased application of recycled waters, biofilm pathogens will presenting a range of pathogen challenges to communities.
Human behavior is also influenced by climate change, with increased consumption of uncooked foods and water during warmer conditions, as well as increased travel (trans-boundary transport of pathogens). Increased rainfall events also promote zoonotic diseases (e.g., cryptosporidiosis, E. coli O157:H7 infections) via contamination of drinking water sources and waters used to irrigate crops.
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| PRESENTATION |
Comparison of the Urinary Metabolites of Rats, Mice, and Humans After Oral Arsenic Exposure Focusing on Thioarsenicals |
03/25/2007
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CONKLIN, S., B. ADAIR, P. A. CREED, J. T. CREED, M. F. HUGHES, AND D. J. THOMAS. Comparison of the Urinary Metabolites of Rats, Mice, and Humans After Oral Arsenic Exposure Focusing on Thioarsenicals. Presented at American Chemical Society Meeting, Chicago, IL, March 25 - 29, 2007.
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Urinary metabolites of arsenic are useful as biomarkers of exposure because ingested arsenic is excreted primarily in urine1. Complete urinary arsenic speciation can provide insight into possible metabolic pathways as well as potential exposure sources. The pattern of excreted metabolites depends upon which arsenicals are ingested. For example, both arsenosugars and inorganic arsenic generate dimethylarsinate (DMA) as a major urinary metabolite. For arsenosugars, this involves cleavage of As-C bonds, while for inorganic arsenic, As-C bonds are formed. In comparison, other arsenicals such as arsenobetaine and arsenocholine are excreted unchanged in urine. The recent finding of sulfur analogs of some arsenic oxides indicates that comprehensive urinary speciation has not yet been achieved. These sulfur analogs pose a unique analytical challenge because of the possible solution-phase interconversion between the oxygen and sulfur containing metabolites. The observed distribution between the two forms may be influenced by sample preparation and handling because of this equilibrium. It is with this in mind that a series of urine samples from rats, mice and humans have been analyzed specifically for the thiolated arsenicals dimethylthioarsinic acid (DMTA) and trimethylarsine sulfide (TMAS). The results are compiled here as an overview of the thio-arsenicals detected in urine (other arsenicals such as DMA and arsenate (As(V)) were detected, but are not included in this presentation).
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| PRESENTATION |
Understanding and Applying the Environmental Relative Moldiness Index (Ermi)SM in Dod Facilities |
08/07/2007
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VESPER, S. J. Understanding and Applying the Environmental Relative Moldiness Index (Ermi)SM in Dod Facilities. Presented at 2007 DoD 10th Annual Force Health Protection, Louisville, KY, August 07 - 10, 2007.
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Mold burdens in the indoor environment are a growing concern for the Department of Defense and other government agencies, as well as, the general public. Most recently mold in Walter Reed outpatient facilities became a significant issue. Yet there has been no standardized, objective method to quantify the mold burden indoors. This situation has now been corrected with the development of a DNA-based method of mold analysis called Mold Specific Quantitative Polymerase Chain Reaction (MSQPCR) (US Patent No.6,387,652). Using this technology to measure 36 indicator mold species in dust allowed for the development of a scale of relative mold burdens indoors called the Environmental Relative Moldiness Index (ERMI). This scale may be a useful tool to evaluate the mold levels in DOD facilities.
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| PRESENTATION |
Understanding and Applying Environmental Relative Moldiness Index Ermi |
04/03/2007
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VESPER, S. J. Understanding and Applying Environmental Relative Moldiness Index Ermi. Presented at National EPA-Tribal Science Council Meeting, Chicago, IL, April 03, 2007.
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This study compared two binary classification methods to evaluate the mold condition in 271 homes of infants, 144 of which later developed symptoms of respiratory illness. A method using on-site visual mold inspection was compared to another method using a quantitative index of moldiness, calculated from mold specific quantitative PCR (MSQPCR) measurements on the concentration of 36 species of molds in floor dust samples called the EPA relative moldiness index© (ERMI©).
The binary classification of homes as either moldy or non-moldy by on-site visual home inspection was not predictive of the development of respiratory illness (wheeze and/or rhinitis) (p = 0.27). Conversely, a method developed using the ERMI© index fit to a logistic function, can be used to predict the occurrence of illness in homes and allows stake-holders the choice among various levels of risk. An example is given where an ERMI© value of -4.29 is used as a threshold for binary classification of homes producing an odds ratio of 2.53 (95% confidence limits of 1.35, 4.77). The ERMI© based method provides a new and more flexible platform to support mold remediation decisions based on quantitative estimates using cost and risk tolerance.
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| PRESENTATION |
Internal Amplification Control for Use in Quantitative Polymerase Chain Reaction Fecal Indicator Bacteria Assays |
04/13/2007
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SIEFRING, S., E. ATIKOVIC, R. A. HAUGLAND, M. SIVAGANESAN, AND O. C. SHANKS. Internal Amplification Control for Use in Quantitative Polymerase Chain Reaction Fecal Indicator Bacteria Assays. Presented at Ohio Branch of the American Society for Microbiology Annual Meeting, Canton, OH, April 13 - 14, 2007.
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Quantitative polymerase chain reaction (QPCR) can be used as a rapid method for detecting fecal indicator bacteria. Because false negative results can be caused by PCR inhibitors that co-extract with the DNA samples, an internal amplification control (IAC) should be run with each sample. Currently available controls used in QPCR analyses for the fecal indicator bacterial groups Enterococcus and Bacteroidetes were designed primarily to determine variability in DNA yields from environmental samples and cannot be used to directly demonstrate PCR inhibition. Therefore, a competitive IAC plasmid DNA was constructed to detect the presence of PCR inhibitors in QPCR assays for both Enterococcus and Bacteroidetes rRNA gene targets. The IAC was designed to contain a single site for hybridization with a unique probe sequence that is flanked by multiple primer-hybridizing sites that corresponded to the same primers used in the Enterococcus, Bacteroidetes and several additional QPCR assays. The IAC construct was prepared by overlap extension PCR, inserted into the pCR4®TOPO plasmid vector (Invitrogen) and cloned. Gel electrophoresis, QPCR and sequencing analyses were performed to confirm the presence of the correct IAC sequences in the plasmid. Slope and intercept values of standard curves generated from genomic DNA in simplex analyses were not significantly different (p > 0.05) from the values generated during multiplex analyses with a fixed number of 25 IAC plasmid copies. Ranges of genomic DNA concentrations that did not significantly affect the IAC results under the same conditions were also established. Multiplex analyses with the IAC were used in a study of the relative levels of Enterococcus and Bacteroidetes DNA in fecal samples from cattle. In these analyses the Enterococcus IAC assay results showed highly consistent cycle threshold values (mean = 34.15, std. deviation = 0.69, N = 159) where only three results failed to occur within the 95% confidence interval established from analyses of control samples with IAC plasmid but no fecal extracts present. Greater variability in the Bacteroidetes IAC assay results was consistent with the relatively high levels of genomic DNA from these organisms in the samples. These studies indicate that the IAC plasmid DNA performs well as an inhibition control and also may be useful as an alternative to genomic DNA standards for quantifying fecal bacteria target DNA sequences.
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| PRESENTATION |
Estimating Pathogen Exposures the Critical Challenge for Qmra to Support Regulation and Management of Waters |
04/23/2007
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ASHBOLT, N. Estimating Pathogen Exposures the Critical Challenge for Qmra to Support Regulation and Management of Waters. Presented at Toxicology and Risk Assessment Conference, West Chester, OH, April 23 - 26, 2007.
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Pathogen and indicator concentrations normally vary by several orders of magnitude in raw waters, and to an even greater extent during hazardous event periods. This variation in concentration typically dominate the estimate of infection generated in a quantitative microbial risk assessment (QMRA), particularly if results are not averaged over a year. In addition to this variation, numerous uncertainties result from our attempts to assay pathogens and model their behaviour through water systems.
Raw water [oo]cyst concentrations are typically presented with little or no reporting of specific recovery data representative of the site sampled. The uncertainty resulting from limited recovery data was estimated for the MicroRisk Project systems using available data to improve the quality of Cryptosporidium and Giardia raw water concentration estimates. Recovery datasets ranging from 3-99 data points were examined by Bayesian statistics representing three Approaches: I - no recovery data, II - limited, unpaired recovery data from samples, and III - paired recovery data. No useful relationships were seen between water turbidity or type and recoveries reported. Critically, Approach I underestimated [oo]cyst concentrations by about 100%, with little difference between Approaches II & III. Using the smallest (n=3) recovery dataset, the upper band of uncertainty were on average more than 10-times (and on occasion up to 100-times) greater than when using the fullest (n=99) dataset; however, limited reduction in uncertainty occurred beyond n = 20. Nonetheless, for QMRA purposes, recovery data should be collected as a pair with count data for an initial period at least, so that any relationships (priors in Bayesian statistics) may be ascertained without the need to rely on and apply trends from elsewhere. When conveying QMRA results for risk management, output uncertainty should be incorporated into the results via either a two-dimensional (variability and uncertainty) risk assessment or a sensitivity analysis that includes recovery uncertainties.
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| PRESENTATION |
Cryptosporidium Source Tracking to Enhance Source Water Protection Implementation in the Potomac River Watershed: A Regional Applied Research Efforts (Rare) Project |
04/30/2007
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YANG, W., P. CHEN, R. B. LANDY, C. KANETSKY, E. VILLEGAS, AND L. XIAO. Cryptosporidium Source Tracking to Enhance Source Water Protection Implementation in the Potomac River Watershed: A Regional Applied Research Efforts (Rare) Project. Presented at U.S. EPA Region 5 Laboratory Technical Information Group Conference, Chicago, IL, April 30 - May 03, 2007.
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The Potomac River watershed is a critical drinking water supply for the Washington DC metropolitan area. In 2004, the Drinking Water Source Protection Partnership (DWSPP) was formed to help coordinate efforts by local drinking water utilities and government agencies to protect this watershed from various contaminants, including Cryptosporidium. This organism is a protozoan parasite that is excreted in feces by infected animals and humans. There are many different species of Cryptosporidium, but only two, C. parvum and C. hominis, are commonly associated with human infection. Although the current standard method used to detect Cryptosporidium in water is effective in enumerating oocysts, it cannot differentiate human from animal forms of Cryptosporidium. This limitation thus prevents the identification of specific Cryptosporidium species/genotypes that are contaminating environmental waters. Recently, a nested-PCR-based genotyping method has been developed and used to detect and identify specific Cryptosporidium species/genotypes in the watershed. Since Cryptosporidium is host specific, this technique has also been useful in tracking potential sources of Cryptosporidium contamination within the watershed. For this RARE project, a nested-PCR- method is used to detect and identify various Cryptosporidium genotypes that are present and to determine their likely sources in the Potomac River watershed. Results from this project will help DWSPP and local drinking water utilities evaluate current management practices that are aimed at minimizing Cryptosporidium contamination of their watershed.
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| PRESENTATION |
Analysis of Water Soluble Volatile Organic Chemicals in Drinking Water When Volatiles Aren't Purgeable |
05/10/2007
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MUNCH, J. W. AND P. GRIMMETT. Analysis of Water Soluble Volatile Organic Chemicals in Drinking Water When Volatiles Aren't Purgeable. Presented at 45th Annual Water Workshop, Columbus, OH, May 10, 2007.
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| Abstract:
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Slide and poster to be presented at the 45th Annual Water Workshop in Columbus, OH, May 10, 2007
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| PRESENTATION |
Urban Water System Pathogen Assessments: Significance of Distribution Biofilms |
05/04/2007
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ASHBOLT, N. Urban Water System Pathogen Assessments: Significance of Distribution Biofilms. Presented at Urban Water System Pathogen Risk Assessments: Signficance of distribution of biofilms, Cincinnati, OH, May 04, 2007.
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Quantitative microbial risk assessment (QMRA), while not new to science is now providing a fundamental role in framing water guidelines internationally as well as identifying research gaps to be filled. Professor Ashbolt has been instrumental in working QMRA concepts into WHO guidelines for drinking, recreational and reuse waters, as well as within the Swedish Urban Water program and a recently completed EU project (MicroRisk). The process steps in undertaking a QMRA will be discussed along with Bayesian statistical methods that are appropriate to handle small pathogen datasets from watersheds through water treatment. Finally, the role of biofilms in potentially enhancing pathogen risks (through pathogen accumulation and in some cases growth) will be discussed, for both drinking and recycled water applications.
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| PRESENTATION |
Urban Water System Pathogen Assessment: Significance of Distribution Biofilms |
05/10/2007
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ASHBOLT, N. Urban Water System Pathogen Assessment: Significance of Distribution Biofilms. Presented at Center for Disease Control Meeting, Atlanta, GA, May 10, 2007.
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| Abstract:
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Quantitative microbial risk assessment (QMRA), while not new to science is now providing a fundamental role in framing water guidelines internationally as well as identifying research gaps to be filled. Professor Ashbolt has been instrumental in working QMRA concepts into WHO guidelines for drinking, recreational and reuse waters, as well as within the Swedish Urban Water program and a recently completed EU project (MicroRisk). The process steps in undertaking a QMRA will be discussed along with Bayesian statistical methods that are appropriate to handle small pathogen datasets from watersheds through water treatment. Finally, the role of biofilms in potentially enhancing pathogen risks (through pathogen accumulation and in some cases growth) will be discussed, for both drinking and recycled water applications.
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| PRESENTATION |
A Faster Method of Measuring Recreational Water Quality |
05/14/2007
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HAUGLAND, R. A. A Faster Method of Measuring Recreational Water Quality. Presented at EPA Region 10 Bacteria Conference, Tacoma, WA, May 14 - 15, 2007.
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To be presented at the EPA Region 10 Bacteria Conference, Tacoma, Washington, May 14-15, 2007
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| PRESENTATION |
Sample Preparation for Metallomics Studies |
05/21/2007
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PAWLECKI-VONDERHEIDE, A. M. Sample Preparation for Metallomics Studies. Presented at CERMACS 2007, Covington, KY, May 21, 2007.
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To be presented at the CERMACS 2007 - Central Regional Meeting of the American Chemical Society, Northern Kentucky Convention Center, Covington, KY May 21, 2007
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| PRESENTATION |
Laboratory Analyses: Water and Environmental Samples |
06/01/2007
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FOUT, G. Laboratory Analyses: Water and Environmental Samples. Presented at Workshop for Improving the Recognition, Investigation, and Reporting of Waterborne Disease Outbreaks Associated with Drinking, Recreational and Other Waters, Nashville, TN, May 27 - June 01, 2007.
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| Abstract:
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To be presented at the Workshop for Improving the Recognition, Investigation, and Reporting of Waterborne Disease Outbreaks Associated with Drinking, Recreational and Other Waters in Nashville, TN, May 29 - June 1, 2007
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| PRESENTATION |
Presentations at California Water Environment Association Specialty Conference on Chemicals of Emerging Concern |
05/22/2007
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GLASSMEYER, S., E. T. FURLONG, AND D. KOLPIN. Presentations at California Water Environment Association Specialty Conference on Chemicals of Emerging Concern. Presented at California Water Environment Association Specialty Conference on Chemicals of Emerging Concern, Van Nuys, CA, May 22 - 24, 2007.
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| Abstract:
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1. Introduction to Emerging Contaminants
2. Why are emerging Contaminants Important?
3. Regulatory Response to Emerging Contaminants
4. Identifying Chemical Compounds from Wastewater Discharge
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| PRESENTATION |
Temporal Variability of Microbial Indicators of Fecal Contamination of Marine and Freshwater Beaches |
05/21/2007
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WYMER, L. J., A. P. DUFOUR, AND C. MCGEE. Temporal Variability of Microbial Indicators of Fecal Contamination of Marine and Freshwater Beaches. Presented at American Society for Microbiology General 107th Meeting, Toronto, BC, CANADA, May 21 - 25, 2007.
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| Abstract:
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Monitoring methods for microbial indicators of fecal contamination are an integral component for protecting the health of swimmers exposed to potentially contaminated bathing beach waters. The design of monitoring systems which will accurately characterize the quality of water is dependent on knowledge of the variability associated with water sampling. This study was conducted to determine the
temporal variability encountered at bathing beaches. High frequency sampling was conducted at marine and freshwater beaches over a sixty day period. Multiple samples were taken at minute, 10 minute, hourly and daily time intervals. Water samples were assayed for E. coli or enterococci using membrane filter methods. In general, as the time interval between samples increased so did their variability. Based on the average freshwater and marine results, samples taken within seconds of each other had a coefficient of variation (CV) of 0.7. Samples taken at 10 minute intervals had a CV of 1.1. Samples taken at hourly intervals had a CV 1.7, and samples taken daily had a CV of 2.2. The results of this study show that single samples do not adequately characterize the quality of beach waters and that temporal variability must be given serious consideration when developing sampling plans for beach waters.
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| PRESENTATION |
Examination of the Protein Profile of Helicobacter Pylori Under Different Growth Conditions Using Matrix-Assisted Laser Desorption Mass Spectrometry |
05/21/2007
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STELMA, G. N., D. J. FLANIGAN, L. A. BOCZEK, D. J. LYE, AND M. J. DONOHUE. Examination of the Protein Profile of Helicobacter Pylori Under Different Growth Conditions Using Matrix-Assisted Laser Desorption Mass Spectrometry. Presented at American Society for Microbiology 107th Meeting, Toronto, ON, CANADA, May 21 - 25, 2007.
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| Abstract:
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US EPA currently has H. pylori on its Contaminant Candidate List 2 (CCL 2), methods are needed to detect the occurrence of viable H. pylori in drinking water. H. pyloi is an interesting microorganism because it can change from a cultural and metabolically active state with a helical morphology to a non-culturable, dormant state with a coccoid morphology. There is currently no culture medium for isolating H. pyloi from drinking water. Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry
(MALDI-MS) was used to look at protein profiles of H. pyloi over the course of 10 days. After four days on blood agar, more than 50% of the H. pyloi cells were coccoid and non-culturable and a noticeable shift of protein expression was observed. This altered protein profile was continuously observed up to day 10, at which time all culturability was lost but ATP was still present. Efforts are currently focused on identifying the H. pyloi proteins observed between day 4 and day 10. We hope to use these proteins as possible markers of viability and develop more sensitive detection methods using these markers as targets.
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| PRESENTATION |
Real-Time Quantitative Pcr Detection of Mycobacterium Avium Complex Organisms in Drinking Water |
05/21/2007
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KING, D. N., A. BEUMER, AND S. L. PFALLER. Real-Time Quantitative Pcr Detection of Mycobacterium Avium Complex Organisms in Drinking Water. Presented at American Society for Microbiology 107th General Meeting, Toronto, ON, CANADA, May 21 - 25, 2007.
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| Abstract:
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The Mycobacterium avium Complex (MAC) includes the species M. avium (MA), M. intracellulare (MI), and others. MAC are listed on the U.S. Environmental Protection Agency's Contaminant Candidate List (CCL) due to their association with human disease and occurrence in public drinking water systems. Current methods for detecting MAC organisms in drinking water are culture-based. However evidence suggests that culture-based methods have severe limitations including long incubation periods, loss of target due to overgrowth of background organisms, up to 70% loss of target due to harsh decontamination techniques, and inability to recover MAC in a viable-but-non-culturable state. Because of these drawbacks and the need for more accurate and comprehensive occurrence data, we have developed real-time QPCR assays for the detection and quantification of MA, MI, and M. avium subspecies paratuberculosis (MAP) in drinking water. Real-time QPCR assays were developed using primers and TaqMan® probes designed to amplify a region of the 16S rDNA in MA and MI, and regions of IS900 and Target 251 in MAP. Primer/probe sets were found to be highly specific when compared to
sequences in nucleotide databases and confirmed experimentally by screening 104 MAC strains. No false negatives occurred when each species was tested with its own primer/probe set, 2.3% (1/42) MA strains were false positive with the MI prober/probe set, and 2.5% (1/40) MI strains were false positive with the MA primer/probe set. No false positives were obtained when nine non-MAC species were screened with all primer/probe sets. Quantification is linear over a minimum range of six logs of target
concentration in all four assays. Additionally, a control has been developed to measure PCR inhibition due to compounds in the water matrix. We are currently evaluating the QPCR assays for use on actual drinking water samples as a rapid alternative to culture methods in order to generate a more complete understanding of MAC occurrence in drinking water.
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| PRESENTATION |
Efficient Recovery of Enterococci from Marine and Fresh Water Beaches By a 30,000 Molecular Weight Cutoff Hollow Fiber Ultrafilter |
05/21/2007
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MCDANIELS, A. E., C. C. RANKIN, L. J. WYMER, A. P. DUFOUR, AND K. OSHIMA. Efficient Recovery of Enterococci from Marine and Fresh Water Beaches By a 30,000 Molecular Weight Cutoff Hollow Fiber Ultrafilter. Presented at American Society for Microbiology General 107th Meeing, Toronto, ON, CANADA, May 21 - 25, 2007.
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| Abstract:
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Ultrafiltration systems have been used to concentrate pathogens from various types of fresh water samples. However, less work has been done with marine waters for the concentration of pathogens or indicator bacteria. An ultrafiltration approach to concentrate indicator bacteria such as Enterococci may be advantageous because of the ability to smaple larger volumes and concentrate multiple target organisms simultaneously. Each ten liter sample was seeded with approximately 1 x 104 cells. A peristaltic pump was used to force water through a 30,000 molecular weight cutoff hollow fiber ultrafilter (Fresenius). During the filtration process, Enterococci remained in the retentate while the filtered water was collected in the permeate. Cells were eluted from the filter by forward flushing with 0.1% Tween 80 and 0.05M glycine in PBS at 7.2 pH, followed by reverse flushing and filter shaking. Nitrogen gas was used to remove all remaining fluid. Ten liters were reduced to approximately 500 mL at a rate of l liter per minute. Aliquots from seeded initial and final concentrates were filtered, the filtgers placed on mEI agar for growth and colony counts used to calculate filter mL at a rate of 1 liter per minute. Aliquots from seeded initial and final concentrates were filtered, the filters placed on mEI agar for grown and colony counts used to calculate recoveries. Recoveries from seeded marine waters averaged 93% (n=17) from the Pacific and 64.7 (n=13) from the Atlantic Oceans, which was a significant difference (p-value=0.0185). Seeded fresh water recoveries averaged 81% (n=12). The physical-chemcial parameters of pH, turbidity, specific conductivity and salinity were within their expected ranges. In conclusion, the ultrafiltration process produced high recoveries of Enterococci from large volumes of beach waters within a reasonable time frame, was simple to operate, used inexpensive disposable filters and can be operated in the field as well as in the laboratory.
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| PRESENTATION |
Dietary Arsenic Exposure Assessment Using Enzymatic Based Extraction Conditions and Detection of Urinary Thio-Arsenicals as Metabolites of Exposure Mceard2 |
05/21/2007
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CREED, J. T., P. A. CREED, J. ELLIS, S. CONKLIN, C. GALLAWA, A. YOUNG, C. A. SCHWEGEL, AND J. CARUSO. Dietary Arsenic Exposure Assessment Using Enzymatic Based Extraction Conditions and Detection of Urinary Thio-Arsenicals as Metabolites of Exposure Mceard2. Presented at Central Regional Meeting of the American Chemical Society (CERMACS), Covington, KY, May 21, 2007.
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| Abstract:
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Inorganic arsenic is classified as a carcinogen and has been linked to lung and bladder cancer as well as other non-cancerous health effects. Because of these health effects the U.S. EPA has set a Maximum Contaminant Level (MCL) at 10ppb based on a linear extrapolation of risk and a Health Risk Reduction and Cost Analysis (HRRCA). A re-evaluation of the HRRCA will require the Agency to review scientific developments regarding arsenic's Mode of Action, treatment options and the aggregate exposure assessment. The scientific basis for any drinking water MCL is reviewed every six years.
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| PRESENTATION |
Children's Exposure to Pesticides: Unconventional Routes of Introduction |
05/18/2007
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PAWLECKI-VONDERHEIDE, A. M., L. J. MELNYK, J. N. MORGAN, T. E. HIEBER, AND P. KAUFFMAN. Children's Exposure to Pesticides: Unconventional Routes of Introduction. Presented at Seminar for the University of Cincinnati, College of Engineering, Cincinnati, OH, May 18, 2007.
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| Abstract:
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To be presented at Seminar for the University of Cincinnati, College of Engineering, May 18, 2007
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| PRESENTATION |
Decision-Making and Needs for Support: Protecting Groundwater Supplies from Pathogen Health Risks International Perspective |
06/12/2007
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ASHBOLT, N. Decision-Making and Needs for Support: Protecting Groundwater Supplies from Pathogen Health Risks International Perspective. Presented at Pathogen-In-Ground Research Consortium - Kick Off Workshop - Canadian Water Network, Toronto, ON, CANADA, June 12 - 13, 2007.
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| Abstract:
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To be presented at the Pathogen-In-Ground Research Consortium - Kick Off Workshop - Canadian Water Network, Toronto, Canada, June 12-13, 2007
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| PRESENTATION |
Development of An EPA Method for Perfluorocalkyl Compounds in Drinking Water |
06/25/2007
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SHOEMAKER, J. A. Development of An EPA Method for Perfluorocalkyl Compounds in Drinking Water. Presented at 2007 American Water Resources Association, Vail, CO, June 25 - 27, 2007.
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Perfluoroalkyl compounds (PFCs) have been manufactured for over 50 years and their use has dramatically increased over the years. Due to their unique properties of repelling both water and oil, PFCs have been used in a wide variety of applications. In 2001, identification of organic fluorine as PFCs was confirmed in human sera through the use of liquid chromatography/mass spectrometry (LC/MS). These compounds received world-wide attention when the presence of perfluorooctane sulfonate and perfluorooctanoic acid were reported in blood and liver samples of animals in urban and remote locations.
Of particular interest is the global detection of PFCs in ground and surface waters as these can be potential drinking water sources. The 1996 Amendments to the Safe Drinking Water Act required the Environmental Protection Agency (EPA) to establish a Drinking Water Contaminant Candidate List (CCL) that contains a list of drinking water contaminants that the Agency will consider for future regulation. Because of the recent interest in PFCs, it is likely that they will be listed on a future CCL. One of the key pieces of information necessary to make a regulatory determination is nationwide occurrence data for the chemical contaminants under consideration. Historically, EPA's Office of Ground Water and Drinking Water has collected the necessary occurrence data under its Unregulated Contaminant Monitoring Regulations. To gather occurrence data, a rugged analytical method is needed for these PFCs.
While several methods have been reported, these methods do not adequately address issues specific to analyzing PFCs in drinking water. Issues, such as preservatives, internal and surrogate standards, and establishing acceptable background levels, will be studied in this method development effort. The target analyte list includes the C6 through C14 perfluorinated carboxylic and sulfonic acids, as well as the perfluorooctanesulfonamidoacetates. Drinking water samples are concentrated by solid phase extraction using styrene-divinyl benzene sorbents and analyzed using LC/tandem mass spectrometry (LC/MS/MS). Recovery and precision data for the target PFCs will be presented using various drinking water matrices. Detection limits below 10 ng/L will be demonstrated.
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| PRESENTATION |
Development and Evaluation of a Microarray Approach to Detect and Genotype Noroviruses in Water |
06/18/2007
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BRINKMAN, N. Development and Evaluation of a Microarray Approach to Detect and Genotype Noroviruses in Water. Presented at The U.S. EPA Workshop on Innovative Approach to Detecting Microorganisms in Water, Cincinnati, OH, June 18 - 20, 2007.
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| Abstract:
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Noroviruses are the leading cause of nonbacterial gastroenteritis outbreaks in the United States, some of which are caused by the ingestion of contaminated water. These viruses are usually detected and genotyped using reverse transcription-polymerase chain reaction (RT-PCR) based methods followed by sequencing. Unfortunately, the accurate detection of noroviruses in environmental samples is often hindered by the co-amplification of non-specific DNA, which can result in the need for further purification of PCR products before accurate sequence information can be obtained. As an alternative to direct sequencing, a generic microarray was evaluated for its ability to genotype norovirus RT-PCR products by probe hybridization. With this approach, RT-PCR amplicons were first mixed with a range of genotype specific probes and single base extension (SBE) reactions were run. This resulted in the labeling of those probes that have sequences complementary to specific RT-PCR products. These genotype-specific probes were then hybridized to an Affymetrix GenFlex Tag Array for detection. Using a standardized, multiplex SBE reaction, the genotyping of representative strains was accomplished and resulted in the generation of specific hybridization patterns, or fingerprints, on the microarray that were diagnostic for the genotype of norovirus detected. Furthermore, the SBE-GenFlex array method was shown to be successful in the genotype identification of noroviruses seeded into tap and Ohio River water samples. This study demonstrates the utility of using a microarray to genotype noroviruses in complex environmental matrices.
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| PRESENTATION |
An Overview of Pathogen Research in the Microbiological and Chemical Exposure Assessment Research Division |
06/18/2007
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GRIMM, A. An Overview of Pathogen Research in the Microbiological and Chemical Exposure Assessment Research Division. Presented at USEPA Workshop on Innovative Approaches for Detecting Microorganisms in Water, Cincinnati, OH, June 18 - 20, 2007.
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| Abstract:
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The Microbiological and Chemical Exposure Assessment Research Division of the EPA Office of Research and Development's National Exposure Research Laboratory has a robust in-house research program aimed at developing better occurrence and exposure methods for waterborne pathogens. In particular, research is aimed toward developing and improving occurrence methods so that they are more rapid, sensitive and inexpensive. To conduct this research, a diverse array of detection technologies is used, including real-time PCR, microarrays, proteomics, and cell culture, among others. In addition, the division has more recently invested in evaluating new approaches to collecting and concentrating samples more effectively. The long-term goal of this work is to enable the Agency to conduct better, more accurate risk assessments that will aid regulatory decision-making.
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| PRESENTATION |
Global Water Research Coalition |
06/25/2007
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GLASSMEYER, S., F. S. HAUCHMAN, AND D. TILLMAN. Global Water Research Coalition. Presented at 2007 AWRA Summer Specialty Conference, Vail, CO, June 25 - 27, 2007.
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The Global Water Research Coalition (GWRC) is a collaboration of 14 member drinking and wastewater research organizations. The USEPA is currently a partner to the GWRC membership. Through the GWRC, the members are able to leverage research funds on mutually desired efforts to maximize outcomes and minimize duplication of efforts. Currently, these research efforts cover topics such as algal toxins, endocrine disrupting chemicals, pharmaceuticals and personal care products, nitrosodimethylamine, water quality in distribution systems, asset management and water reuse.
Issues that cannot be adequately addressed by individual organizations
Exchange of knowledge and expertise needed
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| PRESENTATION |
Dna Based Method of Mold and Applying the Environmental Relative Moldiness Index (Ermi) in Dod Facilities |
08/07/2007
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VESPER, S. J. Dna Based Method of Mold and Applying the Environmental Relative Moldiness Index (Ermi) in Dod Facilities. Presented at DoD Seminar - Force Health Protection Conference, Louisville, KY, August 07, 2007.
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| Abstract:
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To be presented at the Department of Defense - Force Health Protection Conference in Louisville, KY, August 7, 2007
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| PRESENTATION |
Emerging Contaminants in the Drinking Water Cycle |
06/25/2007
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GLASSMEYER, S., P. E. STACKELBERG, E. T. FURLONG, AND D. W. KOLPIN. Emerging Contaminants in the Drinking Water Cycle. Presented at 2007 AWRA Summer Specialty Conference, Vail, CO, June 25 - 27, 2007.
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| Abstract:
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PRESENTATION OUTLINE: I. General overview of the water cycle;
II. USEPA and USGS Research;
a. Wastewater treatment plant (WWTP) effluents and downstream surface waters;
b. Groundwater down gradient from WW lagoon;
c. Source and finished water from a drinking water treatment plant (DWTP);
d. Chlorination laboratory studies;
III. Future directions
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| PRESENTATION |
Dna Based Method of Mold and Applying the Environmental Relative Moldiness Index (Ermi) |
07/12/2007
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VESPER, S. J. Dna Based Method of Mold and Applying the Environmental Relative Moldiness Index (Ermi). Presented at NASA Seminar, Cincinnati, OH, July 12, 2007.
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NASA facilities can potentially have mold contamination problems. The EPA has created an Environmental Relative Moldiness Index based on the analysis of dust by Mold Specific Quantitative PCR (MSQPCR). In this presentation, the scientific background for the ERMI will be presented. The goal of the ERMI is to be able to provide a simple, comparative value that will define the mold burden in an indoor environment.
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| PRESENTATION |
Emerging Contaminants in the Water Cycle: Fate and Transport |
07/16/2007
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GLASSMEYER, S. Emerging Contaminants in the Water Cycle: Fate and Transport. Presented at USEPA Workshop on Building an Integrated Surveillance System for Emerging Chemicals in the Great Lakes and Nationwide, Chicago, IL, July 16 - 18, 2007.
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In the past decade, the scientific community and general public have become increasingly aware of the potential for the presence of unregulated, and generally unmonitored contaminants, found at low concentrations in surface, ground and drinking water. The most common pathway for the introduction of these chemicals is from an upstream direct discharge of wastewater effluent. In the US, there are more than two dozen communities that draw their drinking water from streams that consist of more than 50 % wastewater during low flow conditions. The US Geological Survey (USGS) and US Environmental Protection Agency (USEPA) have been working on a series of collaborative research projects to determine the complex mixtures of chemicals that are commonly present in wastewater effluent, the persistence of these chemicals in surface and ground waters, the removal of these chemicals during drinking water treatment, the formation of by-products during chlorination and the presence of these chemicals in finished drinking water. In effluents collected at eleven wastewater treatment plants (WWTPs) across the US, 72 out of 110 monitored chemicals were detected at least once, documenting incomplete removal during wastewater treatment. Downstream of the WWTPs, the chemicals exhibited varying environmental persistence. In the source water of one conventional drinking water facility, 45 out of 113 monitored chemicals were detected at least once, with 21 chemicals still detectable in the finished drinking water. This documents the incomplete removal of such chemicals during treatment. In companion laboratory studies on the effects of chlorination, eight of the 14 chemicals investigated were oxidized by the disinfectant, two of which were at least partially chlorinated. Taken as a whole, these studies demonstrate that to understand the comprehensive environmental impact of emerging contaminants, their persistence, removal efficiencies during waste and drinking water treatment, as well as the potential for by-product formation, must be known.
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| PRESENTATION |
Use of a Molecularly Imprinted Polymer in the Determination of Pyrethroid Pesticides in Composite Foods |
07/22/2007
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MORGAN, J. N., T. E. HIEBER, A. M. PAWLECKI-VONDERHEIDE, P. KAUFFMAN, L. J. MELNYK, B. BOYD, A. RYBERG, AND E. YLMAZ. Use of a Molecularly Imprinted Polymer in the Determination of Pyrethroid Pesticides in Composite Foods. Presented at 2007 Florida Pesticide Residue Workshop, St. Pete Beach, FL, July 22 - 25, 2007.
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| Abstract:
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The U.S. Environmental Protection Agency's (EPA) National Exposure Research Laboratory (NERL) measures the exposure of individuals to chemical pollutants through the diet, as well as other media. In support of this research, methods are being evaluated for determination of various classes of pesticides in composite diet samples. Existing methods for pesticides generally have been developed for regulatory purposes and often do not have sufficiently low detection limits for exposure studies. Consequently, there is a need to improve the performance of these methods for analysis of pesticides in composite dietary samples. The objective of this work is to evaluate the applicability of a molecularly imprinted polymer (MIP) based solid phase extraction material for use in the analysis of pyrethroid pesticides in composite dietary samples collected in exposure studies.
A MIP-based solid phase extraction material, specific for pyrethroid pesticides, was developed by MIP Technologies AB, under contract to the EPA. This material was evaluated in combination with a variety of extraction and clean-up techniques in an effort to optimize its performance in the determination of pyrethroid pesticides in composite foods. This presentation will outline the various experiments conducted until acceptable performance was achieved.
The final method consisted of a modified QuEChERS1 procedure followed by a clean-up step using the MIP. Detection was performed using GC/µECD. The procedure was evaluated for cis and trans-permethrin, cypermethrin and cyfluthrin in high, medium and low fat (10%, 5% and 2%, respectively) food samples. Recoveries ranged from 83-126%. Detection limits ranged from 7 to 35 ppb using GC/ECD. This work was designed primarily to evaluate performance of the MIP. Future work will focus on ways to improve detection limits to sub-ppb levels.
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| PRESENTATION |
U.S. Environmental Protection Agency's Regulation and Management of Waterborne Viruses |
08/27/2007
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GRIMM, A. U.S. Environmental Protection Agency's Regulation and Management of Waterborne Viruses. Presented at International Symposium on Waterworks Technology, Seoul, SOUTH KOREA, August 27 - 28, 2007.
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The U.S. Environmental Protection Agency (USEPA) manages waterborne viruses and other pathogens through the establishment of rules and regulations that are designed to ensure public health protection. The rules that currently regulate pathogens focus on the management of viruses, Cryptosporidium, Giardia, Legionella and coliforms, and use a variety of approaches to reducing the level of these organisms in water. In addition, a Contaminant Candidate List (CCL) is maintained by the Agency as a way of identifying unregulated pathogens for which more data is needed before a regulatory determination can be made. Gathering more data on both regulated and unregulated pathogens is a major driver of the microbial research at the USEPA and each of the Laboratories and Centers that make up the Office of Research and Development (ORD) contribute to this effort by conducting complementary research that is organized according to the risk assessment paradigm.
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| PRESENTATION |
Microarray for Detection of Waterborne Pathogens |
09/05/2007
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VILLEGAS, E. AND N. BRINKMAN. Microarray for Detection of Waterborne Pathogens. Presented at 2007 Clean Water Partnership Summit, Cincinnati, OH, September 05 - 06, 2007.
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| Abstract:
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To be presented at the 2007 Clean Water Partnership. September 5-6, 2007 at the USEPA, Cincinnati, OH
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| PRESENTATION |
EPA's Expertise in Microbiological and Chemical Detection |
09/05/2007
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DUFOUR, A. P. EPA's Expertise in Microbiological and Chemical Detection. Presented at 2007 Clean Water Partnership Summit, Cincinnati, OH, September 05 - 06, 2007.
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To be presented at the 2007 Clean Water Partnership Summit held September 5-6, 2007 in Cincinnati, OH. The Challenge -Maintaining and Protecting Water Quality. Contaminants come from a variety of natural and man-made sources. Annually, 300 million tons of synthetic compounds derived from industrial and consumer products enter water systems.
Worldwide, over 2 million deaths each year can be attributed to biological contamination of water. In the wake of 9/11, there is heightened concern over the security of our nation's water infrastructure.
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| PRESENTATION |
Real-Time Microbial Ensor for Recreational Water |
09/05/2007
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DUFOUR, A. P. Real-Time Microbial Ensor for Recreational Water. Presented at 2007 Clean Water Summit, Cincinnati, OH, September 05, 2007.
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| Abstract:
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Presented at the 2007 Clean Water Summit, Cincinnati, OH. September 5-6,2007
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| PRESENTATION |
Quantitative Pcr Analysis of Waterborne Legionella Species |
09/05/2007
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VESPER, S. J. AND R. A. HAUGLAND. Quantitative Pcr Analysis of Waterborne Legionella Species. Presented at 2007 Clean Water Summit, Cincinnati, OH, September 05 - 06, 2007.
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| Abstract:
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Presented at the 2007 Clean Water Summit held in Cincinnati, OH, September 5-6, 2007
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| PRESENTATION |
Cryptosporidium Research Projects Within the Biohazard Assessment Research Branch |
09/13/2007
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VILLEGAS, E. Cryptosporidium Research Projects Within the Biohazard Assessment Research Branch. Presented at Workshop on Cryptosporidium Research Associated with Monitoring and Analytical Efforts in the Mid-Atlantic, Philadelphia, PA, September 13, 2007.
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| Abstract:
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Presented at the Workshop on Cryptosporidium Research Associated with Monitoring and Analytical Efforts in the Mid-Atlantic, September 13, 2007
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| PRESENTATION |
Biofilm-Like Water Sampling Device |
09/05/2007
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ASHBOLT, N. Biofilm-Like Water Sampling Device. Presented at 2007 Clean Water Summit, Cincinnati, OH, September 05 - 06, 2007.
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| Abstract:
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Presented at the 2007 Clean Water Summit, Cincinnati,OH, September 05-06, 2007
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| PRESENTATION |
Mi Medium |
09/18/2007
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BRENNER, K. P. Mi Medium. Presented at Microbiology Drinking Water Certification Training Course, Cincinnati, OH, September 18, 2007.
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| Abstract:
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To be presented at the Microbiology Drinking Water Certification Training Course, Cincinnati, OH, September 18, 2007
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| PRESENTATION |
Membrane Filter Methods |
09/18/2007
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BRENNER, K. P. Membrane Filter Methods. Presented at Microbiology Drinking Water Certification Training Course, Cincinnati, OH, September 18, 2007.
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| Abstract:
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To be presented at the Microbiology Drinking Water Certification Training
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| PRESENTATION |
Enteroccocci and Fecal Streptococci Membrane Filter Methods |
09/19/2007
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BRENNER, K. P. Enteroccocci and Fecal Streptococci Membrane Filter Methods. Presented at Microbiological Drinking Water Certification Training Course, Cincinnati, OH, September 19, 2007.
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| Abstract:
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To be presented at the Microbiology Drinking Water Certification Training Course, Cincinnati, OH, September 19, 2007
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| PRESENTATION |
Statistical Framework for Recreational Water Quality Criteria and Monitoring |
10/03/2007
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WYMER, L. J. Statistical Framework for Recreational Water Quality Criteria and Monitoring. Presented at 7th Annual Great Lakes Beach Association Meeting, Cincinnati, OH, October 03 - 05, 2007.
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| Abstract:
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Discussion between the EPA Office of Research and Development (ORD) and the EPA Office of Water (OW), which is charged with setting criteria in accordance with the BEACH Act of 2000, have made it clear that in-depth statistical guidance for such criteria is needed. In January 2004, a workshop was conducted at ORD's research center in Cincinnati, Ohio, which attempted to address many of the statistical concerns expressed by OW and to develop an outline for a book on the subject. The demand for such a book was evident given that OW's questions were not unique, but were in fact being asked by all involved in assessing recreational water quality. Consequently, the book is aimed at public health officials, government regulators and the water microbiology science community in general. Contributors consist of preeminent leaders in environmental statistics and recreational water modeling. Individual chapters were written by those with expertise in the respective subject matter. European perspectives are included, along with those of the United States, Canada, and our colleagues "down under." Chapters are arranged in a logical progression covering the history of recreational water quality management, present-day management perspectives, rationale for the use of indicators, statistical process control and quality control concepts, sampling designs, uses of arithmetic and geometric means, case study in sampling, risk assessment modeling, concentration-response modeling, modeling and forecasting techniques as adjuncts to monitoring, and sensitivity analysis.
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| PRESENTATION |
Statistical Framework for Recreational Water Quality Criteria and Monitoring, a New Title in the John Wiley & Sons Series, "STATISTICS in Practice" |
10/03/2007
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WYMER, L. J. Statistical Framework for Recreational Water Quality Criteria and Monitoring, a New Title in the John Wiley & Sons Series, "STATISTICS in Practice". Presented at 7th Annual Great Lakes Beach Association Meeting, Traverse City, MI, October 03 - 05, 2007.
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| Abstract:
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To be presented at the 7th Annnual Great Lakes Beach Association Meeting, October 3-5, 2007
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| PRESENTATION |
Community Duplicate Diet Methodology |
10/14/2007
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MELNYK, L. J., C. Clarke, S. McNutt, AND J. J. QUACKENBOSS. Community Duplicate Diet Methodology. Presented at ISEA Annual Meeting, Durham, NC, October 14 - 18, 2007.
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| Abstract:
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To be presented at the ISEA Annual Meeting in Durham, NC, October 14-18, 2007
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| PRESENTATION |
Investigation of Reagent Gases for the Positive Chemical Ionization of the Polybrominated Diphenyl Ethers |
10/18/2007
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PAWLECKI-VONDERHEIDE, A. M., T. E. HIEBER, P. KAUFFMAN, J. N. MORGAN, AND L. J. MELNYK. Investigation of Reagent Gases for the Positive Chemical Ionization of the Polybrominated Diphenyl Ethers. Presented at 2007 Federation of Analytical Chemistry & Spectroscopy Societies (FACSS) Conference, Memphis, TN, October 14 - 18, 2007.
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To be presented at the 2007 Fedration of Analytical Chemistry and Spectroscopy Societies (FACSS) Conference, October 14-18, 2007, Memphis, TN
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| PRESENTATION |
Surface-to-Food Pesticide Transfer as a Function of Fat and Moisture Content |
10/14/2007
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PAWLECKI-VONDERHEIDE, A. M., T. E. HIEBER, P. KAUFFMAN, J. N. MORGAN, AND L. J. MELNYK. Surface-to-Food Pesticide Transfer as a Function of Fat and Moisture Content. Presented at 2007 ISEA Meeting, Research Triangle Park, NC, October 14 - 18, 2007.
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To be presented at the 2007 ISEA Meeting, Durham/Research Triangle Park, NC October 14-18, 2007
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| PRESENTATION |
Monitoring of Microbes in Drinking Water |
10/15/2007
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ASHBOLT, N. Monitoring of Microbes in Drinking Water. Presented at 2007 International Society for Exposure Analysis Annual Meeting, Raleigh, NC, October 15 - 18, 2007.
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Internationally there is a move towards managing the provision of safe drinking water by direct assessment of the performance of key pathogen barriers (critical control points), rather than end point testing (i.e. in drinking water). For fecal pathogens that breakthrough the various barriers in a drinking water supply system, a residual chlorine concentration may provide further health protection. However, short-term failures (the most likely in systems) are likely to overwhelm the chlorine residual. Of even greater concern is the view that short-term failures are unlikely to be detected by either compliance-type routine monitoring of drinking waters or by pathogen-specific monitoring, such as monthly and more frequent for Cryptosporidium oocysts. On the other hand, a rapid drop in chlorine residual (preferably measured on-line) provides a good trigger for follow-up investigation, well ahead of results from routine microbiological assessment. An emerging issue, however, is the role of distribution system biofilms, which are ubiquitous and a potential site for pathogen accumulation and growth post water treatment. Biofilms are largely extracellular bacterial mucilage that can protect the microbial inhabitants from a residual disinfectant. Further, amoeba that predate on biofilm bacteria are known hosts for the selection of opportunistic bacterial pathogens, such as strains of Legionella, Mycobacterium and other pathogens. The role of biofilms/amoeba for antibiotic gene amplification/transfer has yet to be quantified. Hence, within the risk management paradigm for water system monitoring, assessing biofilm-pathogens deserves greater attention. In addition, we need to move on from only assessing diarrheal impact and include more severe and longer-lasting diseases (sequelae), such as arthritis and various cancers possibly caused by waterborne pathogens. In the future, perhaps we would be better protected by monitoring biofilms via a more metagenomic approach seeking to identify changes in community ecology and key genes? Views expressed are not necessarily those of the US-EPA.
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| PRESENTATION |
Incidental Water Ingestion During Recreational Swimming |
10/15/2007
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DUFOUR, A. P. Incidental Water Ingestion During Recreational Swimming. Presented at 2007 International Society for Exposure Analysis Annual Meeting, Raleigh, NC, October 15 - 18, 2007.
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To be presented at the 2007 International Society for Exposure Analysis Annual Meeting
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| PRESENTATION |
Identification of Mycobacterium Avium Subsp. Hominissuis Isolated from Drinking Water |
11/04/2007
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KING, D. N., A. Beumer, AND S. L. PFALLER. Identification of Mycobacterium Avium Subsp. Hominissuis Isolated from Drinking Water. Presented at Water Technology Conference, Charlotte, NC, November 04 - 08, 2007.
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Mycobacterium avium (MA) is divided into four subspecies based primarily on host-range and consists of MA subsp. avium (birds), MA subsp. silvaticum (wood pigeons), MA subsp. paratuberculosis (broad, poorly-defined host range), and the recently described MA subsp. hominissuis (humans and swine). MA are ubiquitous in the environment and evidence suggests water is a possible source of human exposure. Routes of exposure to waterborne pathogens include ingestion, inhalation of water vapor, and ingestion of produce irrigated or washed with contaminated water. The goal of this study was to determine how specific MA subspecies hominissuis is to humans by subspeciating human clinical isolates of MA and also to determine the proportion of MA isolated from food and water that belong to this subspecies. Understanding the occurrence of MA subsp. hominissuis in water and food will aid the U.S. Environmental Protection Agency in assessing the risk of human exposure to these sources. Temperature growth range, IS1245 copy number, sequencing of the 16S-23S internal transcribed spacer region or hsp65 gene and other methods have been used to differentiate subspecies hominissuis from the other subspecies. MA subsp. hominissuis can grow on Lowenstein Jensen (LJ) medium supplemented with pyruvate at 45 ۫C while bird type isolates cannot (Mijs et al., 2002). Clinical and environmental MA isolates from our collection were inoculated on this medium and allowed to incubate for up to 90 days. Ninety percent of the isolates grew at 25 ۫C and 67% grew at 45 ۫C. Isolates that grew at 45 ۫C also grew at 25 ۫C. Nearly 100% of the human isolates included in this study grew at 45 ۫C, suggesting they are strains of MA subsp. hominissuis. In addition, we performed PCR and sequence analysis on the 3′ end of the hsp65 gene as a means to confirm MA subsp. hominissuis identification. Human isolates had hsp65 sequences closely related to hsp65 sequences from MA subspecies hominissuis described previously and several hsp65 sequences from environmental isolates were identical or very closely related to those of hominissuis. None of the human or environmental isolates in our study had hsp65 sequences that clustered with MA subsp. avium or paratuberculosis. These data suggest treated drinking water and produce are potential sources of exposure to the type of MA that can infect humans.
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| PRESENTATION |
Presence of Pharmaceuticals in Groundwater Down Gradient from Wastewater Lagoons Receiving Partially Treated Wastewater |
11/11/2007
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GLASSMEYER, S., C. Kenah, E. T. Furlong, AND D. Kolpin. Presence of Pharmaceuticals in Groundwater Down Gradient from Wastewater Lagoons Receiving Partially Treated Wastewater. Presented at SETAC North American 28th Annual Meeting, Milwaukee, WI, November 11 - 15, 2007.
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Wastewater can contain traces of the pharmaceutical compounds that are used within a given household or community. These chemicals can act as markers of the wastewater, and their presence can help determine potential sources of contamination of water resources. For this study, groundwater samples were collected up and down gradient from an unlined wastewater treatment plant lagoon serving 400 households. All samples were analyzed for 16 pharmaceuticals using liquid chromatography/ mass spectrometry. The study consisted of two phases. In Phase I (August 2005), 13 different boring locations were drilled, and samples were collected at two to three depths within each hole to determine the boundaries of the leachate plume. In addition, the treated effluent, the municipal lagoon, and the community’s drinking water were also sampled. Monitoring wells (MW) were installed adjacent to three of the bore holes, one up gradient, and two down gradient. During Phase II, the three MWs, the lagoon, and the drinking water source were sampled quarterly (January, April, July and October 2006) to examine seasonal trends in chemical concentrations. Thirteen of the 16 target pharmaceutical compounds were detected at least once. Carbamazepine and sulfamethoxazole were the most commonly detected, occurring in every sample collected from the two down gradient MWs, The carbamazepine concentrations in the MW proximate to the lagoon (MW 2) had concentrations which ranged from 0.316 to 0.642 µg/L, median 0.513 µg/L, which were greater than the concentrations we measured in the lagoon samples (0.02 to 0.557 µg/L, median 0.044 µg/L). The concentrations in MW 2 were even greater than the treated wastewater effluent samples collected in 2002 (max = 0.270 µg/L). The samples collected from multiple depths within each boring location exhibited the highest concentrations of pharmaceuticals in the shallower samples (25-45 feet below the surface), with decreasing cocentrations with increasing depths.
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| PRESENTATION |
Evaluation of a Microarray for Genotyping Noroviruses |
11/10/2007
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FOUT, G. AND N. BRINKMAN. Evaluation of a Microarray for Genotyping Noroviruses. Presented at Third International Calcivirus Conference, Cancun, MEXICO, November 10 - 13, 2007.
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Noroviruses that infect humans are divided into three genogroups based upon their sequence diversity. Of these, genogroups I and II have been identified as leading causes of waterborne disease outbreaks worldwide and are frequently found in rivers and lakes that serve as drinking water sources. To develop a better understanding of the risk posed by these organisms, a more rapid procedure for identifying and genotyping noroviruses in the environment was investigated. Currently, noroviruses are usually detected in water samples using reverse transcriptase-polymerase chain reaction (RT-PCR) based assays. Although sequencing of RT-PCR amplicons is the gold standard for genotyping these viruses, RT-PCR assays of water concentrates often produce a range of non-specific amplicons in a single reaction. This usually necessitates the cloning of products prior to sequencing.
To determine whether genotyping could be performed more rapidly and without the need for cloning, we evaluated the use of a generic microarray format using multiple strain specific probes that would be able to hybridize to region B of the RNA polymerase gene. With this method, a single base extension (SBE) reaction was used to label genotype-specific probes that had annealed to a matching virus sequence. This labeling was done in the presence of single and multiple norovirus templates. These probes were then hybridized to an Affymetrix GenFlex Tag Array and the labeled probes were detected. The specificity of this approach was examined by designing probes with and without nucleotide mismatches. Our results show that the perfect-matched tag-probes were specifically labeled in SBE reactions for all strains examined. In addition, most of the mismatched tag-probes were not labeled and those that were resulted in specific fingerprints that could be used to subtype strains. We also showed that norovirus seeded into source water concentrates can be genotyped. These results demonstrate the utility of the GenFlex Tag Array for genotyping noroviruses from water samples. This approach also could be adapted to include the simultaneous identification of multiple waterborne pathogens.
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| PRESENTATION |
The Enterococcus Qpcr Method for Recreational Water Quality Testing: Testing Background, Performance and Issues |
10/03/2007
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HAUGLAND, R. A. The Enterococcus Qpcr Method for Recreational Water Quality Testing: Testing Background, Performance and Issues. Presented at 7th Annual Great Lakes Beach Association Meeting, Traverse City, MI, October 03 - 05, 2007.
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Currently accepted culture-based monitoring methods for fecal indicator bacteria in surface waters take at least 24 hr to determine if unacceptable levels of fecal pollution have reached our recreational beaches. During this waiting period changing water conditions may result either in unnecessary beach closings or exposures to unsafe conditions. Newer molecular based technologies such as real-time quantitative PCR (QPCR) have the ability to provide measurements of fecal indicator bacteria nucleic acids within a few hours and thus could lead to more accurate and health protective advisories to the public due to their greater timeliness. The National Epidemiological and Environmental Assessment of Recreational (NEEAR) Waters Studies, performed by U.S. EPA and CDC in 2003-2005 demonstrated a correlation between swimming-related gastrointestinal illness rates and QPCR measurements of Enterococcus fecal bacteria at Great Lakes fresh water and Gulf Coast marine water beaches. These results suggest that the QPCR method can provide determinations of fecal pollution levels that are predictive of swimming-related health risks. This presentation will provide an overview of the Enterococcus QPCR method including a brief description of the method and its history, a synopsis of studies that have been conducted to determine its performance characteristics and a discussion of unresolved issues associated with its potential implementation on a national scale.
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| PRESENTATION |
Frank and Opportunistic Pathogen Exposure Risks from Distribution System Biofilms |
11/09/2007
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ASHBOLT, N. Frank and Opportunistic Pathogen Exposure Risks from Distribution System Biofilms. Presented at Meeting at Columbia University, New York, NY, November 09, 2007.
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To be presented at Columbia University, New York, NY, November 9, 2007
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| PRESENTATION |
The Use of Technologies: Exposure (Cross Contamination), Risk Assessment, and Guidelines |
10/17/2007
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ASHBOLT, N. The Use of Technologies: Exposure (Cross Contamination), Risk Assessment, and Guidelines. Presented at Institute of Medicine Rountable: Global Environmental Health, Washington, DC, October 17 - 18, 2007.
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Presentation given at the Institute of Medicine Roundtable: Global Environmental Health, Washington, DC, October 17-18, 2007
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| PUBLISHED REPORT |
Preliminary Comparative Study of Methods to Extract Virus from Raw and Processed Sewage Sludges |
09/28/2007
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RHODES, E., B. MCMINN, N. BRINKMAN, J. CASHDOLLAR, AND G. FOUT. Preliminary Comparative Study of Methods to Extract Virus from Raw and Processed Sewage Sludges. U.S. Environmental Protection Agency, Washington, D.C., EPA/600/R-07/118, 2007.
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Two simple virus extraction techniques were compared to an EPA standard method for detection of human enteric viruses in raw sewage sludge and class A biosolids. The techniques were used to detect both indigenous and seeded virus from a plant that distributes class A material produced by a heat drying process. Virus titers were measured using a plaque assay, a quantal assay and an integrated cell culture-reverse transcription-polymerase chain reaction (ICC-PCR) assay. The best extraction technique overall for detection of indigenous and seeded virus by plaque, quantal, and ICC-PCR assays was a simple chloroform-based technique. The detection of indigenous virus in raw sludge was similar for all three methods by the plaque assay, giving an average of 50±21 PFU/4 grams of sludge. Recovery of seeded virus from raw sludge averaged 0.7% for the EPA versus 22±17% for the other two techniques. Recovery of seeded virus from class A biosolids was similar for all techniques, averaging 95±24%. Both the quantal and the ICC-PCR assays outperformed the plaque assay for virus detection. Virus titers were generally 2->10 fold higher by these assays than by the plaque assay.
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