The assurance of safe food and water is paramount to the health and performance of the warfighter. Any technology to assess the chemical and microbial purity of food and water under field conditions must meet rigorous criteria: it must be readily portable, provide timely results (no more than 4 hours), have adequate sensitivity (1 cfu/100 mL for potable water), be compatible with military power sources, and be of complexity appropriate for operation by a Preventive Medicine Specialist. The nomination of an Army Science and Technology Objective (STO) leads to assessment of existing technologies and commercial products; identification of users, regulators and developers; definition of essential capabilities; and consideration of potential obstructions. The U.S. Army Center for Environmental Health Research has identified a number of technologies for detecting microbial contaminants in food and water and has pursued development of the more promising examples. This paper examines developmental risks in the context of the STO and offers some insight and strategies to manage them.

The Water Environment Research Foundation (WERF) is a not- for-profit organization established in 1989 to advance the science and technology of a broad spectrum of environmental and human health concerns to the wastewater industry and the public. It is a unique public/private partnership between utilities, academia, government, and industry, committed to funding research by leveraging resources and expertise to develop and disseminate sound scientific and technological information. Funded by subscribers, grants and contributions, WERF manages a broad array of research projects aimed at protecting human health and the environment. While WERF funds and manages projects, the actual research is carried out by individual organizations or teams composed of utilities, consultants, universities, and industrial or commercial firms. Examples of WERF's current research program include the investigation of on- line monitoring techniques for microbial and chemical contaminants in water and wastewater, optimization of processes for pathogen removal and inactivation, improved treatment of toxic compounds, and assessing the potential risks to public health from exposure to these microbial and chemical contaminants. This paper will provide an overview of the program, research funded to date, and technology needs for the future.

Operational environments for military forces are becoming potentially more dangerous due to the increased number, use, and misuse of toxic chemicals across the entire range of military missions. Defense personnel may be exposed to harmful chemicals as a result of industrial accidents or intentional or unintentional action of enemy, friendly forces, or indigenous populations. While there has been a significant military effort to enable forces to operate safely and survive and sustain operations in nuclear, biological, chemical warfare agent environments, until recently there has not been a concomitant effort associated with potential adverse health effects from exposures of deployed personnel to toxic industrial chemicals. To provide continuous real-time toxicity assessments across a broad spectrum of individual chemicals or chemical mixtures, an Environmental Sentinel Biomonitor (ESB) system concept is proposed. An ESB system will integrate data from one or more platforms of biologically-based systems and chemical detectors placed in the environment to sense developing toxic conditions and transmit time-relevant data for use in risk assessment, mitigation, and/or management. Issues, challenges, and next steps for the ESB system concept are described, based in part on discussions at a September 2001 workshop sponsored by the U.S. Army Center for Environmental Health Research.

Robowell is an automated system and method for monitoring ground-water quality. Robowell meets accepted manual- sampling protocols without high labor and laboratory costs. Robowell periodically monitors and records water-quality properties and constituents in ground water by pumping a well or multilevel sampler until one or more purge criteria have been met. A record of frequent water-quality measurements from a monitoring site can indicate changes in ground-water quality and can provide a context for the interpretation of laboratory data from discrete samples. Robowell also can communicate data and system performance through a remote communication link. Remote access to ground-water data enables the user to monitor conditions and optimize manual sampling efforts. Six Robowell prototypes have successfully monitored ground-water quality during all four seasons of the year under different hydrogeologic conditions, well designs, and geochemical environments. The U.S. Geological Survey is seeking partners for research with robust and economical water-quality monitoring instruments designed to measure contaminants of concern in conjunction with the application and commercialization of the Robowell technology. Project publications and information about technology transfer opportunities are available on the Internet at URL http://ma.water.usgs.gov/automon/

Particulates that move with ground water and those that are artificially mobilized during well purging could be incorporated into water samples during collection and could cause trace-element concentrations to vary in unfiltered samples, and possibly in filtered samples (typically 0.45-um (micron) pore size) as well, depending on the particle-size fractions present. Therefore, measured concentrations may not be representative of those in the aquifer. Ground water may contain particles of various sizes and shapes that are broadly classified as colloids, which do not settle from water, and particulates, which do. In order to investigate variations in trace-element concentrations in ground-water samples as a function of particle concentrations and particle-size fractions, the U.S. Geological Survey, in cooperation with the U.S. Air Force, collected samples from five wells completed in the unconfined, oxic Kirkwood-Cohansey aquifer system of the New Jersey Coastal Plain. Samples were collected by purging with a portable pump at low flow (0.2-0.5 liters per minute and minimal drawdown, ideally less than 0.5 foot). Unfiltered samples were collected in the following sequence: (1) within the first few minutes of pumping, (2) after initial turbidity declined and about one to two casing volumes of water had been purged, and (3) after turbidity values had stabilized at less than 1 to 5 Nephelometric Turbidity Units. Filtered samples were split concurrently through (1) a 0.45-um pore size capsule filter, (2) a 0.45-um pore size capsule filter and a 0.0029-um pore size tangential-flow filter in sequence, and (3), in selected cases, a 0.45-um and a 0.05-um pore size capsule filter in sequence. Filtered samples were collected concurrently with the unfiltered sample that was collected when turbidity values stabilized. Quality-assurance samples consisted of sequential duplicates (about 25 percent) and equipment blanks. Concentrations of particles were determined by light scattering.

Since September 11, 2001, the threat of terrorist attack and biological warfare within U.S. borders has become a sobering reality. In an effort to aid military personnel and the public at large, we have been investigating the utility of surface-enhanced Raman spectroscopy (SERS) to provide rapid identification of chemical agents directly, and biological agents through their chemical signatures. This approach is based on the ability of Raman spectroscopy to identify molecular structure through the abundant vibration information provided in spectra and the ability of SERS to detect extremely low concentrations (e.g. part-per-billion) through the enhancement of Raman scattering by six orders of magnitude or more. Toward the goal of developing a portable analyzer, we have been studying the ability of two SER media to obtain continuous (i.e., reversible) and quantitative (i.e., reproducible) measurements. Here we compare measurements of nucleic acid bases, adenosine monophosphate, and ribonucleic acid extracted from Escherichia coli, Bacillus subtilis and Staphylococcus aureus obtained by electrolytic SERS and metal-doped sol-gel SERS. The capabilities of these SER media are summarized in terms of rapid detection of B. anthracis and dipicolinic acid.

Spatial, temporal, and spectral characteristics of stimulated scatterings (SS) of light, excited in water, have been investigated both in nano- and picosecond range in different experimental conditions and compared with the analogous properties in other liquids. The results can give important information about the structure of water, its purity, and additions, which may lead to its pollution. Conditions have been defined for stimulated excitation of SS in one spatial mode and with maximum pulse conversion. Stimulated scatterings of light can be used for water quality control, which may be fulfilled in a very short time, and for information processing: amplitude-phase structure of complex light fields may be registered in water as dynamic hologram and reconstructed in a real-time scale.

Nomadics and Professor Timothy Swager have collaborated extensively on the development of Amplifying Fluorescent Polymer (AFP) sensing technologies. AFPs are fully conjugated polymers which link many fluorphores into what is in effect, a molecular wire. Once linked, the ensemble of fluorophores will respond to a transduction event at any individual fluorophore. As a result, each individual analyte interaction will produce an effect in a much larger ensemble of fluorphores, amplifying each sensing event. Nomadics capitalized on this effect to construct a handheld landmine detector, which can identify buried landmines by sensing the trace signature of TNT vapor emanating from a mine. This presentation will describe efforts of Nomadics and the Swager group to leverage the intrinsic amplification of AFP systems for bioanalyte detection.

Rapid detection and identification of bacteria and other pathogens is important for many civilian and military applications. The profiles of biological markers such as fatty acids can be used to characterize biological samples or to distinguish bacteria at the gram-type, genera, and even species level. Common methods for whole cell bacterial analysis are neither portable nor rapid, requiring lengthy, labor intensive sample preparation and bench-scale instrumentation. These methods chemically derivatize fatty acids to produce more volatile fatty acid methyl esters (FAMEs) that can be separated and analyzed by a gas chromatograph (GC)/mass spectrometer. More recent publications demonstrate decreased sample preparation time with in situ derivatization of whole bacterial samples using pyrolysis/derivatization. Ongoing development of miniaturized pyrolysis/GC instrumentation by this department capitalizes on Sandia advances in the field of microfabricated chemical analysis systems ((mu) ChemLab). Microdevices include rapidly heated stages capable of pyrolysis or sample concentration, gas chromatography columns, and surface acoustic wave (SAW) sensor arrays. We will present results demonstrating the capabilities of these devices toward fulfilling the goal of portable, rapid detection and early warning of the presence of pathogens in air or water.

Detection of multiple chemical and biological weapons (CBW) agents and/or complex mixtures of toxic industrial chemicals (TIC) is imperative for both the commercial and military sectors. In a military scenario, a multi-CBW attack would create confusion, thereby delaying decontamination and therapeutic efforts. In the commercial sector, polluted sites invariably contain a mixture of TIC. Novel detection systems capable of detecting CBW and TIC are sorely needed. While it may be impossible to build a detector capable of discriminating all the possible combinations of CBW, a detection system capable of statistically predicting the most likely composition of a given mixture is within the reach of current emerging technologies. Aquatic insect-gene activity may prove to be a sensitive, discriminating, and elegant paradigm for the detection of CBW and TIC. We propose to systematically establish the expression patterns of selected protein markers in insects exposed to specific mixtures of chemical and biological warfare agents to generate a library of biosignatures of exposure. The predicting capabilities of an operational library of biosignatures of exposures will allow the detection of emerging novel or genetically engineered agents, as well as complex mixtures of chemical and biological weapons agents. CBW and TIC are discussed in the context of war, terrorism, and pollution.

In many circumstances, the ability to perform on-site, point-of-collection analysis can play a pivotal role in the goals or requirements of the inquiry. Toward this end, the use of commercial or customized kits, which require the analyst to manually perform the metering and mixing of reagents with the sample and the subsequent visual, spectrophotometric or other interpretation of the results, has become widespread. Often, these methods can suffer from poor reproducibility and sensitivity in addition to being tedious and time consuming. Flow analysis methods, such as traditional flow injection analysis (FIA) and the more recent sequential injection analysis (SIA), have found widespread use in the automation of sample and reagent handling and subsequent analysis for many important analytes. These methods can be completely automated and offer excellent reproducibility, minimized analysis time, and in certain configurations, very high sensitivity. We have developed a miniaturized, fully portable SIA-based instrument for on-site screening for chemical weapons degradation products during challenge inspections under the Chemical Weapons Convention, as well as for the sensitive analysis of other important environmental analytes. In this paper, we will discuss our portable SIA design, the analytical approaches utilized, and results obtained for the analysis of representative chemical weapons degradation compounds.

Developments in rapid detection technologies have made countless improvements over the years. However, because of the limited sample that these technologies can process in a single run, the chance of capturing and identifying a small amount of pathogens is difficult. The problem is further magnified by the natural random distribution of pathogens in foods. Methods to simplify pathogenic detection through the identification of bacteria specific VOC were studied. E. coli O157:H7 and Salmonella typhimurium were grown on selected agar medium to model protein, and carbohydrate based foods. Pathogenic and common spoilage bacteria (Pseudomonas and Morexella) were screened for unique VOC production. Bacteria were grown on agar slants in closed vials. Headspace sampling was performed at intervals up to 24 hours using Solid Phase Micro-Extraction (SPME) techniques followed by GC/MS analysis. Development of unique volatiles was followed to establish sensitivity of detection. E. coli produced VOC not found in either Trypticase Soy Yeast (TSY) agar blanks or spoilage organism samples were - indole, 1-decanol, and 2-nonanone. Salmonella specific VOC grown on TSY were 3-methyl-1-butanol, dimethyl sulfide, 2-undecanol, 2-pentadecanol and 1-octanol. Trials on potato dextrose agar (PDA) slants indicated VOC specific for E. coli and Salmonella when compared to PDA blanks and Pseudomonas samples. However, these VOC peaks were similar for both pathogens. Morexella did not grow on PDA slants. Work will continue with model growth mediums at various temperatures, and mixed flora inoculums. As well as, VOC production based on the dynamics of bacterial growth.

An artificial nose based on microsphere sensor arrays has been developed for the discrimination of numerous volatile organic compounds. Sensor elements consist of 3-5 micron diameter silica and polymer spheres that have a fluorescent, solvatochromic dye adsorbed to the microsphere surface. These sensors respond to changes in the local polarity of the environment by shifting their excitation and/or emission characteristics, thereby indicating the presence of different volatile compounds. High-density microsphere arrays are fabricated which contain thousands of individual sensor elements and multiple copies of each sensor type. By monitoring the sensors temporal fluorescence responses with a CCD camera, unique patterns are recorded that identify individual analytes or are characteristic of a complex mixture. By summing over the redundant sensor elements within an array, the signal-to-noise ratio can be enhanced. These types of sensor arrays have been used to detect and discriminate between different bacterial strains such as Escherichia coli based on characteristic odors from the live and dead bacteria.

The Auburn University Detection and Food Safety Center has demonstrated real-time biosensor for the detection of Salmonella typimhurium, consisting of a thickness shear-mode (TSM) quartz resonator with antibodies immobilized in a Langmuir-Blodgett surface film. Scanning Electron Microscopy (SEM) images of bound Salmonella bacteria to both polished and unpolished TSM resonators were taken to correlate the mass of the bound organism to the Sauerbrey equation. Theoretical frequency shifts for unpolished TSM resonators predicted by the Sauerbrey equation are much smaller than experimentally measured frequency shift. The Salmonella detector operates in a liquid environment. The viscous properties of this liquid overlayer could influence the TSM resonator's response. Various liquid media were studied as a function of temperature (0 to 50 degree(s)C). The chicken exudate samples with varying fat content show coagulation occurring at temperatures above 35 degree(s)C. Kinematic viscosity test were performed with buffer solutions containing varying quantities of Salmonella bacteria. Since the TSM resonators only entrain a boundary layer of fluid near the surface, they do not respond to these background viscous property changes. Bilk viscosity increases when bacteria concentrations are high. This paper describes investigations of TSM resonator surface acoustic interactions - mass, fluid viscosity, and viscoelasticity - that affect the sensor.

The inductor-capacitor (LC) sensor, comprised of a thick- film printed LC resonant circuit the resonant frequency of which can be remotely detected with a loop antenna, has been used for the monitoring of temperature, humidity, atmospheric pressure, salt concentration, and complex permittivity, as well as the detection of bacteria in a liquid medium based upon changes in the complex permittivity due to the bacteria growth. Due to its low unit cost and wireless detection, the LC sensor is potentially suitable for commercial scale monitoring of food quality. This paper includes the operational principles and design criteria of the LC sensor, and illustrates the monitoring of bacteria growth in milk, meat, and beer.

Using a chemiluminescent fiber optic biosensor and magnetic particles, a simple, sensitive and rapid method to determine Staphylococcus aureus enterotoxin A (SEA) in military ration components was developed. Anti-staphylococcal enterotoxin A (Anti-SEA) was immobilized on magnetic particles and incubated with SEA. The beads were then collected and rinsed on a membrane filter (0.45um). The captured toxin was then selectively labeled with a monoclonal-horseradish peroxidase (POD) conjugate. SEA concentration was detected with a luminometer and a chemiluminescent enhancing reagent. Total assay time was 1.25 hours. Chemiluminescent signal due to nonspecific binding was tested with various blocking agents. Phosphate buffered saline with casein had the lowest background signal. Primary antibody concentration, secondary labeled antibody concentration and chemiluminescent substrate type were also evaluated to optimize signal intensity. The chemiluminescent fiber optic biosensor assay was compared to the Analyte 2000, a commercial fluorescent fiber optic biosensor. This assay consisted of immobilizing Anti-SEA on polystyrene waveguides, and incubating the waveguides with the toxin. The waveguide was incubated with a selectively labeled monoclonal-CY5 Dye conjugate. The sensitivity of chemiluminescent and fluorescent immunoassays were 1 ng, significantly lower than the levels needed to cause illness.