SUMMARY Recent events have made public health officials acutely aware of the importance of rapidly and accurately detecting acts of bioterrorism. Because bioterrorism is difficult to predict or prevent, reliable platforms to rapidly detect and identify biothreat agents are important to minimize the spread of these agents and to protect the public health. These platforms must not only be sensitive and specific, but must also be able to accurately detect a variety of pathogens, including modified or previously uncharacterized agents, directly from complex sample matrices. Various commercial tests utilizing biochemical, immunological, nucleic acid, and bioluminescence procedures are currently available to identify biological threat agents. Newer tests have also been developed to identify such agents using aptamers, biochips, evanescent wave biosensors, cantilevers, living cells, and other innovative technologies. This review describes these current and developing technologies and considers challenges to rapid, accurate detection of biothreat agents. Although there is no ideal platform, many of these technologies have proved invaluable for the detection and identification of biothreat agents.

Aims:  Concentration of pathogens diluted in large volumes of water is necessary for their detection. An automated concentration system placed online in drinking water distribution systems would facilitate detection and mitigate the risk to public health. Methods and Results:  A prototype concentrator based on dead‐end hollow fibre ultrafiltration was used to concentrate Bacillus atrophaeus spores directly from tap water. Backflush was used to recover accumulated particulates for analysis. In field tests conducted on a water utility distribution system, 3·2 × 104–1·4 × 106 CFU ml−1 (6·1 × 106–3·0 × 108 CFU) were recovered from the filter when 2·9 × 107–1·0 × 109 CFU were spiked into the system. Per cent recovery ranged from 21% to 68% for flow volumes of 15–21 l. Tests using spore influent levels <10 CFU l−1 (spike < 1000 CFU) yielded 23–40% recovery for volumes >100 l. Conclusions:  B. atrophaeus spores at levels <10 CFU l−1 were concentrated directly from tap water using an automated dead‐end hollow‐fibre ultrafiltration system. Significance and Impact of the Study:  The prototype concentrator represents a critical step towards an autonomous system that could be installed in drinking water distribution lines or other critical water lines to facilitate monitoring. Recovered samples can be analysed using standard or rapid biosensor methods.

An automated concentration system (ACS) based on dead-end ultrafiltration was used in this study to concentrate bacteria, including Escherichia coli O157:H7, from 50-liter produce washes (PWs, sieved produce wash). Cells trapped in the filters were recovered in approximately 400 ml of buffer to create PW retentates (PWRs). Extent of concentration was determined by analyzing PWs and PWRs for total coliform bacteria and E. coli O157:H7 using standard methods. In addition, an electrochemiluminescence immunoassay was evaluated for detection of E. coli O157:H7 in spiked PWs and PWRs to demonstrate usefulness of the ACS for same-day detection. The levels of total coliform bacteria and E. coli O157:H7 in PWRs were higher than those in PWs by 1.85 ± 0.41 log most probable number per 100 ml and 1.82 ± 0.24 log CFU/ml, respectively. Electrochemiluminescence detection of E. coli O157:H7 was accomplished within 2 h using ACS concentration of lettuce and spinach wash water artificially spiked with the pathogen at levels as low as 0.36 log CFU/ml and 1.39 log CFU/ml, respectively. Detection of E. coli O157:H7 at -0.93 ± 0.15 log CFU/ml in lettuce wash occurred within approximately 6 h when a 4-h enrichment step was added to the procedure. Use of dead-end ultrafiltration increased bacterial concentrations in PWR and allowed same-day detection of low levels of E. coli O157:H7 in PW. This concentration system could be useful to improve the sensitivity of current rapid methods for detection of low levels of foodborne pathogens in PW water.

Aims: A Portable Multi-use Automated Concentration System (PMACS) concentrates micro-organisms from large volumes of water through automated dead-end ultrafiltration and backflushing. The ability to detect microbial targets from ground, surface and cooling tower waters collected using standard methods was compared with samples from the PMACS in this study. Methods and Results: PMACS (100 l) and standard grab samples (100-500 ml) were collected from sites in Florida and South Carolina, USA. Samples were analysed for the presence of faecal indicator bacteria (FIB; ground and surface water) or Legionella pneumophila (Lp; cooling tower water). FIB were enumerated by growth on selective media following membrane filtration or in IDEXX defined substrate media. Lp cells were detected by direct fluorescence immunoassay using FITC-labelled monoclonal antibodies targeting serogroups 1, 2, 4 and 6. FIB were found in PMACS samples from ground and surface waters when their concentrations were below detection limits in grab samples. The concentrations of Lp in cooling tower samples collected over 5 months were more consistent in PMACS samples than grab samples.Conclusions: These data demonstrate that PMACS concentration is advantageous for water monitoring. FIB were detected in PMACS samples when their concentrations were below the detection limits of the standard methods used. PMACS processing provided more representative samples of cooling tower waters reducing sample variability during long-term monitoring. Significance and Impact of the Study: This study highlights the utility of PMACS processing for enhanced monitoring of water for low-level microbial targets and for reducing sample variability in long-term monitoring programmes.

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