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.

Current and Developing Technologies for Monitoring Agents of Bioterrorism and Biowarfare

In the spring and summer of 2001, 47 fathers, 48 mothers and 117 children of Iowa farm and non-farm households were recruited to participate in a study investigating take-home pesticide exposure. On two occasions approximately 1 month apart, urine samples from each participant and dust samples from various rooms were collected from each household and were analyzed for atrazine, metolachlor, glyphosate and chlorpyrifos or their metabolites. The adjusted geometric mean (GM) level of the urine metabolite of atrazine was significantly higher in fathers, mothers and children from farm households compared with those from non-farm households (P < or = 0.0001). Urine metabolites of chlorpyrifos were significantly higher in farm fathers (P = 0.02) and marginally higher in farm mothers (P = 0.05) when compared with non-farm fathers and mothers, but metolachlor and glyphosate levels were similar between the two groups. GM levels of the urinary metabolites for chlorpyrifos, metolachlor and glyphosate were not significantly different between farm children and non-farm children. Farm children had significantly higher urinary atrazine and chlorpyrifos levels (P = 0.03 and P = 0.03 respectively) when these pesticides were applied by their fathers prior to sample collection than those of farm children where these pesticides were not recently applied. Urinary metabolite concentration was positively associated with pesticide dust concentration in the homes for all pesticides except atrazine in farm mothers; however, the associations were generally not significant. There were generally good correlations for urinary metabolite levels among members of the same family.

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Urinary Pesticide Concentrations Among Children, Mothers and Fathers Living in Farm and Non-Farm Households in Iowa

Public concern over pesticide residues has been increasing dramatically owing to the high toxicity and bioaccumulation effects of pesticides and the serious risks that they pose to the environment and human health. It is therefore crucial to monitor pesticide residues by using various analytical methods and techniques, especially highly sensitive, highly selective, simple, rapid, cost-effective, and portable ones. Biosensor strategies have become research hotspots and ideal candidates for pesticide detection, having such features as high sensitivity, fast response, robustness, low cost and miniaturization, as well as in situ and real-time monitoring. This review covers advances in the design and fabrication of biosensors for pesticide detection since 2005. Special emphasis is placed on the state-of-art selection of receptors, the use of different transduction techniques and fast screening strategies, and the application of various biosensors developed in food and environmental safety. Both advantages and drawbacks of these techniques are then summarized. Finally, challenges, strategies, and perspectives in further developing pesticide biosensors are also discussed.

Advances in pesticide biosensors: current status, challenges, and future perspectives.

Biomonitoring of exposure to pesticides

The presence of trace contamination of soil and groundwater with explosives is an ongoing concern, for which improved methods are required to facilitate their detection and quantification. This is true both for the monitoring of remediation and for site characterization. Immunosensors have been found effective for solution-phase detection of environmental contaminants. Our work utilized the Luminex100 (flow cytometer) to detect TNT in a multiplexed displacement immunoassay format. The Luminex100 can perform a multiplexed assay by discriminating between up to 100 different bead sets. We used this capability to evaluate four different TNT monoclonal antibodies, two recombinant TNT antibodies, and a control antibody simultaneously for the rapid detection of TNT and other nitroaromatics. TNT could be detected at 0.1 ppb and quantified over the range of 1.0 ppb to 10 ppm. In addition, the assay was shown to be effective in various matrixes such as lake water, seawater, and acetone extracts of soil. Seawater re...

TNT detection using multiplexed liquid array displacement immunoassays

Body burdens from exposures to pesticides may be estimated from urinary analyses of pesticide parent/metabolite concentrations. Pesticide applicators and others are often exposed to numerous unrelated pesticides, either sequentially or simultaneously. Classically, body burdens of pesticides are analyzed using chemical/instrumental analysis (CIM) or enzyme immunoassays (EIAs). Both of these technologies can usually be used to quantitate one analyte (or closely related groups of analytes) per analysis. Alternatively, multiple analytes can be measured simultaneously using a multiplexed fluorescence covalent microbead immunoassay (FCMIA). We developed a multiplexed FCMIA to simultaneously measure glyphosate (Gly), atrazine (Atz), and metolachlor mercapturate (MM) in water and urine. The assay had least detectable doses (LDDs) in water/diluted urine of 0.11/0.09 ng/ml (Gly, water/urine LDD), 0.10/0.07 ng/ml (Atz) and 0.09/0.03 ng/ml (MM). The sensitivity for the measurement of Gly was enhanced by derivatization. All assays gave linear responses from the LDDs for each respective pesticide to 300 ng/ml. There was no cross-reactivity between the three analytes. Using a 96-well microplate and an autosampler, as many as 288 separate analyses can be completed in approximately 120 min with precision, sensitivity, and specificity equivalent to, if not better, than that found when these same analytes are measured by CIM or EIA.

Development of a sensitivity enhanced multiplexed fluorescence covalent microbead immunosorbent assay (FCMIA) for the measurement of glyphosate, atrazine and metolachlor mercapturate in water and urine.

Aims: To evaluate potential exposure to Bacillis anthracis ( Ba ) spores in sampling/decontamination workers in the aftermath of an anthrax terror attack. Methods: Fifty six serum samples were obtained from workers involved in environmental sampling for Ba spores at the American Media, Inc. (AMI) building in Boca Raton, FL after the anthrax attack there in October 2001. Nineteen sera were drawn from individuals both pre-entry and several weeks after entrance into the building. Nine sera each were drawn from unique individuals at the pre-entry and follow up blood draws. Thirteen donor control sera were also evaluated. Individuals were surveyed for Ba exposure by measurement of serum Ba anti-protective antigen (PA) specific IgG antibodies using a newly developed fluorescent covalent microsphere immunoassay (FCMIA). Results: Four sera gave positive anti-PA IgG results (defined as anti-PA IgG concentrations ⩾ the mean μg/ml anti-PA IgG from donor control sera (n = 13 plus 2 SD which were also inhibited ⩾ 85% when the serum was pre-adsorbed with PA). The positive sera were the pre-entry and follow up samples of two workers who had received their last dose of anthrax vaccine in 2000. Conclusion: It appears that the sampling/decontamination workers of the present study either had insufficient exposure to Ba spores to cause the production of anti-PA IgG antibodies or they were exposed to anthrax spores without producing antibody. The FCMIA appears to be a fast, sensitive, accurate, and precise method for the measurement of anti-PA IgG antibodies.

https://oem.bmj.com/content/oemed/61/8/703.full.pdf

Determination of serum IgG antibodies to Bacillus anthracis protective antigen in environmental sampling workers using a fluorescent covalent microsphere immunoassay.

Background: Asthma in bakery workers is one of the most frequently occurring forms of occupational asthma in the world. Experience from other countries has shown the prevalence of sensitization (IgE) to bakery-associated allergens (BAAs) (wheat [W], flour dust [FD], -amylase [AA]) in bakery workers to be 5% to 53%, whereas the prevalence in nonoccupationally exposed individuals was estimated to be 1.2% to 6.4%. Objective: To estimate the prevalence of BAA sensitization by measuring BAA specific IgE in the residual serum tubes of volunteer blood donors. Methods: Serum samples from 534 volunteer blood donors were measured for anti-W, anti-FD, and anti-AA specific IgE antibodies (in duplicate) using the AlaSTAT microplate assay. Samples with BAA IgE concentrations of 0.35 kU/L or greater were considered positive. Results: Nineteen of 530 serum samples (3.6%; 95% confidence interval [CI], 3.3%–3.9%) were positive for W (range, 0.38–3.61 kU/L), whereas 31 of 534 (5.8%; 95% CI, 5.3%–6.3%) were positive for FD (range, 0.35–2.34 kU/L) and 5 of 529 (1.0%; 95% CI, 0.9%–1.1%) were positive for AA (range, 0.38–1.59 kU/L). Thirteen serum samples were positive for both W and FD; 1 sample each was positive for W and AA and FD and AA. Conclusions: The prevalence of IgE sensitization in serum samples from a relatively large unselected population of volunteer blood donors is 1.0% for AA, 3.6% for W, and 5.8% for FD, which agrees well with data from other countries for sensitization prevalence rates for nonoccupationally exposed individuals. Ann Allergy Asthma Immunol. 2004;92:649–653.

C. A. F. Striley

Papers

Development of a sensitivity enhanced multiplexed fluorescence covalent microbead immunosorbent assay (FCMIA) for the measurement of glyphosate, atrazine and metolachlor mercapturate in water and urine.

Determination of serum IgG antibodies to Bacillus anthracis protective antigen in environmental sampling workers using a fluorescent covalent microsphere immunoassay.

Evaluation of the prevalence of anti-wheat-, anti—flour dust-, and anti-alpha amylase-specific IgE antibodies in blood donors