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Demonstration of the feasibility of novel rapid field tests for the detection of antimicrobial growth promoters in animal feed. - VM02154

Antibiotics have been routinely used in animal production for decades worldwide. Various antimicrobial compounds have been found to be effective growth promoting agents when added in low doses to the feed of farm animals.
Previously, the EC banned certain antibiotics used in human medicine from being added to animal feeds. As from January 1st 2006 this was extended and an EU-wide ban on the use of antibiotics as growth promoters in animal feed became effective. The remaining four antibiotics, which had been previously permitted as feed additives are now no longer allowed to be marketed, or used from this date. This ban is the final step in the phasing out of antibiotics used for prophylactic purposes and is part of the EC’s overall strategy to address the increasing emergence of bacteria and other microbes resistant to antibiotics, due to their overexploitation or misuse.
In order for this ban to be applied effectively, analytical systems are required to detect potential abuse.Traditionally this has been a laboratory based activity, however, the possibility of undertaking analysis of samples at the farm is highly attractive. For example, multiple feed types may be present and the collection, labelling and transportation of these all have cost implications and it is likely that only a limited number would be sampled. The ability to carry out a rapid test at site would allow a more rigorous sampling regime to be used. A positive finding, while it would require confirmatory analysis using a lab based technology, could also trigger a more reactive response than laboratory testing allows, for example, additional sampling of livestock to prove the band substance was indeed being abused.
The attraction of applying analysis for field analysis is well understood in a number of fields including forensic analysis, anti-terrorism, healthcare and crop protection to name a few and a number of developments allowing rapid testing without the need for laboratory based equipment have resulted. Where high sensitivity is required, the majority of these utilise immunochemistry based detection systems, for example lateral flow devices (as used for home pregnancy tests) or surface plasmon resonance (SPR) technologies. Lateral flow devices have the attraction of low cost, ease of use and high portability. However, they are best suited to qualitative yes/no applications. SPR based systems have the capability of high sensitivity and, despite their technological complexity, can be operated in simple turn-key mode. Significant disadvantages of this technology are the relatively high capital / running costs and limited portability.

An alternative and highly attractive approach is potentiometric biosensors. In these devices an antibody or other recognition molecule is immobilised to a polypyrrole coated electrode and the binding of an appropriate substrate to this antibody causes a change in potential at the electrode. The presence of the analyte being measured affects the amount of enzyme bound and in turn the measured voltage. This technology has the capability of attaining high sensitivity using low tech, low cost and highly portable instrumentation consisting of, at its most basic level, a millivolt meter. With appropriate software on a laptop or hand held computer either quantitative or qualitative yes/no response data can be generated. An important aspect of any device designed for field application is ease of use and ready interpretation of results. Incorporating a computer system also allows the user to follow simple step by step instructions for use of the device, further software can be included to interpret the results of the readouts from the sensors and provide straightforward output to the user. The actual electrodes are very compact, low cost disposable devices and several electrodes can be placed on a single device allowing the concurrent analysis of multiple analytes.
As the sensor works in the liquid phase and most of the materials being tested are solids methods are required to extract the antimicrobial agents. These methods must be generic, allowing the extraction of multiple analytes from a wide range of feed types, for example grain or compounded pellet feeds. The methods must also be capable of use in the field and result in a liquid sample compatible with the biosensor, typically an aqueous medium of near physiological pH and concentration.

The primary objective of this research project is therefore to develop a portable biosensor and extraction method for one or more antimicrobial agents in animal feedstuffs and to demonstrate applicability in a field setting. The ability to detect several agents using a single test stick will also be evaluated.
Objective 1
Selection of analytes based upon relevance and availability of antibody and enzyme labelled antigen. (HFL). Duration 1 month.

Objective 2
Develop a “proof of principle” potentiometric biosensor based assay for at least one antibiotic. (HFL). Duration 8 months

Objective 3
Develop initial generic extraction method suitable for field use (CSL). Duration 8 months

Objective 4
Transfer extraction / biosensor technologies from CSL to HFL and vice versa. (HFL / CSL). Duration 1 month

Objective 5
Optimise sensor / extraction technology (HFL / CSL). Duration 2 months

Objective 6
Decision point - Progress to multianalyte biosensor (HFL / CSL). Duration 1 month

Objective 7
Complete investigation of the feasibility of transfering the prototype single assays onto a multi-electrode device (HFL). Duration 3 months

Objective 8
Decision point - Progress to robustness testing (HFL / CSL). Duration 1 month

Objective 9
Demonstrate applicability of the prototype biosensor technology to field-test usage (HFL / CSL). Duration 2 months

Objective 10
Conduct a full cost benefit analysis for the prototype potentiometric biosensor assay. (HFL / CSL). Duration 1 month

Objective 11
Produce a final report. (HFL / CSL). Duration 1 month

Time-Scale and Cost
From: 2007

To: 2009

Cost: £191,811
Contractor / Funded Organisations
Central Science Laboratory, HFL Limited
Animal Feed              
Animal Health              
Animal Production              
Antimicrobial Growth Promotors              
Antimicrobial Resistance              
Livestock Farming              
Method Development              
Plants and Animals              
Veterinary Medicines              
Fields of Study
Animal Health
Veterinary Medicine