LITERATURE SURVEY ON SELECTIVE PURIFICATION AND CONCENTRATION OF PATHOGENS (WMI 4587) Final Report to the Department of the Environment
Report No DWI0254

April 1994

EXECUTIVE SUMMARY

The Department of the Environment has identified the selective purification and concentration of pathogenic micro-organisms from water as a topic for investigation. However, before allocating funds for experimental work in this area, it was necessary to arrive at an informed judgement of the potential approaches. Accordingly, WRc plc was invited to review recent developments in the selective concentration of micro-organisms, and to make recommendations for future research.

The objectives of the review were:

1. Critically review the conventional separation and concentration techniques for pathogens which are currently in routine use.

2. Identify novel techniques, from published work and by personal communication, which might offer advantages over the existing methods.

3. Critically review the novel techniques in comparison with the existing ones, taking into account considerations such as speed, materials costs, and ease of use.

4. Draft recommendations for further investigations, taking into account the protozoan parasites Cryptosporidium and Giardia, both of which are a focus of concern in drinking water.

WRc has drawn both on its in-house expertise and on the experience of its many external contacts to identify a wide range of potential separation techniques. This has been supplemented by comprehensive literature searches on internal and external databases. WRc has reached two major conclusions in the course of the review. Firstly, that the failure to develop rapid, efficient, and convenient methods to purify and concentrate pathogens has acted as a brake on the introduction of novel diagnostic procedures in water microbiology. Secondly, that the characteristics of each class of pathogen protozoa, viruses, and bacteria, are substantially different, and therefore warrant different approaches to purification and concentration. For example, the oocysts of the protozoan parasite Cryptosporidium are larger and more robust than bacterial cells, and can therefore be subjected to harsher separation techniques.

To summarise WRc's findings, the following specific recommendations have been made to the Department of the Environment concerning the three classes of pathogen:

Protozoa

• Continue evaluation of advanced large-volume filtration techniques, such as tangential flow filtration.

• Evaluate the applicability of immunomagnetic separation techniques to Cryptosporidium and Giardia purification.

• Initiate research into the structure, biochemical, and biophysical properties of the oocyst wall of Cryptosporidium, with the aim of finding robust and specific epitopes for antibody attachment.

• Collaborate with the manufacturers of flow cytometers to optimise the performance of fluorescence-activated cell sorting for protozoan pathogens in water samples.

Viruses

• Enter into dialogue with centres of expertise in the clinical and pharmaceutical fields. Obtain detailed information on the surface structure of viruses. Identify molecules which could be used to selectively recover viruses from water samples.

Bacteria

• Incorporate commercial products for immunomagnetic separation, which are already available for the bacterial pathogens Salmonella and E. coli0157, into the existing EC methods.

• Investigate the use of immunomagnetic separation with concentrates of water samples. Evaluate alternative methods of using antibodies to selectively purify bacterial pathogens, such as immunoaffinity columns.

• Investigate the effects of environmental stresses and disinfection processes on the potential binding sites for antibodies on the cell surfaces of bacterial pathogens.

The approaches recommended for protozoa and bacteria are based on materials and techniques which are already commercially available. As a result, a reasonable degree of success is ensured in initial investigations, which would provide a sound basis for further, more fundamental work. Unfortunately, the situation with viruses is less satisfactory, with almost no demonstrated alternatives to the existing separation methods in existence. In this area, therefore, a thorough investigation of novel drug-based binding molecules is warranted.

Copies of this report may be available as an Acrobat pdf download under the 'Find Completed Research' heading on the DWI website.