Deliberately induced infections in well-defined animal models have a central place in the study of infectious diseases. Central to the use of animals in research is the promotion of the 3Rs: replacement, refinement and reduction. If an alternative is available it must be used and where it is not, scientists are compelled to get the best data using the fewest animals and causing the minimum pain, suffering and distress.

I am a scientist currently researching enteropathogenic Escherichia coli (EPEC), which causes infantile diarrhoea in developing countries, and enterohaemorrhagic E. coli (EHEC), responsible for a wide spectrum of illnesses. While EHEC and EPEC are poorly pathogenic in mice, a close relative, Citrobacter rodentium, is a natural mouse pathogen that uses the same mechanisms to infect the murine colon.

As a researcher using mice, I firmly believe that animals should not be considered as mere laboratory consumables but as the sentient creatures they are. I find working with animals stressful; I do not enjoy seeing an animal in pain and so feel a responsibility to ensure that this does not happen. As a result the 3Rs are central to my everyday thinking. However, for me, refinement also goes beyond minimising pain and suffering: it is an ongoing process and includes constantly re-evaluating the design of the experiments to make sure the model used is the best representation of the disease under study.

My work on C. rodentium began with the development of a bioluminescent derivative, which we used to follow in vivo colonisation dynamics after oral gavage with bioluminescence imaging. One of the striking findings of our first study was that prior to colonisation of the colon, C. rodentium colonises a patch of lymphoid tissue known as the caecal patch. This suggests the caecal patch is where the organism adapts to the murine gastrointestinal tract.

Like EPEC and EHEC, C. rodentium is spread through the faecal–oral route. We then wondered what would happen if we orally infected one mouse and housed it with uninfected littermates. Would C. rodentium be transmitted and, if so, would the infection be any different? We found that after passage through a mouse C. rodentium becomes 'hyperinfectious', colonising further mice without the need for adaptation in the caecal patch and with a 1000-fold lower infectious dose than bacteria grown in laboratory media.

The most important impact of these findings was the development of an animal model that more realistically modelled the human infection, while requiring far fewer animals to undergo the invasive oral gavage procedure. Furthermore, the use of bioluminescence imaging allows infection to be followed in individual animals in real time, reducing the numbers of animals required and allowing pathogen burden to be measured before clinical symptoms appear.

Our application of the 3Rs has made our model more relevant to the disease we are studying. This has wider implications. In almost all models of bacterial infections, animals are deliberately infected with high doses of bacteria grown in artificial laboratory media, rather than being exposed to the organism in a way that resembles the natural route of transmission. Surely the unique pathogenic phenotypes that result from residence in their natural microenvironments are impossible to duplicate by such artificial means? While it may not be possible or practical to include transmission in many disease models, it is imperative that researchers attempt to ensure that the inoculating cultures used are appropriate to the process under study.

Siouxsie Wiles has been a researcher at Imperial College London since 2000. Her work with Citrobacter rodentium won the inaugural 3Rs prize from the National Centre for the Replacement, Refinement and Reduction of Animals in Research.

External links

• Wiles S et al. Organ specificity, colonization and clearance dynamics in vivo following oral challenges with the murine pathogen Citrobacter rodentium. Cell Microbiol 2004;6(10):963–72.
• Wiles S et al. Emergence of a 'hyperinfectious' bacterial state after passage of Citrobacter rodentium through the host gastrointestinal tract. Cell Microbiol 2005;7(8):1163–72.
• Wiles S et al. In vivo bioluminescence imaging of the murine pathogen Citrobacter rodentium. Infect Immun 2006;74(9):5391–6.
• Wiles S et al. Modelling infectious disease – time to think outside the box? Nat Rev Microbiol 2006;4(4):307–12.