Legionnaires' disease outbreak investigation toolbox

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Microbiology: Legionella laboratory diagnosis and detection

Good microbiological analysis enables investigators to identify and epidemiologically link Legionella cases to an environmental source. It is important, wherever possible, that confirmatory testing is performed, ideally in accredited reference laboratories with experience of detecting Legionella, to provide a high level of confidence. There are many detection methods and many variations of each method. The practical details of performing each of these methods is beyond the scope of this toolbox, but instead a brief summary is provided to inform judgement about the pros and cons of each type of test methodology that might be useful when responding to an outbreak. There are many peer-reviewed references in academic journals which contain the practical details for each method and their variations as well as a number of review articles (for example [1]).

There are essentially five types of microbiological methodology that can be applied to detect and identify Legionella; culture; antibody-based, DNA-based, serology and fluorescence staining. The choice of which technique to use is based upon sample type and availability, experience and further preference which may be guided by the need to fulfil regulatory or quality requirements. The gold standard however remains culture, which as well as confirming the presence of Legionella in a sample, also provides a supply of material to do a more in depth analysis down to individual strain level if necessary. Increasingly, there are DNA amplification-based techniques which allow a similar level of discrimination but these are, at present, technically more complex and therefore have less widespread use.

It is important to note that detection of Legionella in a particular environmental sample does not, in itself, necessarily prove that the site from which this sample was taken was responsible for the outbreak. It does however allow the reasonably quick elimination of sites from the investigation, if there is confidence that the site was examined appropriately and that all samples from that site were negative. However, if the Legionella in an environmental sample can be "matched" with Legionella found in a clinical sample from one of the patients involved in the outbreak then this provides strong evidence for a causative link. To be confident that the Legionella from the two samples "match" from a microbiological point of view, samples would need to undergo additional microbiological identification and typing following initial detection, to either confirm or discriminate between them. The choice of techniques and the level of confidence required is, as mentioned previously, driven by a number of factors. Several molecular subtyping techniques are in use to subtype L. pneumophila strains in epidemiological investigations e.g Sequence Based Typing method (SBT) (for more information please visit http://www.ewgli.org/). Other epidemiological subtyping tools include, monoclonal antibody typing, macrorestriction analysis (MRA) and amplified fragment length polymorphism (AFLP) typing [2].

The current techniques available for analysing clinical and environmental samples and, where appropriate, the level of discrimination that they provide are discussed on their respective pages.

  1. BARTRAM J. (2007) Legionella and the prevention of legionellosis WHO, Geneva ISBN 92 4 156297 http pdf
  2. HEUNER K. & SWANSON M. (2008) Legionella: Molecular Microbiology ISBN: 978-1-904455-26-4 http