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The current knowledge about the epidemiology of Legionella spp. is based mainly on data gathered from studies of outbreaks. The standard method for the detection of Legionella in environmental samples is culture on BCYE agar supplemented with L-cysteine. However it is very time-consuming and fraught with limitations, since it cannot detect viable but non-culturable cells (VBNC). In other hand, PCR based-methods overestimate results as it detects both live and killed Legionella. Among the numerous methods described, the most promising for the future is FISH.
Nevertheless, the use of fluorescently-labeled probes limits the application of this technique to laboratories that either have an epifluorescent microscope or a flow cytometer in their facilities. 
Hence we have decided to develop an alternative ISH procedure for Legionella detection, using biotinylated NAM probes and an enzymatic complex as a reporter molecule. With this method development it is expected a direct and simple visualisation of the Legionella presence in water avoiding the need for samples manipulation such as DNA extraction and signal amplification required for PCR. In the future, this may allow the in loco water samples read-out by the use of a simple and portable optical detection equipment such as spectrophotometer.    

Laura Cerqueira
Laura CerqueiraJunior Researcherlcerqueira@fe.up.pt
Nuno Guimarães
Nuno GuimarãesJunior Researchernguimaraes@fe.up.pt
Nuno Azevedo
Nuno AzevedoSenior Researchernazevedo@fe.up.pt
Montse Vásquez
Montse VásquezResearch Fellowmontserratnv@gmail.com
Gonçalo Almeida
Gonçalo AlmeidaSenior Researchergoncalo.almeida@iniav.pt
Carina Almeida
Carina AlmeidaSenior Researchercarina.almeida@iniav.pt
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LEPABE
FEUP
FCT
Portugal 2020
Compete 2020
Norte 2020
União Europeia