Detection of the free living amoeba Naegleria fowleri by using conventional and real-time PCR based on a single copy DNA sequence

• Detection of N. fowleri by conventional and qPCR based on a single copy DNA region.
• One copy corresponds to one N. fowleri cell.
• The qPCR results give a detection limit of 1 copy per reaction.
• The procedure method does not require a Taqman probe.

The amoeba-flagellate Naegleria fowleri is a causative agent of primary amoebic meningoencephalitis (PAM). This thermophilic species occurs worldwide and tends to proliferate in warm aquatic environment. The PAM cases remain rare but this infection is mostly fatal. Here, we describe a single copy region which has been cloned and sequenced, and was used for both conventional and real-time PCR. Targeting a single-copy DNA sequence allows to directly quantify the N. fowleri cells. The real-time PCR results give a detection limit of 1 copy per reaction with high reproducibility without the need of a Taqman probe. This procedure is of interest as compared to other procedures which are mostly based on the detection of multi-copy DNA associated with a Taqman probe.

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