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Development of a rapid DNA extraction method and one-step nested PCR for the detection of Naegleria fowleri from the
Arine Fadzlun Ahmad1, James Lonnen, Peter W Andrew
1Department of Infection, Immunity & Inflammation, University of Leicester, Medical Sciences Building, PO Box 138, University Road, Leicester LE1 9HN, UK.
Abstract:
Naegleria fowleri is a small free-living amoebo-flagellate found in natural and manmade thermal aquatic habitats worldwide. The organism is pathogenic to man causing fatal primary amoebic meningoencephalitis (PAM). Infection typically results from bathing in contaminated water and is usually fatal. It is, therefore, important to identify sites containing N. fowleri in the interests of preventive public health microbiology. Culture of environmental material is the conventional method for the isolation of N. fowleri but requires several days incubation and subsequent biochemical or molecular tests to confirm identification. Here, a nested one-step PCR test, in conjunction with a direct DNA extraction from water or sediment material, was developed for the rapid and reliable detection of N. fowleri from the environment. Here, the assay detected N, fowleri in 18/109 river water samples associated with a nuclear power plant in South West France and 0/10 from a similar site in the UK. Although culture of samples yielded numerous thermophilic free-living amoebae, none were N. fowleri or other thermophilic Naegleria spp. The availability of a rapid, reliable and sensitive one-step nested PCR method for the direct detection of N. fowleri from the environment may aid ecological studies and enable intervention to prevent PAM cases.
Insights
A new PCR test can rapidly detect Naegleria fowleri (N. fowleri), the cause of fatal primary amoebic meningoencephalitis (PAM). This method aids in identifying contaminated water sources for public health.
Area of Science:
- Environmental microbiology
- Molecular diagnostics
- Public health
Background:
- Naegleria fowleri is a pathogenic amoeba found in warm aquatic environments.
- Infection with N. fowleri causes primary amoebic meningoencephalitis (PAM), a usually fatal brain infection.
- Early detection of N. fowleri in water is crucial for public health interventions.
Purpose of the Study:
- To develop a rapid and reliable method for detecting N. fowleri in environmental samples.
- To establish a direct DNA extraction and nested one-step PCR assay for N. fowleri identification.
- To assess the prevalence of N. fowleri in specific aquatic environments.
Main Methods:
- Direct DNA extraction from water and sediment samples.
- Development of a nested one-step PCR assay for N. fowleri detection.
- Comparison of PCR results with conventional culture methods.
Main Results:
- The nested PCR assay successfully detected N. fowleri in 18 out of 109 river water samples from a nuclear power plant site in France.
- No N. fowleri was detected in 10 samples from a similar UK site.
- Conventional culture methods isolated various thermophilic amoebae but not N. fowleri or other thermophilic Naegleria species.
Conclusions:
- A rapid, sensitive, and reliable one-step nested PCR method for direct N. fowleri detection from environmental sources has been developed.
- This molecular tool can significantly aid ecological studies and public health efforts to prevent PAM.
- The PCR assay offers a faster alternative to traditional culture methods for identifying N. fowleri in water bodies.
