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Improved Method for the Detection and Quantification of Naegleria fowleri in Water and Sediment Using Immunomagnetic Separation and Real-Time PCR.

Mull BJ, Narayanan J, Hill VR - J Parasitol Res (2013)

Bottom Line: Primary amebic meningoencephalitis (PAM) is a rare and typically fatal infection caused by the thermophilic free-living ameba, Naegleria fowleri.An immunomagnetic separation (IMS) procedure and real-time PCR TaqMan assay were developed to recover and quantify N. fowleri in water and sediment samples.The method was then applied to sediment and water samples with unknown N. fowleri concentrations, resulting in positive direct detections by real-time PCR in 3 out of 16 samples and confirmation of N. fowleri culture in 6 of 16 samples.

View Article: PubMed Central - PubMed

Affiliation: Division of Foodborne, Waterborne and Environmental Diseases, Centers for Disease Control and Prevention, National Center for Emerging and Zoonotic Infectious Diseases, 1600 Clifton Road, NE, Mail Stop D-66, Atlanta, GA 30329-4018, USA.

ABSTRACT
Primary amebic meningoencephalitis (PAM) is a rare and typically fatal infection caused by the thermophilic free-living ameba, Naegleria fowleri. In 2010, the first confirmed case of PAM acquired in Minnesota highlighted the need for improved detection and quantification methods in order to study the changing ecology of N. fowleri and to evaluate potential risk factors for increased exposure. An immunomagnetic separation (IMS) procedure and real-time PCR TaqMan assay were developed to recover and quantify N. fowleri in water and sediment samples. When one liter of lake water was seeded with N. fowleri strain CDC:V212, the method had an average recovery of 46% and detection limit of 14 amebas per liter of water. The method was then applied to sediment and water samples with unknown N. fowleri concentrations, resulting in positive direct detections by real-time PCR in 3 out of 16 samples and confirmation of N. fowleri culture in 6 of 16 samples. This study has resulted in a new method for detection and quantification of N. fowleri in water and sediment that should be a useful tool to facilitate studies of the physical, chemical, and biological factors associated with the presence and dynamics of N. fowleri in environmental systems.

No MeSH data available.


Related in: MedlinePlus

Sediment and water sample processing procedure.
© Copyright Policy - open-access
Related In: Results  -  Collection


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fig1: Sediment and water sample processing procedure.

Mentions: In August and October 2011, 1 L water and sediment samples were collected from 10 lakes in Minnesota and 6 lakes in Florida. Each water and sediment sample was a composite from 4 sites along a beach area at each lake. Sediment samples were washed twice with 500 mL of WB saline and the supernatant was processed using the same procedure as the water samples (Figure 1). Each sample was concentrated by centrifugation as already described, then 1 mL of the resulting pellet was placed on an agar culture plate and the remaining pellet volume was processed using the IMS procedure. The resulting IMS concentrate was split in half: 50 μL was added to an agar culture plate and 50 μL was extracted and assayed using real-time qPCR.


Improved Method for the Detection and Quantification of Naegleria fowleri in Water and Sediment Using Immunomagnetic Separation and Real-Time PCR.

Mull BJ, Narayanan J, Hill VR - J Parasitol Res (2013)

Sediment and water sample processing procedure.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3818898&req=5

fig1: Sediment and water sample processing procedure.
Mentions: In August and October 2011, 1 L water and sediment samples were collected from 10 lakes in Minnesota and 6 lakes in Florida. Each water and sediment sample was a composite from 4 sites along a beach area at each lake. Sediment samples were washed twice with 500 mL of WB saline and the supernatant was processed using the same procedure as the water samples (Figure 1). Each sample was concentrated by centrifugation as already described, then 1 mL of the resulting pellet was placed on an agar culture plate and the remaining pellet volume was processed using the IMS procedure. The resulting IMS concentrate was split in half: 50 μL was added to an agar culture plate and 50 μL was extracted and assayed using real-time qPCR.

Bottom Line: Primary amebic meningoencephalitis (PAM) is a rare and typically fatal infection caused by the thermophilic free-living ameba, Naegleria fowleri.An immunomagnetic separation (IMS) procedure and real-time PCR TaqMan assay were developed to recover and quantify N. fowleri in water and sediment samples.The method was then applied to sediment and water samples with unknown N. fowleri concentrations, resulting in positive direct detections by real-time PCR in 3 out of 16 samples and confirmation of N. fowleri culture in 6 of 16 samples.

View Article: PubMed Central - PubMed

Affiliation: Division of Foodborne, Waterborne and Environmental Diseases, Centers for Disease Control and Prevention, National Center for Emerging and Zoonotic Infectious Diseases, 1600 Clifton Road, NE, Mail Stop D-66, Atlanta, GA 30329-4018, USA.

ABSTRACT
Primary amebic meningoencephalitis (PAM) is a rare and typically fatal infection caused by the thermophilic free-living ameba, Naegleria fowleri. In 2010, the first confirmed case of PAM acquired in Minnesota highlighted the need for improved detection and quantification methods in order to study the changing ecology of N. fowleri and to evaluate potential risk factors for increased exposure. An immunomagnetic separation (IMS) procedure and real-time PCR TaqMan assay were developed to recover and quantify N. fowleri in water and sediment samples. When one liter of lake water was seeded with N. fowleri strain CDC:V212, the method had an average recovery of 46% and detection limit of 14 amebas per liter of water. The method was then applied to sediment and water samples with unknown N. fowleri concentrations, resulting in positive direct detections by real-time PCR in 3 out of 16 samples and confirmation of N. fowleri culture in 6 of 16 samples. This study has resulted in a new method for detection and quantification of N. fowleri in water and sediment that should be a useful tool to facilitate studies of the physical, chemical, and biological factors associated with the presence and dynamics of N. fowleri in environmental systems.

No MeSH data available.


Related in: MedlinePlus