scholarly journals Comparison of real-time PCR methods for the detection of Naegleria fowleri in surface water and sediment

2015 ◽  
Vol 114 (5) ◽  
pp. 1739-1746 ◽  
Author(s):  
Ashleigh Streby ◽  
Bonnie J. Mull ◽  
Karen Levy ◽  
Vincent R. Hill
Keyword(s):  
Surface Water ◽  
Real Time ◽  
Real Time Pcr ◽  
Naegleria Fowleri ◽  
Water And Sediment ◽  
Nægleria Fowleri

A real-time PCR diagnostic method for detection of Naegleria fowleri

2010 ◽  
Vol 126 (1) ◽  
pp. 37-41 ◽  
Author(s):  
Lucia Maďarová ◽  
Katarína Trnková ◽  
Soňa Feiková ◽  
Cyril Klement ◽  
Margita Obernauerová
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Diagnostic Method ◽  
Naegleria Fowleri ◽  
Nægleria Fowleri

A duplex real-time PCR assay for the quantitative detection of Naegleria fowleri in water samples

2007 ◽  
Vol 41 (1) ◽  
pp. 118-126 ◽  
Author(s):  
Jonas Behets ◽  
Priscilla Declerck ◽  
Yasmine Delaedt ◽  
Lieve Verelst ◽  
Frans Ollevier
Keyword(s):  
Real Time ◽  
Water Samples ◽  
Real Time Pcr ◽  
Quantitative Detection ◽  
Naegleria Fowleri ◽  
Pcr Assay ◽  
Nægleria Fowleri

Rapid and specific detection of Salmonella in water samples using real-time PCR and High Resolution Melt (HRM) curve analysis

2011 ◽  
Vol 64 (12) ◽  
pp. 2453-2459 ◽  
Author(s):  
G. N. van Blerk ◽  
L. Leibach ◽  
A. Mabunda ◽  
A. Chapman ◽  
D. Louw
Keyword(s):  
Surface Water ◽  
High Resolution ◽  
Real Time ◽  
Water Samples ◽  
Real Time Pcr ◽  
Curve Analysis ◽  
Pcr Assay ◽  
High Resolution Melt ◽  
Gene Target ◽  
Enrichment Step

A real-time PCR assay combined with a pre-enrichment step for the specific and rapid detection of Salmonella in water samples is described. Following amplification of the invA gene target, High Resolution Melt (HRM) curve analysis was used to discriminate between products formed and to positively identify invA amplification. The real-time PCR assay was evaluated for specificity and sensitivity. The assay displayed 100% specificity for Salmonella and combined with a 16–18 h non-selective pre-enrichment step, the assay proved to be highly sensitive with a detection limit of 1.0 CFU/ml for surface water samples. The detection assay also demonstrated a high intra-run and inter-run repeatability with very little variation in invA amplicon melting temperature. When applied to water samples received routinely by the laboratory, the assay showed the presence of Salmonella in particularly surface water and treated effluent samples. Using the HRM based assay, the time required for Salmonella detection was drastically shortened to less than 24 h compared to several days when using standard culturing methods. This assay provides a useful tool for routine water quality monitoring as well as for quick screening during disease outbreaks.

scholarly journals Detection of Bacterial Indicators and Human and Bovine Enteric Viruses in Surface Water and Groundwater Sources Potentially Impacted by Animal and Human Wastes in Lower Yakima Valley, Washington

2010 ◽  
Vol 77 (1) ◽  
pp. 355-362 ◽  
Author(s):  
Kristen E. Gibson ◽  
Kellogg J. Schwab
Keyword(s):  
Surface Water ◽  
Real Time ◽  
Real Time Pcr ◽  
Enteric Viruses ◽  
Bacterial Indicators ◽  
Tangential Flow ◽  
Surface Water And Groundwater ◽  
Human Wastes

ABSTRACTTangential flow ultrafiltration (UF) was used to concentrate and recover bacterial indicators and enteric viruses from 100 liters of groundwater (GW;n= 10) and surface water (SW;n= 11) samples collected in Lower Yakima Valley, WA. Human and bovine enteric viruses were analyzed in SW and GW concentrates by real-time PCR by using integrated inhibition detection.

scholarly journals Improved Method for the Detection and Quantification ofNaegleria fowleriin Water and Sediment Using Immunomagnetic Separation and Real-Time PCR

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Bonnie J. Mull ◽  
Jothikumar Narayanan ◽  
Vincent R. Hill
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Immunomagnetic Separation ◽  
Taqman Assay ◽  
Water And Sediment ◽  
Potential Risk Factors ◽  
Positive Direct ◽  
Free Living Ameba ◽  
Primary Amebic Meningoencephalitis ◽  
Detection And Quantification

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 ofN. fowleriand to evaluate potential risk factors for increased exposure. An immunomagnetic separation (IMS) procedure and real-time PCR TaqMan assay were developed to recover and quantifyN. fowleriin water and sediment samples. When one liter of lake water was seeded withN. fowleristrain 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 unknownN. fowlericoncentrations, resulting in positive direct detections by real-time PCR in 3 out of 16 samples and confirmation ofN. fowlericulture in 6 of 16 samples. This study has resulted in a new method for detection and quantification ofN. fowleriin 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 ofN. fowleriin environmental systems.

Recirculating Immunomagnetic Separation and Optimal Enrichment Conditions for Enhanced Detection and Recovery of Low Levels of Escherichia coli O157:H7 from Fresh Leafy Produce and Surface Water

2007 ◽  
Vol 70 (12) ◽  
pp. 2717-2724 ◽  
Author(s):  
SUNEE HIMATHONGKHAM ◽  
MARY LEE DODD ◽  
JENNY K. YEE ◽  
DAVID K. LAU ◽  
RAYMOND G. BRYANT ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Surface Water ◽  
Real Time ◽  
Real Time Pcr ◽  
Immunomagnetic Separation ◽  
Escherichia Coli O157 ◽  
Simple Method ◽  
E Coli ◽  
Low Levels ◽  
Spinach Leaves

The objective of this study was to develop a rapid, simple method for enhanced detection and isolation of low levels of Escherichia coli O157:H7 from leafy produce and surface water using recirculating immunomagnetic separation (RIMS) coupled with real-time PCR and a standard culture method. The optimal enrichment conditions for the method also were determined. Analysis of real-time PCR data (CT values) suggested that incubation of lettuce and spinach leaves rather than rinsates provides better enrichment of E. coli O157:H7. Enrichment of lettuce or spinach leaves at 42°C for 5 h provided better detection than enrichment at 37°C. Extended incubation of surface water for 20 h at 42°C did not improve the detection. The optimized enrichment conditions were also employed with modified Moore swabs, which were used to sample flowing water sites. Positive isolation rates and real-time PCR results indicated an increased recovery of E. coli O157:H7 from all samples following the application of RIMS. Under these conditions, the method provided detection and/or isolation of E. coli O157:H7 at levels as low as 0.07 CFU/g of lettuce, 0.1 CFU/g of spinach, 6 CFU/100 ml of surface water, and 9 CFU per modified Moore swab. During a 6-month field study, modified Moore swabs yielded high isolation rates when deployed in natural watershed sites. The method used in this study was effective for monitoring E. coli O157:H7 in the farm environment, during postharvest processing, and in foodborne outbreak investigations.

scholarly journals Recovery and Detection of Escherichia coli O157:H7 in Surface Water, Using Ultrafiltration and Real-Time PCR

2009 ◽  
Vol 75 (11) ◽  
pp. 3593-3597 ◽  
Author(s):  
Bonnie Mull ◽  
Vincent R. Hill
Keyword(s):  
Escherichia Coli ◽  
Surface Water ◽  
Real Time ◽  
Real Time Pcr ◽  
Membrane Filtration ◽  
Analytical Techniques ◽  
Immunomagnetic Separation ◽  
Treated Wastewater ◽  
Sample Collection ◽  
Multiple Pcr

ABSTRACT Enterohemorrhagic Escherichia coli O157:H7 (EHEC O157:H7) outbreaks have revealed the need for improved analytical techniques for environmental samples. Ultrafiltration (UF) is increasingly recognized as an effective procedure for concentrating and recovering microbes from large volumes of water and treated wastewater. This study describes the application of hollow-fiber UF as the primary step for concentrating EHEC O157:H7 seeded into 40-liter samples of surface water, followed by an established culture/immunomagnetic-separation (IMS) method and a suite of real-time PCR assays. Three TaqMan assays were used to detect the stx1, stx2, and rfbE gene targets. The results from this study indicate that approximately 50 EHEC O157:H7 cells can be consistently recovered from a 40-liter surface water sample and detected by culture and real-time PCR. Centrifugation was investigated and shown to be a viable alternative to membrane filtration in the secondary culture/IMS step when water quality limits the volume of water that can be processed by a filter. Using multiple PCR assay sets to detect rfbE, stx1, and stx2 genes allowed for specific detection of EHEC O157:H7 from strains that do not possess all three genes. The reported sample collection and analysis procedure should be a sensitive and effective tool for detecting EHEC O157:H7 in response to outbreaks of disease associated with contaminated water.

Sign in / Sign up

Export Citation Format

Share Document