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 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

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.

Monitoring bacterial indicators and pathogens in cattle feedlot waste by real-time PCR

2010 ◽  
Vol 44 (5) ◽  
pp. 1381-1388 ◽  
Author(s):  
Marcus Klein ◽  
Leearna Brown ◽  
Ben van den Akker ◽  
Gregory M. Peters ◽  
Richard M. Stuetz ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Bacterial Indicators ◽  
Cattle Feedlot

A probe-free four-tube real-time PCR assay for simultaneous detection of twelve enteric viruses and bacteria

2015 ◽  
Vol 118 ◽  
pp. 93-98 ◽  
Author(s):  
Chen Zhang ◽  
Peihua Niu ◽  
Yanying Hong ◽  
Ji Wang ◽  
Jingyun Zhang ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Simultaneous Detection ◽  
Enteric Viruses ◽  
Pcr Assay

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.

scholarly journals Quantitative Detection of Human Adenoviruses in Wastewater and Combined Sewer Overflows Influencing a Michigan River

2009 ◽  
Vol 76 (3) ◽  
pp. 715-723 ◽  
Author(s):  
Theng-Theng Fong ◽  
Mantha S. Phanikumar ◽  
Irene Xagoraraki ◽  
Joan B. Rose
Keyword(s):  
Wastewater Treatment ◽  
Surface Water ◽  
Real Time ◽  
Surface Waters ◽  
Wastewater Treatment Plant ◽  
Real Time Pcr ◽  
Treatment Plant ◽  
Combined Sewer Overflows ◽  
Treated Effluent ◽  
Combined Sewer

ABSTRACT Enteric viruses are important pathogens found in contaminated surface waters and have previously been detected in waters of the Great Lakes. Human adenoviruses were monitored because of their high prevalence and persistence in aquatic environments. In this study, we quantified adenoviruses in wastewater, surface water, and combined sewer overflows (CSOs) by real-time PCR. Between August 2005 and August 2006, adenovirus concentrations in raw sewage, primary-treated effluent, secondary-treated effluent, and chlorinated effluent from a wastewater treatment plant in Michigan were examined. CSO samples (n = 6) were collected from a CSO retention basin in Grand Rapids, MI. Adenoviruses were detected in 100% of wastewater and CSO discharge samples. Average adenovirus DNA concentrations in sewage and CSOs were 1.15 × 106 viruses/liter and 5.35 × 105 viruses/liter, respectively. Adenovirus removal was <2 log10 (99%) at the wastewater treatment plant. Adenovirus type 41 (60% of clones), type 12 (29%), type 40 (3%), type 2 (3%), and type 3 (3%) were isolated from raw sewage and primary effluents (n = 28). Six of 20 surface water samples from recreational parks at the lower Grand River showed virus concentrations above the real-time PCR detection limit (average, 7.8 × 103 viruses/liter). This research demonstrates that wastewater effluents and wastewater-impacted surface waters in the lower Grand River in Michigan contain high levels of viruses and may not be suitable for full-body recreational activities. High concentrations of adenovirus in these waters may be due to inefficient removal during wastewater treatment and to the high persistence of these viruses in the environment.

scholarly journals Detection of Giardia intestinalis in water samples collected from natural water reservoirs and wells in northern and north-eastern Poland using LAMP, real-time PCR and nested PCR

2017 ◽  
Vol 15 (5) ◽  
pp. 775-787 ◽  
Author(s):  
Anna Lass ◽  
Beata Szostakowska ◽  
Krzysztof Korzeniewski ◽  
Panagiotis Karanis
Keyword(s):  
Surface Water ◽  
Real Time ◽  
Water Samples ◽  
Real Time Pcr ◽  
Nested Pcr ◽  
Lamp Assay ◽  
Giardia Intestinalis ◽  
Detection Methods ◽  
Rrna Gene ◽  
Water Reservoirs

Giardia intestinalis is a protozoan parasite, transmitted to humans and animals by the faecal–oral route, mainly through contaminated water and food. Knowledge about the distribution of this parasite in surface water in Poland is fragmentary and incomplete. Accordingly, 36 environmental water samples taken from surface water reservoirs and wells were collected in Pomerania and Warmia-Masuria provinces, Poland. The 50 L samples were filtered and subsequently analysed with three molecular detection methods: loop-mediated isothermal amplification (LAMP), real-time polymerase chain reaction (real-time PCR) and nested PCR. Of the samples examined, Giardia DNA was found in 15 (42%) samples with the use of LAMP; in 12 (33%) of these samples, Giardia DNA from this parasite was also detected using real-time PCR; and in 9 (25%) using nested PCR. Sequencing of selected positive samples confirmed that the PCR products were fragments of the Giardia intestinalis small subunit rRNA gene. Genotyping using multiplex real-time PCR indicated the presence of assemblages A and B, with the latter predominating. The results indicate that surface water in Poland, as well as water taken from surface wells, may be a source of Giardia strains which are potentially pathogenic for humans. It was also demonstrated that LAMP assay is more sensitive than the other two molecular assays.

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