Universal mitochondrial PCR combined with species-specific dot-blot assay as a source-tracking method of human, bovine, chicken, ovine, and porcine in fecal-contaminated surface water

Kortbaoui, Randa; Locas, Annie; Imbeau, Marianne; Payment, Pierre; Villemur, Richard

doi:10.1016/j.watres.2009.01.030

Cited by 35 publications

(24 citation statements)

References 35 publications

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“…Most research on the fate of aqueous macrobial eDNA has not considered settling (Martellini, Payment & Villemur 2005;Kortbaoui et al 2009;Dejean et al 2011;Thomsen et al 2012a), except one experiment that applied water circulation to eliminate it (Thomsen et al 2012b). Our findings emphasize that degradation of DNA molecules and particle settling combine to reduce aqueous eDNA concentration over time, and future research should try to measure both processes.…”

Section: S T a T E A N D F A T E O F A Q U E O U S M A C R O B I A L mentioning

confidence: 83%

Particle size distribution and optimal capture of aqueous macrobial eDNA

et al. 2014

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Summary1. Using environmental DNA (eDNA) to detect aquatic macroorganisms is a new survey method with broad applicability. However, the origin, state and fate of aqueous macrobial eDNA -which collectively determine how well eDNA can serve as a proxy for directly observing organisms and how eDNA should be captured, purified and assayed -are poorly understood. 2. The size of aquatic particles provides clues about their origin, state and fate. We used sequential filtration size fractionation to measure the particle size distribution (PSD) of macrobial eDNA, specifically Common Carp (hereafter referred to as Carp) eDNA. We compared it to the PSDs of total eDNA (from all organisms) and suspended particle matter (SPM). We quantified Carp mitochondrial eDNA using a custom qPCR assay, total eDNA with fluorometry and SPM with gravimetric analysis. 3. In a lake and a pond, we found Carp eDNA in particles from >180 to <0Á2 lm, but it was most abundant from 1 to 10 lm. Total eDNA was most abundant below 0Á2 lm, and SPM was most abundant above 100 lm. SPM consisted of ≤0Á1% total eDNA, and total eDNA consisted of ≤0Á0004% Carp eDNA. 0Á2 lm filtration maximized Carp eDNA capture (85% AE 6%) while minimizing total (i.e. non-target) eDNA capture (48% AE 3%), but filter clogging limited this pore size to a sample volume <250 mL. To mitigate this limitation, we estimated a continuous PSD model for Carp eDNA and derived an equation for calculating isoclines of pore size and water volume that yield equivalent amounts of Carp eDNA. 4. Our results suggest that aqueous macrobial eDNA predominantly exists inside mitochondria or cells, and that settling may therefore play an important role in its fate. For optimal eDNA capture, we recommend 0Á2 lm filtration or a combination of larger pore size and water volume that exceeds the 0Á2 lm isocline. In situ filtration of large volumes could maximize detection probability when surveying large habitats for rare organisms. Our method for eDNA particle size analysis enables future research to compare the PSDs of eDNA from other organisms and environments, and to easily apply them for ecological monitoring.

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Section: S T a T E A N D F A T E O F A Q U E O U S M A C R O B I A L mentioning

confidence: 83%

Particle size distribution and optimal capture of aqueous macrobial eDNA

et al. 2014

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“…The other human marker (HF183) was found in 50% of the samples. Poultry and ovine host-specific mtDNAs were also targeted (Kortbaoui et al, 2009), but generated very few (chicken, ca. 3%) or no (ovine) signals (data not shown).…”

Section: Occurrence and Concentration Of The Fst Markers In Surface Wmentioning

confidence: 99%

“…There are numerous studies that used different FST markers to assess the impact of human activities on the level of fecal contamination in watersheds, which crossed natural, recreational, urban, suburban, rural/agricultural areas. However, only a few field studies have been reported for the detection of mtDNA in watersheds (Baker-Austin et al, 2010;Kapoor et al, 2013;Kortbaoui et al, 2009;Martellini et al, 2005;Tambalo et al, 2012a;Vuong et al, 2013). Such field studies are essential if these new markers are to be accepted as valuable FST markers.…”

Section: Introductionmentioning

confidence: 99%

An environmental survey of surface waters using mitochondrial DNA from human, bovine and porcine origin as fecal source tracking markers

et al. 2015

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“…Moreover, the mtDNA sequences of most species are publicly available in several DNA databases (Caldwell et al, 2011). Some mtDNA-based methods have been developed to identify the sources of fecal contamination, including single/multiplex PCR (Martellini et al, 2005), nested PCR (Kortbaoui et al, 2009), quantitative real-time PCR (qPCR) (Caldwell and Levine, 2009;Caldwell et al, 2007;Schill and Mathes, 2008) and mtDNA microarray (Nguyet-Minh et al, 2013).…”

Section: Introductionmentioning

confidence: 99%