Presence and Distribution of Antibiotic Resistance
Genes in the Great Rivers of the Northern Adirondack
Region
Carrie Shaffer1
and Shane Rogers2
The overuse of antibiotics in human medicine and agriculture is a growing concern for public
health. Over 50 million pounds of antibiotics are used in the US each year. Greater than half of the US
antibiotic consumption is in the agricultural sector, for which a dominant fraction is for the promotion of
more rapid growth of livestock animals (Mellon and Fondriest, 2001). Overuse combined with
inadequate wastewater treatment has lead to the presence of antibiotic resistant bacteria and genes
encoding antibiotic resistance in surface waters, river sediments, and the feces of wild animals exposed to
waste residuals. Exposure to antibiotic resistant bacteria and the corresponding antibiotic resistance genes
(ARG) in surface waters can occur when the waters are used for drinking or recreation. Infection by
antibiotic-resistant strains of bacterial pathogens can have severe health implications for the sickened
individual including more virulent strains and less treatment options (Barza 2002).
Motivated by the increasing use of antibiotics and the public health concern that arises from this
overuse, the goals of this study are three fold. The first is to determine if land use surrounding the Grasse
and St Regis Rivers affects ARG levels in both the river sediment and water. A positive correlation
between agricultural land use, urban discharge sites and heightened levels of ARG would indicate that
source tracking markers can potentially serve as useful biomarkers for identifying the origin of the ARG
within the environment. The second objective strives to determine if relationships exist between hostspecific PCR biomarkers (human and ruminant), Salmonella sp., Fecal bacterial indicators (E. coli and/or
enterococci, qPCR methods for Bacteroidales and enterococci), and ARGs detected in river water and
1
2
Department of Biology, State University of New York, Potsdam
Assistant Professor, Civil and Environmental Engineering, Clarkson University
sediments of the Grasse and/or St. Regis rivers. Finally, this study attempts to investigate the distribution
of ARGs in feces of wild deer in agricultural, urban, and pristine land use areas of the greater northern
Adirondack Region and North Country of New York State that may imply that these animals can serve as
potential vectors of ARGs, to the great rivers of the Northern Adirondack region following potential
exposure to human and livestock wastes and wastewater residuals.
During the ten week course of this study, two rivers (Grasse River and St. Regis River) in the
Adirondack Mountains of New York were monitored for Salmonella spp., E. coli, Enterococci, hostspecific PCR biomarkers, and selected antibiotics on a weekly basis. Both rivers had a minimum of seven
distinct sites encompassing pristine areas with minimal anthropogenic influences, agricultural areas, and
urban areas where there was the potential to be affected by non point source wastewater discharge
contamination. Two liter water samples were composited from three discrete samples across the river
transect at each site then stored on ice until filtration (<8 hours). Water samples were taken as close as
possible to an ideal depth of 12 inches. Sediment samples were taken concurrently with the water samples
near the river banks and stored at -20C for DNA testing in a subsequent study.
Wild deer were investigated as vectors due in part to the abundance in the Northern Adirondack
Region, their herbivorous diet and frequent grazing in and near fields onto which livestock manures have
been applied, and their limited migratory range (commonly within one square mile) (Dewey, 2003). Deer
feces were collected at a variety of locations that were then classified as either urban, agricultural or
pristine areas. The fecal material was collected using sterile techniques then frozen until processing could
occur. Forty two distinct deer fecal samples were collected from agricultural, urban, and pristine areas.
Water samples were tested for E. coli and Enterococci using the Idexx ColiSure and Enterolert
methods. Five 300 mL water samples were also filtered onto 0.2 microliter pore size Supor membranes
using a vacuum filtration system. Four filters were frozen for later DNA extraction and quantitative real
time PCR measurements of fecal indicators, ARGs, and host-specific PCR biomarkers. The fifth filter
was used to determine the presence/absence of Salmonella spp. using enrichment and immunomagnetic
separation techniques as per manufacturer’s instructions (Dynal Dynabeads Salmonella, Invitrogen Corp.,
Carlsbad, CA), except that TaqMan qPCR was used to detect the presence of Salmonella spp. in
secondary enrichments rather than biochemical confirmation. DNA was extracted from the remaining
water filters using the MoBio PowerSoil kit as per manufacturer’s instructions, and qPCR with hydrolysis
probes were used to quantify fecal indicators and ARGs in the sample extracts.
The presence and distribution of ARGs, fecal indicators, and host-specific PCR biomarkers in the
river waters and deer feces will also be presented.
The results of this study will add to a growing body of literature on the presence and distribution
of antibiotic resistant genes as emerging contaminants in the environment, and provide critical
information on the potential for wildlife acting as vectors of these contaminants. The results will also be
useful for evaluating the efficacy of host-specific PCR biomarkers for fecal source allocation, an
important area of research that seeks to improve upon fecal indicator technologies by providing more
rapid and specific information to guide development of total maximum daily loads in impaired waters.
References
Barza, M. and K. Travers 2002. Excess infections due to antimicrobial resistance: the
“Attributable Fraction”, In: The need to Improve Antimicrobial Use in Agriculture: Ecological
and Human Health Consequences, M Barza and SL Gorbach (Eds.,), Clinical Infectious
Diseases, 34(S3): s126-s130
Dewey, T. and Animal Diversity Web Staff. 2003. "Odocoileus virginianus", Animal Diversity
Web. July 20, 2008
htttp://animaldiversity.ummz.umich.edu/site/accounts/information/Odocoileus_virginianus.html.
Mellon M., Fondriest S. 2001. Hogging It: Estimates of antimicrobial abuse in livestock. Union
of Concerned ScientistsMagazine. Vol 23: No.1