We tested 772 cloacal swabs from wild ducks to estimate the prevalence of enteric bacteria resistant to extended-spectrum cephalosporins (ESC). We found a low prevalence of the important ESC resistance genotypes, blaCMY (5.7%) and blaCTX-M (0.3%). This suggests a minor role for wild ducks in the movement of resistant bacteria in the environment.

Antimicrobial resistance is of critical importance, with significant negative consequences for health care (Razazi et al. 2012). Resistance to extended-spectrum cephalosporins (ESC) has been identified as a serious threat to human health (Centers for Disease Control 2013). Livestock have been implicated in the transmission of ESC-resistant bacteria to humans via food-borne transmission (Mora et al. 2010; López-Cerero et al. 2011). Livestock and food products can harbor bacteria with ESC resistance, primarily from plasmid-mediated β-lactamase genes conferring resistance to ESC, including the AmpC blaCMY and the extended spectrum β-lactamase blaCTX-M (Mollenkopf et al. 2014; Davis et al. 2015).

Wild ducks have been associated with the transmission of livestock pathogens and Shiga-toxigenic Escherichia coli (Ewers et al. 2009; Kim et al. 2009). In Europe and Asia, wild waterfowl have been shown to harbor ESC-resistant bacteria in their gastrointestinal (GI) flora (Guenther et al. 2010; Veldman et al. 2013; Mohsin et al. 2016). However, little is known of their role in the transmission or movement of ESC-resistant bacteria in the US. Wild ducks have direct contact with water ecosystems, potentially contaminating many environments and populations. We investigated wild ducks as a reservoir for ESC-resistant Enterobacteriaceae. Specifically, we determined the prevalence of the AmpC (blaCMY) and extended-spectrum β-lactamase (blaCTX-M) resistance genes, which are plasmid-mediated genes that can readily move between bacterial species via horizontal gene transfer (Winokur et al. 2000).

Ducks were caught during routine waterfowl surveillance in conjunction with the Ohio Department of Natural Resources throughout Ohio and in conjunction with Winous Point Marsh Conservancy along the shoreline of Lake Erie in Ohio from July 2014–April 2015. Cloacal swabs were obtained with sterile Stuart's liquid media transport swabs (Becton, Dickinson and Company, Franklin Lakes, New Jersey, USA). Birds were banded with an aluminum band and released. Additionally, cloacal swabs were obtained from hunter-harvested ducks throughout the 2014–15 hunting season during routine surveillance by the Ohio Department of Natural Resources, Winous Point Marsh Conservancy, and by a field technician along the Mississippi flyway in Illinois, Wisconsin, Mississippi, Arkansas, Iowa, and Missouri.

Cloacal swabs were appropriately stored for up to 2 wk and transported to the laboratory at ambient temperature. Swabs were incubated overnight in 10 mL of buffered peptone water (Becton, Dickinson and Company); a 1-mL aliquot was then incubated overnight in 9 mL of MacConkey broth (Becton, Dickinson and Company) modified with 2 μg/mL of cefotaxime (TCI America, Portland, Oregon, USA). MacConkey broth was inoculated onto MacConkey agar (EMD Millipore, Darmstadt, Germany) modified with 8 μg/mL cefoxitin (CHEM-IMPEX INT'L, Wood Dale, Illinois, USA), 4 μg/mL cefepime (CHEM-IMPEX), or 1 μg/mL meropenem (US Pharmacopeial Convention, Rockville, Maryland, USA) to identify blaCMY, blaCTX-M, and carbapenem-resistant phenotypes, respectively, and incubated overnight. Up to three lactose fermenting, indole-positive isolates representing distinct morphologies were presumed to be E. coli and were selected for further characterization from each selective agar. For Salmonella screening, a 100-μL aliquot of buffered peptone water was incubated overnight in 10 mL of Rappaport Vassiliadis broth (Becton, Dickinson and Company), then inoculated onto XLT4 agar (Becton, Dickinson and Company) and incubated overnight. A single, characteristic black colony was selected and grown on nonselective MacConkey agar. A nonlactose-fermenting colony was selected for Salmonella polyvalent O antisera agglutination and inoculated onto a Triple Sugar Iron agar (Becton, Dickinson and Company).

Identification of blaCMY and blaCTX-M was performed using standard PCR techniques with primers previously reported (Mollenkopf et al. 2012). We bidirectionally Sanger-sequenced blaCTX-M genes using the corresponding PCR amplification primers and analyzed them using BLAST (NCBI 2016).

A total of 772 wild ducks were sampled in July 2014–April 2015; 278 were live caught, and 494 were hunter harvested. Samples were collected from 14 different sampling sites in seven states; none of which were in close proximity to any large metropolitan areas (Table 1). Samples were collected from ducks representing 15 different Anseriformes species. Among hunter-harvested ducks, 384 were from Ohio, 38 from Illinois, 22 from Wisconsin, 15 from Mississippi, 16 from Arkansas, 11 from Iowa, and eight from Missouri (Table 1). The most common hunter-harvested ducks sampled were Mallards (Anas platyrhynchos), Green-winged Teal (Anas carolinensis), and Blue-winged Teal (Anas discors). Among live-caught ducks, all sampled in Ohio, the most common ducks sampled were Mallards, Wood Ducks (Aix sponsa), and Redheads (Aythya americana). There were 44 ducks that harbored blaCMY in their GI flora, representing 5.7% of the population: 10.1% of the live-caught ducks and 3.2% of the hunter-harvested ducks (Table 1). These 44 blaCMY harboring ducks were sampled from four states; 37 from Ohio, four from Illinois, two from Wisconsin, and one from Missouri; 9.9% were sampled in the warm season (April–September), and 2.5% were from the cold season (October–March; Tables 1, 2). Among the 44 ducks that harbored blaCMY, 23 were Mallards, nine were Wood Ducks, seven were Blue-wing Teal, two each were Gadwall (Anas strepera) and Green-wing Teal, and one was a Northern Pintail (Anas acuta). Only two ducks, a Mallard and a Northern Shoveler (Anas clypeata), harbored blaCTX-M in their GI flora, representing 0.3% of the population; both were hunter-harvested ducks from Ohio in the cold season (Tables 1, 2). Both blaCTX-M genes were identified as blaCTX-M-15, the most common extended-spectrum β-lactamase found in humans, and occasionally reported in food-producing animal populations (Watson et al. 2012; Chen et al. 2014). Six wild ducks, all of which were hunter harvested from Ohio, were culture positive for Salmonella spp., representing 0.8% of the sampled population; none of which were resistant to ESC.

Table 1

Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing hunter-harvested and live-caught ducks.

Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing hunter-harvested and live-caught ducks.
Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing hunter-harvested and live-caught ducks.
Table 2

Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing ducks sampled in the warm and cold seasons.

Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing ducks sampled in the warm and cold seasons.
Number (prevalence) of wild ducks with enteric bacteria harboring blaCMY and blaCTX-M resistance genes sampled from July 2014–April 2015 along the central flyway in the USA, comparing ducks sampled in the warm and cold seasons.

Our results indicated that wild ducks can harbor clinically relevant antimicrobial-resistance genes in their GI flora. Many ducks are migratory and can travel great distances, leading to the potential for regional or international movement of these resistance genes, as documented with other pathogens (Gaidet et al. 2008). Although ducks have limited direct contact with livestock species, they have been reported to transmit infectious pathogens to food animal populations (Kim et al. 2009), where they can be introduced into the food supply. Given the mobility of ducks, resistance genes can be disseminated to multiple waterways with the potential for direct human contact via recreational water use or to livestock facilities. This type of community-acquired infection can be important because 78% of Canadian extended-spectrum β-lactamase-producing E. coli bloodstream infections reportedly originated in the community (Peirano et al. 2012). Wildlife, especially birds, which can have a role in community exposure by contaminating aquatic ecosystems, represents a potential source of transmission to livestock species and is a direct food source when hunter harvested. Food animal populations have much higher levels of blaCMY and blaCTX-M, most likely because of the high level of antimicrobial selection pressure in those populations. In the US, 70% of dairies have been shown to have >80% of their cows harboring blaCMY, and 76% of dairies have >40% of their cows harboring blaCTX-M (Davis et al. 2015). Hospitalized human populations, however, have a similar prevalence of blaCMY in their enteric flora (10%) and a higher prevalence of blaCTX-M (3%; Landers et al. 2016). However, because of the lower observed prevalence of ESC-resistance genes in this study, the role of wild ducks in the dissemination of clinically relevant, antibiotic-resistant bacteria and resistance genes may be limited, relative to other reservoirs of antimicrobial resistance.

We would like to thank Brendan Shirkey at Winous Point Conservancy and Brian Tucker for their help in live duck sampling and collection of hunter-harvested duck cloacal swabs.

Centers for Disease Control and Prevention
.
2013
.
Antibiotic resistance threats in the United States. http://www.cdc.gov/drugresistance/threat-report-2013. Accessed September 2016
.
Chen
LF,
Freeman
JT,
Nicholson
B,
Keiger
A,
Lancaster
S,
Joyce
M,
Woods
CW,
Cook
E,
Adcock
L,
Louis
S,
et al.
2014
.
Widespread dissemination of CTX-M-15 genotype extended-spectrum-β-lactamase-producing Enterobacteriaceae among patients presenting to community hospitals in the southeastern United States
.
Antimicrob Agents Chemother
58
:
1200
1202
.
Davis
MA,
Sischo
WM,
Jones
LP,
Moore
DA,
Ahmed
S,
Short
DM,
Besser
TE.
2015
.
Recent emergence of Escherichia coli with cephalosporin resistance conferred by blaCTX-M on Washington State dairy farms
.
Appl Environ Microbiol
81
:
4403
4410
.
Ewers
C,
Guenther
S,
Wieler
LH,
Schierack
P.
2009
.
Mallard ducks—A waterfowl species with high risk of distributing Escherichia coli pathogenic for humans
.
Env Microbiol Rep
1
:
510
517
.
Gaidet
N,
Newman
SH,
Hagemeijer
W,
Dodman
T,
Cappelle
J,
Hammoumi
S,
De Simone
L,
Takekawa
JY.
2008
.
Duck migration and past influenza A (H5N1) outbreak areas
.
Emerg Infect Dis
14
:
1164
1166
.
Guenther
S,
Grobbel
M,
Beutlich
J,
Bethe
A,
Friedrich
ND,
Goedecke
A,
Lübke-Becker
A,
Guerra
B,
Wieler
LH,
Ewers
C.
2010
.
CTX-M-15-type extended-spectrum beta-lactamases-producing Escherichia coli from wild birds in Germany
.
Env Microbiol Rep
2
:
641
645
.
Kim
JK,
Negovetich
NJ,
Forrest
HL,
Webster
RG.
2009
.
Ducks: The “Trojan horses” of H5N1 influenza
.
Influenza Other Respir Viruses
3
:
121
128
.
Landers
TF,
Mollenkopf
DF,
Faubel
RL,
Dent
A,
Pancholi
P,
Daniels
JB,
Wittum
TE.
2016
.
Extended spectrum β-lactam resistance in the enteric flora of patients at a tertiary care medical centre
.
Zoonoses Public Health
64
:
161
164
.
López-Cerero
L,
Egea
P,
Serrano
L,
Navarro
D,
Mora
A,
Blanco
J,
Doi
Y,
Paterson
DL,
Rodríguez-Baño
J,
Pascual
A.
2011
.
Characterisation of clinical and food animal Escherichia coli isolates producing CTX-M-15 extended-spectrum β-lactamase belonging to ST410 phylogroup A
.
Int J Antimicrob Agents
37
:
365
367
.
Mohsin
M,
Raza
S,
Roschanski
N,
Schaufler
K,
Guenther
S.
2016
.
First description of plasmid-mediated colistin-resistant extended-spectrum β-lactamase-producing Escherichia coli in a wild migratory bird from Asia
.
Int J Antimicrob Agents
48
:
463
464
.
Mollenkopf
DF,
Cenera
JK,
Bryant
EM,
King
CA,
Kashoma
I,
Kumar
A,
Funk
JA,
Rajashekara
G,
Wittum
TE.
2014
.
Organic or antibiotic-free labeling does not impact the recovery of enteric pathogens and antimicrobial-resistant Escherichia coli from fresh retail chicken
.
Foodborne Pathog Dis
11
:
920
929
.
Mollenkopf
DF,
Weeman
MF,
Daniels
JB,
Abley
MJ,
Mathews
JL,
Gebreyes
WA,
Wittum
TE.
2012
.
Variable within- and between-herd diversity of CTX-M cephalosporinase-bearing Escherichia coli isolates from dairy cattle
.
Appl Environ Microbiol
78
:
4552
4560
.
Mora
A,
Herrera
A,
Mamani
R,
López
C,
Alonso
MP,
Blanco
JE,
Blanco
M,
Dahbi
G,
Garcia-Garrote
F,
Pita
JM,
et al.
2010
.
Recent emergence of clonal group O25b:K1:H4-B2-ST131 ibeA strains among Escherichia coli poultry isolates, including CTX-M-9-producing strains, and comparison with clinical human isolates
.
Appl Environ Microbiol
76
:
6991
6997
.
NCBI (National Center for Biotechnology Information)
.
2016
.
Basic local alignment search tool (BLAST)
.
Peirano
G,
Van Der Bij
AK,
Gregson
DB,
Pitout
JDD.
2012
.
Molecular epidemiology over an 11-year period (2000 to 2010) of extended-spectrum β-lactamase-producing Escherichia coli causing bacteremia in a centralized Canadian region
.
J Clin Microbiol
50
:
294
299
.
Razazi
K,
Derde
LPG,
Verachten
M,
Legrand
P,
Lesprit
P,
Brun-Buisson
C.
2012
.
Clinical impact and risk factors for colonization with extended-spectrum β-lactamase-producing bacteria in the intensive care unit
.
Intensive Care Med
38
:
1769
1778
.
Veldman
K,
Van Tulden
P,
Kant
A,
Testerink
J,
Mevius
D.
2013
.
Characteristics of cefotaxime-resistant Escherichia coli from wild birds in The Netherlands
.
Appl Environ Microbiol
79
:
7556
7561
.
Watson
E,
Jeckel
S,
Snow
L,
Stubbs
R,
Teale
C,
Wearing
H,
Horton
R,
Toszeghy
M,
Tearne
O,
Ellis-Iversen
J,
et al.
2012
.
Epidemiology of extended spectrum beta-lactamase E. coli (CTX-M-15) on a commercial dairy farm
.
Vet Microbiol
154
:
339
346
.
Winokur
PL,
Brueggemann
A,
Desalvo
DL,
Hoffmann
L,
Apley
MD,
Uhlenhopp
EK,
Pfaller
MA,
Doern
GV.
2000
.
Animal and human multidrug-resistant, cephalosporin-resistant Salmonella isolates expressing a plasmid-mediated CMY-2 AmpC β-lactamase
.
Antimicrob Agents Chemother
44
:
2777
2783
.