IJERD – International Journal of Environmental and Rural Development (2023) 14-2

Research article
erdAntimicrobial Resistance Profiles Found in a Case Study ofEscherichia coli from Cohabitant Pets and Environmental
Surfaces at Animal Clinics, Phnom Penh CityBAN NAIHEAK*
Faculty of Veterinary Medicine, Royal University of Agriculture, Cambodia
Email: naiheakban@gmail.com

CHEA BUNNA
Faculty of Veterinary Medicine, Royal University of Agriculture, Cambodia

VENN VUTEY
Faculty of Veterinary Medicine, Royal University of Agriculture, Cambodia

KANG KROESNA
Faculty of Veterinary Medicine, Royal University of Agriculture, Cambodia
Received 20 November 2023

Accepted 22 December 2023(*Corresponding Author)

Abstract Escherichia coli (E. coli) can be transmitted from dogs to surfaces through direct
exposure to dog faeces, pus, and urine; alternatively, it can live in the environment for a shortperiod. We aimed to determine the prevalence and antibiotic resistance profile of E. coli fromdogs and environmental surfaces of animal clinics in Phnom Penh, Cambodia. We swabbed
twenty-four samples from dogs (18 faeces and 6 pus) and twenty-three from environmental
surfaces (8 cage-floors, 7 cage-walls, 1 feeding-plate, and 7 treatment-tables) at animal clinics.
The bacterial culture method was used to isolate E. coli from both dog and environmental
surface samples. An antibiogram of the isolates was tested using the disk diffusion method with
six antibiotics (amoxicillin, ampicillin, ceftriaxone, gentamycin, levofloxacin, andtetracycline), following the standard of the Clinical and Laboratory Standard Institute. The
results showed that E. coli was present in approximately 54% of dog samples and 22% ofenvironmental surface samples. The E. coli isolates from dog samples showed high resistanceto amoxicillin, ampicillin, and tetracycline (54%) and lower resistance to ceftriaxone, gentamycin(27%), and levofloxacin (8%). E. coli isolates from environmental surfaces demonstrated highresistance to amoxicillin, ampicillin, ceftriaxone, and tetracycline (80%-100%), and lower
resistance to gentamicin and levofloxacin (60%). In conclusion, E. coli was found in both dogs
and veterinary clinic surfaces and exhibited resistance to numerous antibiotics. This study
suggests that further research is necessary to identify the specific genes responsible for theantibiotic resistance of E. coli found in dogs, on clinic surfaces, as well as in humans –including clinic technicians and dog owners – in order to better understand how this resistanceis affected by the transmission of E. coli between different carriers.
Keywords dogs, environmental surfaces, E. coli, antibiotic resistance, antibiogram
INTRODUCTION
The use of antimicrobial drugs does not only increase the antimicrobial resistance (AMR) of
pathogenic bacteria but also the endogenous commensal flora. (Berge et al., 2006; Dancer, 2004;
Goosens, 2009). It is reported that the potential problem of AMR among pet animals may cause
human health problems, due to the increasing utilization of the same antimicrobial substances inhuman medicine and to the close contact between pets and their human cohabitants (Guardabassi etal., 2004; Moyaert et al., 2006; Schwarz et al., 2001).

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The spread of antimicrobial-resistant bacteria can occur directly, by skin-to-skin contact andcontact with bacteria-containing material (e.g., saliva, feces, urine), or indirectly via the household
environment (Guardabassi et al., 2004; Schwarz et al., 2001). Leite et al. (2013) reported that direct,
close contact between all the cohabitants including pets and humans, and the touch of contaminatedhousehold surfaces and objects could lead to shared antimicrobial-resistant bacteria. In the new host,resistant bacteria can both colonize and infect and remain in that particular environment for a shorttime. During this period, the resistant bacteria are capable of either spreading their resistance genes
to other bacteria residing in the new host (endogenous or pathogen bacteria), but also accepting
resistance genes from other bacteria (Livermore, 2003; Schwarz et al., 2001). E. coli has possible
biology in spreading resistance not only to the acceptor and donor of the transmissible drug-resistantgene but also commonly found in the intestinal tract of both humans and animals (Costa et al., 2008).
E. coli can also be implicated in various intestinal and extra-intestinal diseases including urinary tract
infection (Johnson et al., 2008; Johnson et al., 2001) and Pyometra (Bassessar et al., 2013). Likewise,
Markey et al. (2013) showed that E. coli can cause diseases such as Colisepticaemia, Pyometra, andUrinary tract infections in dogs. No study has been conducted in Cambodia regarding the E. coli
resistance in pets and its associated environment.
OBJECTIVE
The present study aimed to determine the prevalence and resistance profiles among E. coli fromcohabitant pets and environmental surfaces at animal clinics in Phnom Penh City, Cambodia.
METHODOLOGY
Study Site and Sampling
The study was conducted in three animal clinics in Phnom Penh city, Cambodia from March to May
2019. In total, forty-seven samples were swabbed from dogs and environmental surfaces. The petsamples were selected based on the diagnosis of bloody diarrhea (3 to 8 months old), canine
parvovirus, and Pyometra disease. The environmental samples were selected based on the casereported animals above which animals were exposed to. Twenty-four samples (18 faeces and 6 pus)
were collected from dogs. Twenty-three samples (8 cage-floors, 7 cage-walls, 1 feeding plate, and 7
treatment-tables) were collected from the environmental surface. All samples were collected andtransported on the same day to the Veterinary Microbiology Laboratory of the Faculty of Veterinary
Medicine, Royal University of Agriculture, Cambodia.
E. coli Isolation and Identification
Samples were collected and the bacterial culture was followed by Markey et al. (2013). Briefly, theswap was subcultured into 5 mL of Buffered Peptone Water (BPW) and incubated at 370C for 24hours. Then the presumptive was streaked on MacConkey agar and incubated at 370C for 24 hours.
The suspected colonies with large/medium dark pink colonies were confirmed with biochemical tests
such as gram staining, Catala's test, Triple Sugar Iron (TSI), and Motility-Indole-Lysine (MIL) test.
Antimicrobial Susceptibility Testing
Antimicrobial susceptibility testing was performed using the disk diffusion method following
Clinical and Laboratory Standards Institute (CLSI, 2017) guidelines. Briefly, fresh E. coli isolates
were plated on Trypton-Soja-Agar (TSA) plates and incubated at 370C for 24 hours. Then 2-3
colonies on TSA were inoculated on NaCl suspension to a turbidity equivalent to 0.5 McFarland
standards. One hundred microliter suspension was plated on 15cm depth Mueller-Hinton agar
(MHA) and standard discs were applied using a dispenser and incubated for 16-18 hours at 370C. AⒸ ISERD

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total of six antimicrobial agents including Tetracycline (30 µg), Ampicillin (10 µg), Amoxicillin (10µg), Gentamycin (10 µg), Ceftriaxone (30 µg), and Levofloxacin (5 µg) were tested. All antibiotics
used for the study are based on the treatment protocol of each animal clinic.
RESULTS AND DISCUSSION
The present study found that a high prevalence of E. coli (54.16%) presented in dogs; whereas, a low
prevalence (21.74%) presented in environmental surfaces isolated from the animal clinics in Phnom
Penh city. E. coli presented in about 67% and 17% of dog faeces and pus, respectively. The result issimilar to the previous findings of Das et al. (2012) on the faeces of dogs. E. coli is the major
pathogen in the genital tract (Bassessar et al., 2013). Some studies have shown 50-85% prevalenceof E. coli in bitches suffering from pyometra disease (Bjurström et al., 1993; Sandholm et al., 1975;
and Trevena et al., 1996). Sharif et al. (2017) found that the prevalence of E. coli in the pus of bitcheswith pyometra was higher than the present study as the sample is much larger. Dog age is a factor
contributing to the prevalence of E. coli in faeces. Greene et al. (1998) have reported two strains ofE. coli were associated with gastrointestinal disease in young dogs. 77% of E. coli is present in young
dogs at age 1-3 months (Younis et al., 2015). However, Torkan et al. (2016) showed a significantly
lower prevalence of E. coli (47%) in the adult dog's faeces. The present study was on dogs aged 3-8months and showed a prevalence somewhere between the high and low values reported by the
aforementioned studies.
Table 1 The prevalence of E. coli distributed the animal clinics
Dogs (faeces and pus)
Number (n/N)
Percent (%)
01/50
20.00
05/10
50.00
07/90
77.78
13/24
54.16

Animal Clinics
A
B
C
Total

Environmental surface
Number (n/N)
Percent (%)
00/20
00.00
05/16
31.25
00/40
00.00
05/23
21.74

Table 2 The prevalence of E. coli distributed in animal and environmental surface samples
SampleDogs
Environmental
surfaces

FaecesPus
Cage-floor
Cage-wall
Feeding plate
Treatment table

Prevalence
Number (n/N)
12/18
01/60
3/8
1/7
0/1
1/7

Percent (%)
66.67
16.67
37.50
14.28
00.00
14.28

Total
Number (n/N)

Percent (%)

13/24

54.16

05/23

21.74

Table 3 The antimicrobial resistance profiles of E. coli in dogs and environmental surfaces
Antibiotics
Tetracycline
Ampicillin
Amoxicillin
Gentamycin
Ceftriaxone
Levofloxacin

Dogs
Number (n/N)
7/13
7/13
7/13
3/13
4/13
1/13

Percent (%)
53.85
53.85
53.85
23.08
30.77
07.69

Environmental surfaces
Number (n/N)
Percent (%)
5/5
100.004/5
080.005/5
100.003/5
060.004/5
080.003/5
060.00The prevalence of E. coli on the environmental surface varies based on the animals that havebeen exposed, according to Sidjabat et al. (2006). The highest prevalence of E. coli (37.50%) foundon environmental surfaces of the animal clinics in this study was isolated from the cage floor, whichis the most contaminated between faeces or pus of the dogs on the cage floor during hospitalization.
Instead, it might also be attributed to poor hygiene of the animal clinics and the cross-contamination
between dogs and the surfaces. Madubuike et al. (2016) reported a lower prevalence of E. coli
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bacteria on environmental surfaces (3.60%), especially the cage floor (0%) of the animal clinics inNigeria might be the biosecurity measures taken in the animal clinics were able to reduce thecontamination of E. coli.
E. coli showed high resistance to amoxicillin, ampicillin, and tetracycline (54%); while lessresistant to ceftriaxone (31%), gentamycin (23%), and levofloxacin (8%) in dogs. Similarly, these
isolates also had high resistance to amoxicillin, ampicillin, ceftriaxone, and tetracycline (80-100%);
while less resistance to gentamicin and levofloxacin (60%) on environmental surfaces. The highresistance to antibiotics on environmental surfaces was higher than the dog samples in the present
study. This might be fewer samples in the environment and the possibility of E. coli surviving on thesurfaces. Leite et al. (2013) and Carvalho et al. (2016) reported that pet dogs were shown to be apotential household source of multi-resistant E. coli strains to humans through the environment.
Resistance to antibiotics in dogs in the present study is consistent with the study of Nam et al. (2010).
CONCLUSION
In conclusion, both dogs and environmental surfaces showed identical prevalence of E. coli and its
resistance to several important antimicrobials. This is correlated with the spreading of antimicrobial
resistance into the environmental surfaces of the animal clinics from hospitalized dogs and could bespreading to owners and veterinarians, a cause of concern with regard to animal and public health.
ACKNOWLEDGEMENTS
The study is supported by the Center of Excellence on Sustainable Agricultural Intensification and
Nutrition (CE SAIN) of the Royal University of Agriculture (RUA) receives financial support fromthe U.S. Agency for International Development (USAID) through Feed the Future Innovation Lab
for Sustainable Intensification (SIIL).
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