Antimicrobial Resistance among Escherichia coli Strains Isolated from Healthy and Septicemic Chickens
There is a clear association between heavy antimicrobial consumption in poultry industry and the recovery of resistant bacteria. This was a case-control study of 396 E. coli strains isolated from clinically affected broiler chickens and 132 strains from healthy controls to compare the antimicrobial resistance rates. Antimicrobial resistance testing of 525 avian E. coli strains isolated in Kashan-Iran showed very high levels of resistance to 11 antimicrobials tested, especially to trimethoprim-sulfamethoxazole (98.7%) and to ciprofloxacin (69.7%). The prevalence rate of resistant E. coli to ciprofloxacin and erythromycin in the samples isolated from chickens with colibacillosis was significantly higher than healthy controls. In addition, to prevent the emergence of cross-resistance with human enteric pathogens, controlled use of these antimicrobial agents in veterinary practice is recommended.
Colibacillosis in chickens is frequently manifested as a septicemia resulting in a subacute serositis characterized by fibrinoheterophilic infiltration producing a fibrinous pericarditis, perihepatitis, airsacculitis and pneumonia. It is one of the most serious threats to broiler chicken flock between 4 and 5 weeks of age with respiratory signs, omphalitis and septicemia in baby chicks causing high mortality rates. It is trasmitted through contamination of egg shellwith pathogenic E. coli from poultry house orhatcher environment (AAAPG, 2005).
Antibiotics have been used on poultry husbandry farms to treat and control diseases in chickens and to improve chicken productivity (Peter, 1999). This practice is reported to have caused high resistance to antimicrobial agents in normal intestinal flora and pathogenic organisms of chickens (Tollefson et al., 1997; Manie et al., 1998). E. coli is commonly found in the intestinal tracts of animals, but only pathogenic serotypes cause avian collibacillosis (Aarestrup and Wegener, 1999; Gorbach, 2001). The transfer of resistant E. coli from chickens to humans is a common event, as has been demonstrated by several groups of researchers (Kanai et al., 1983; Van den Bogaard and Stobberingh, 1999). Antimicrobial agents are widely used in poultry husbandry in Iran as therapy for an infection or, in the absence of disease, in sub-therapeutic doses with the goals of growth promotion and enhanced feed efficiency. Multidrug-resistant strains of E. coli are prevalent in both human and animal isolates in different parts of the world and Iran, too (Nazer, 1980; Blanco Jesus et al., 1997; Al-Ghamdi Mastour et al., 1999). To look into these issues with more details, the current study was designed to evaluate the frequency of resistant E. coli isolated from septicemic broiler chickens and healthy controls.
MATERIALS AND METHODS
This was a case-control study of 396 E. coli strains isolated from clinically
affected broiler chickens and 132 strains from healthy controls were collected
in 20 commercial poultry husbandry fields in province of Isfahan (Kashan, Iran)
in 2004. All strains isolated from diseased chickens came from confirmed cases
of collibacillosis in which bacteria were obtained in culture from liver and
heart tissues. Fecal strains were isolated from the cloacal contents of healthy
chickens. Antimicrobial susceptibilities were assessed for amikacin, ampicillin,
chloramphenicol, ciprofloxacin, gentamicin, tobramicin, doxycycline, nalidixic
acid, erythromycin, nitrofurantoin and trimethoprim/sulfamethoxazole by the
standard disk diffusion method in Muller-Hinton agar with disks provided by
Difco and Biomerieux according to standards developed by the National Committee
for Clinical Laboratory Standards (NCCLS, 1999), which its name recently changed
to CLSI (Clinical Laboratory Standards Institute), guidelines. E. coli
ATCC 25922 and E. coli ATCC 35218 were quality-control organisms. Three
samples of healthy controls were excluded from the study. The statistical procedures
were: Fischers exact test and Chi-square. We considered differences significant
at p<0.05. The ethical committee of the Kashan University of Medical Sciences
approved the study.
RESULTS AND DISCUSSION
The resistance frequencies for 11 antimicrobial agents tested are shown in
Table 1. As the Table 1 indicates, resistance
frequency rates were very high. The resistance rates for 525 E. coli
isolates were very high to amikacin and doxycycline (99.4%), nalidixic acid
(99%), followed by ampicillin, trimethoprim-sulfamethoxazole (SXT), erythromycin
and chloramphenicol. Ciprofloxacin showed high level of resistance rates (69.7%).
The antibiotic resistance rates to tobramycin and gentamicin were at moderate
level (11.6 to 12.6%). The resistant rates of E. coli to ciprofloxacin
(p<0.00002) and erythromycin (p<0.014) in the samples from chickens with
collibacillosis was significantly higher than healthy controls.
We have investigated the resistance of the large proportion of E. coli isolated with reduced susceptibility to antimicrobials could be the frequent exposure of chicken to abundant use and misuse of these drugs in commercial poultry production and colonized the resistant E. coli to 11 antimicrobial agents commonly used in the poultry industry and/or for human medicine in Kashan, Iran. The isolated E. coli from chickens clearly demonstrated high resistance rates to all tested antibiotics commonly used in the poultry industry.
In vitro antibiotic sensitivity results obtained in this study agreed
with several various reports (Blanco Jesus et al., 1997; Al-Ghamdi Mastour
et al., 1999), which have indicated increasing incidence rates of antibiotic
resistant E. coli strains isolated from healthy controls and diseased
chickens (Blanco Jesus et al., 1997). However, the high percentage of
E. coli strains that were resistant to amikacin, doxycyline, nalidixic
acid, trimethoprim/sulfamethoxazole and erythromycin in present study was surprising.
An interesting finding in this study was that the percentage of E. coli
resistant strains to erythromycin and ciprofloxacin (69.7%) in diseased group
were significantly higher than in control. Lee et al. (2005), reported
high resistance to ciprofloxacin (CIP; 60.2%), enrofloxacin (ENO; 73.4%) and
norfloxacin (NOR; 60.2%) in one hundred and twenty-eight isolated E. coli
The existence of high resistant E. coli in this study, like other results
of other researches is alarming and aware us to use these drugs according to
results of antibiogram tests and use the narrow spectrum instead of broad spectrum
antibiotics (Blanco Jesus et al., 1997; Al-Ghamdi Mastour et al.,
1999). Kijima-Tanaka et al. (2003), reported higher resistance rates
in Escherichia coli isolated from food-producing animals in Japan against
oxytetracycline and dihydrostreptomycin, followed by ampicillin and kanamycin.
Resistance was more frequently showed among broiler isolates, followed by isolates
from pigs. Almost 10% of broiler isolates were resistant to fluoroquinolones
and extremely high MICs (100 mg L‾1) were observed. As Asai
et al. (2005), mentioned the antimicrobial susceptibility of the isolates
was examined against 7 classes of 11 antimicrobials. The rates of antimicrobial
resistance among the isolates were found to correlate significantly with the
usage of antimicrobial agents in cattle, pigs and broiler and layer chickens.
Therefore, the overall usage of veterinary antimicrobials appears to contribute
to the appearance of antimicrobial resistance in E. coli isolates from
apparently healthy food-producing animals. In other study isolated E. coli
from diseased piglets (n = 89) and chickens (n = 71) in China were characterized
for antimicrobial susceptibility. The isolates displayed resistance
to nalidixic acid (100%), tetracycline (98%), sulfamethoxazole (84%),
ampicillin (79%), streptomycin (77%) and trimethoprim-sulfamethoxazoole (76%).
|| Antimicrobials resistance among Escherichia coli strains
isolated from healthy and septicemic chickens in Kashan-Iran in 2004
|Total incidences of resistance of strains from healthy (n
= 129) and septicemic (n = 396) chickens were 1092 and 3490, respectively
(total for both groups, 4582). Incidences of resistance per strain were
8.46 and 8.8 for healthy and septicemic groups, respectively (average for
both groups, 8.6)
Multiple-antimicrobial-resistant E. coli isolates, including fluoroquinolone-resistant
variants, are commonly present among diseased swine and chickens in China (Yang
et al., 2004).
Ninety-five avian pathogenic E. coli (APEC) isolates recovered from diagnosed cases of avian colibacillosis from North Georgia. Multiple antimicrobial-resistant phenotypes (> or = 3 antimicrobials) were observed in 92% of E. coli isolates, with the majority of isolates displaying resistance to sulfamethoxazole (93%), tetracycline (87%), streptomycin (86%), gentamicin (69%) and nalidixic acid (59%) (Zaho et al., 2005).
In present study the large proportion of E. coli isolated with reduced susceptibility to antimicrobials could be result of the frequent exposure of chicken to abundant use and misuse of these drugs in commercial poultry production and colonized the resistant E. coli. The high recovery rate of antimicrobial resistant E. coli from broilers in Iran was troubling, but not surprising. Given the routine application of the antimicrobials at subtherapeutic doses for prophylactic and therapeutic purposes by farmers, without prescription and for treatment by veterinary prescription in absence of documented laboratory findings.
In this study, current frequent usage of particular antibiotics in chickens seemed to correlate not only with high prevalence of resistance to those antibiotics, but also with a higher prevalence rates of drug resistance in the population of chickens. Frequent usage of an antibiotic in the past was also associated with relatively high prevalence of resistance in this population.
In conclusion, the increasing problem of bacterial resistance due to rational usage of antibiotics including the implementation of a veterinary antibiotic policy is of utmost importance to safegurad the efficacy of veterinary antibiotic therapy for the future and to minimize the public health risks from veterinary practice. Surveillance is an essential part of antibiotic administration policy and should consist of the regular or continuous monitoring of antimicrobial resistance.
This study was financially supported by a research grant from Deputy for Researches, Kashan University of Medical Sciences and Health Services, Kashan, Iran. We thank Dr. Mohsen Shafiee for his assistance with the preparation of the samples used in this study.
Aarestrup, F.M. and H.C. Wegener, 1999. The effects of antibiotic usage in food animals on the development of antimicrobial resistance of importance for humans in Campylobacter and Escherichia coli. Microbes Infect., 1: 639-644.
Al-Ghamdi, M.S., F. El-Morsey, Z.H. Al-Mustafa, M. Al-Ramadhan and M. Hanif, 1999. Antibiotic resistance of Escherichia coli isolated from poultry workers, patients and chicken in the eastern province of Saudi Arabia. Trop. Med. Int. Health, 4: 278-283.
American Association of Avian Pathologists Guidelines to Judicious Therapeutic Use of Antimicrobials in Poultry, 2005. (Approved by the AVMA Executive Board November 2000; revised by Executive Board April 2005).
Asai, T., A. Kojima, K. Harada, K. Ishihara, T. Takahashi and Y. Tamura, 2005. Correlation between the usage volume of veterinary therapeutic antimicrobials and resistance in Escherichia coli isolated from the feces of food-producing animals in Japan. Jpn. J. Infect. Dis., 58: 369-372.
Direct Link |
Blanco, J.E., M. Blanco, A. Mora and J. Blanco, 1997. Prevalence of bacterial resistance to quinolones and other antimicrobials among avian Escherichia coli strains isolated from septicemic and healthy chickens in Spain. J. Clin. Microbiol., 35: 2184-2185.
PubMed | Direct Link |
Collignon, P.J., 1999. Vancomycin-resistant Enterococci and use of avoparcin in animal feed: Is there a link? Med. J. Aust., 171: 144-146.
Gorbach, S.L., 2001. Anitmicrobial Use in Animal Feed? Time to Stop. Vol. 18. J. Med. Editorial, New England.
Kanai, H., H. Hashimoto and S. Mitsuhashi, 1983. Drug resistance and conjugative R plasmids in fecal Escherichia coli strains isolated from healthy younger animals (chickens, piglets, calves) and children. Microbiol. Immunol., 27: 1013-1041.
Kijima-Tanaka, M., K. Ishihara, A. Morioka, A. Kojima and T. Ohzono et al., 2003. A national surveillance of antimicrobial resistance in Escherichia coli isolated from food-producing animals in Japan. J. Antimicrob Chemother., 51: 447-451.
Direct Link |
Lee, Y., J.K. Cho, K. Kim, R. Tak, A. Kim, J. Kim, S. Im and B. Kim, 2005. Fluoroquinolone resistance and gyrA and parC mutations of Escherichia coli isolated from chicken. J. Microbiol., 43: 391-397.
Manie, T., S. Khan, V.S. Brozel, W.J. Veith and P.A. Gouws, 1998. Antimicrobial resistance of bacteria isolated from slaughtered and retail chickens in South Africa. Lett. Applied Microbiol., 26: 253-258.
NCCLS, 1999. Performance Standards for Antimicrobial Susceptibility Testing. National Committee for Clinical Laboratory Standards, Wayne, PA., USA.
Nazer, A.H., 1980. Transmissible drug resistance in Escherichia coli isolated from poultry and their carcasses in Iran. Cornell Vet., 70: 365-371.
Tollefson, L., S.F. Altekruse and M.E. Potter, 1997. Therapeutic antibiotics in animal feeds and antibiotic resistance. Revue Scientifique et Technique, 16: 709-715.
Van den Bogaard, A.E. and E.E. Stobberingh, 1999. Antibiotic usage in animals: Impact on bacterial resistance and public health. Drugs, 58: 589-607.
PubMed | Direct Link |
Yang, H., S. Chen, D.G. White, S. Zhao, P. McDermott, R. Walker and J. Meng, 2004. Characterization of multiple-antimicrobial-resistant Escherichia coli isolates from diseased chickens and swine in China. J. Clin. Microbiol., 42: 3483-3489.
CrossRef | Direct Link |
Zhao, S., J.J. Maurer, S. Hubert, J.F. De Villena and P.F. McDermott et al., 2005. Antimicrobial susceptibility and molecular characterization of avian pathogenic Escherichia coli isolates. Vet. Microbiol., 107: 215-224.
CrossRef | Direct Link |