Evaluation of the antimicrobian resistance of Escherichia coli isolated of healthy hens (Gallus gallus)/Avaliacao da resistencia antimicrobiana de Escherichia coli isoladas de galinhas (Gallus gallus) saudaveis/Evaluacion de la resistencia antimicrobiana de Escherichia coli aislada de gallinas (Gallus de gallus) sanas.
Escherichia coli (E. coli) is a Gram negative bacillus, facultative anaerobic, belonging to the family Enterobacteriaceae, found colonizing the healthy gut of birds and mammals (1). The pathogenic lineages for birds are named Avian Pathogenic Escherichia coli (2), responsible for infections characterized by signs of septicemia, peritonitis, pneumonia, aerossaculitis, pericarditis, onfalitis and salpingitis. The infection caused by E. coli represent one of the main problem of the industrial poultry breeding, responsible for economic losses as mortality, decrease of weight and condemnation of carcasses in the slaughterhouse (3). The use of antibiotics in the animal food as "promoters of growth" or in an indiscriminate way under the form of sub or over dosage (3), increases the resistance genes selection pressure to the antibiotics for multiresistant bacterial lineages (4).
Studies have shown the resistance genes transmission between the micro-organisms. This fact has become a concerning a public health problem, as the genes of resistance may be transfered to pathogenic bacteria that can infect the humans (5). Many species of the family Enterobacteriaceae present in the gut of healthy birds are frequently exposed to several antibiotics during the life of the animal (6). The present study evaluated the profile of resistance to 12 antibiotics in isolated of E. coli obtained from the excrements of adult healthy hens, without intestinal signs.
MATERIAL AND METHODS
One hundred samples of E. coli obtained from the healthy posture hens excrements were used. The samples were isolated utilizing swab of cloaca, and cultivated in agar MacConkey, in conditions of aerobiosis, to 37[degrees] C, for 24 hours. The characteristic colonies were submitted to the biochemical analysis. For the achievement of the sensitiveness test to the antibiotics, the colonies identified like E. coli were submitted to the diffusion standard test with disks (7), utilizing 12 antibiotics: cefalexin (30[micro]g), gentamicin (10[micro]g), enrofloxacin (5[micro]g), norfloxacin (10[micro]g), doxiciclin (30[micro]g), florfenicol (30[micro]g), cloranfenicol (30[micro]g), neomycin (30[micro]g), amoxicillin (10[micro]g), tetraciclin (30[micro]g), ampicillin (10[micro]g) and penicillin (10[micro]g).
The results were submitted to the statistical analysis, using the program SAS (table 1).
RESULTS AND DISCUSSION
Table 1 shows the antibiogram results. From 100 samples of E. coli, just two (2%) showed itself sensitive to all the tested antibiotics, 15 presented resistance to one or two antibiotics and the other samples were resistant to three or more antibiotics; two of them were resistant simultaneously to the 12 tested antibiotics. Zao et al (8) described similar results when evaluating 95 isolated samples of E. coli, in which 71% were resistant to five or more antibiotics.
Neomicin and ampicillin were not effective for the isolated samples. Blanco et al. (4) observed resistance in 46% of the isolated sample for ampicillin and 79% for neomicin when evaluating 310 isolated samples, differing from the results found in this study. The antibiotic which showed the best effectiveness facing the 100 isolated samples was cloranfenicol with 85% of sensitiveness. Zao et al. (8) observed 89% of sensitiveness of E. coli to this antibiotic. The prescription of antibiotic for healthy birds, in subtherapeutic concentrations for prolonged time, exposes microbiota to a selective pressure, enabling the acquisition of resistance genes to the antibiotics. The intestinal lumen complex, where a great amount of bacteria interacts in very limited space, represents an ideal niche for studies in vivo of the resistance genes transfer between different species and gender (9).
The occurrence of multiresistant samples of E. coli from animal origin is concerning to the world health organization (10). Another prominent point resides in the fact of the majority of the antibiotics available in the market are based on ampicillin or neomicin that, coincidentally are broadly used in the human and veterinary medicine. However, these drugs of the group showed have low or no efficiency against all the samples tested, fact that can be consequence of the inappropriate use of the antibiotics in the poultry breeding.
Escherichia coli samples isolated from excrements of healthy birds were resistant to the antibiotics, fact that is a great problem for the poultry breeding and public health with dissemination risk of multiresistant gene to the antimicrobians to other microorganisms including the pathogenic ones.
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(3.) Ferreira AJP, Knobl T. Colibacilose aviaria. In: Berchieri Junior A, Macari M. Doengas das aves. Campinas: Facta; 2000. p.197-207.
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(8.) Zhao S, Maurer JJ, Hubert S, De Villena JF, McDermott PF, Meng J, et al. Antimicrobial susceptibility and molecular characterization of avian pathogenic Escherichia coli isolates Vet Microbiol. 2005;107(3-4):215-24.
(9.) Schjprring S, Struve CE, Krogfelt KA. Transfer of antimicrobial resistance plasmids from Klebsiella pneumoniae to Escherichia coli in the mouse intestine. J Antimicrob Chemother. 2008;62(5):1086-93.
(10.) Martins IS, Nogueira IA, Conceicao M, Brasil P. Recomendacoes para o uso adequado dos antimicrobianos. Rio de Janeiro: Secretaria de Estado de Saude; 1998 [cited 2009 Set 21]. Available from: <http://www.saude.rj.gov.br/cecih/Antimicrobianos.doc>.
Recebido em: 13/01/11
Aceito em: 27/03/12
Adriano Sakai Okamoto 
Raphael Lucio Andreatti Filho 
Ana Angelita Sampaio Baptista 
Ticiana Silva Rocha 
[1,2] Professor Doctor discipline Avian Pathology FMVZ-UNESP-Botucatu-SP Brazil E-mail: firstname.lastname@example.org
Address: Department of Veterinary Clinic FMVZ UNESP District of Rubiao Junior S/N Botucatu--SP
 Doctoral Student Avian Pathology FMVZ-UNESP-BOTUCATU-SP
 Msc. Student Avian Pathology FMVZ--UNESP-BOTUCATU-SP
Table 1. Sensitivity and resistance by 100 bacterial strains of Escherichia coli from the normal intestinal microbiota of hens. Antimicrobial % sensibility/ Risk of bacterial resistance % resistance RAP CI 95% Value P Penicillin 0/100 Reference Reference NA Amoxicillin 17/83 0,049 0,01-0,38 <0,004 Ampicillin 0/100 1,0 0,06-16,21 >0,1 Cefalexin 25/75 0,03 0,004-0,23 <0,001 Cloranfenicol 85/15 0,002 0,001-0,01 <0,0001 Doxiciclin 50/50 0,01 0,001-0,07 <0,0001 Enrofloxacin 75/25 0,003 0,001-0,02 <0,0001 Florfenicol 67/33 0,005 0,001-0,04 <0,0001 Gentamicin 75/25 0,003 0,001-0,02 <0,0001 Neomicin 0/100 1,0 0,06-16,21 >0,1 Norfloxacin 50/50 0,01 0,001-0,07 <0,0001 Tetraciclin 50/50 0,01 0,001-0,07 <0,0001 RAP: Ratio of adjusted proportions; CI 95%: Confidence interval 95% probability; NA: Not Applicable.