Isolation of lytic bacteriophages against pathogenic Escherichia coli strains in poultry in the northwest of Iran

Document Type : Original Articles

Author

Department of Biology, Faculty of Basic Sciences, University of Maragheh, Maragheh, Iran

Abstract

In this study, 90 internal organ samples of poultry with symptoms of colibacillosis were obtained from Maragheh poultry farms in East Azerbaijan, Iran. In total, 70 bacterial isolates were confirmed as Escherichia coli (E. coli) strains using standard biochemical tests, and antibiotic sensitivity was determined by the disk diffusion method. Antibiotics used in this study included ampicillin, penicillin, nitrofurantoin, tetracycline, amoxicillin, ciprofloxacin, nalidixic acid, and sulfamethoxazole (n=8). Ciprofloxacin showed the highest susceptibility, while the lowest susceptibility was observed with penicillin and amoxicillin. Among the bacterial isolates, 50% showed resistance to at least five antibiotics, and 10 isolates with multidrug resistance were selected for bacteriophage (phage) isolation against recent E. coli isolates using spot test and double-layer agar overlay technique. In addition, water samples for phage isolation were provided from rivers, poultry farm sewages, and an urban sewage treatment center. In total, eight phages were successfully isolated from the urban sewage treatment center (total: 10). After enrichment, purification and titration, phages were further concentrated by polyethylene glycol precipitation. Lowest and highest bacteriophage titers were determined to be 1.05×106 and 1.9×109 PFU/ml, respectively. Host range of the isolated phages was assayed by spot testing, and antibacterial effects against four E. coli isolates were observed in one of the isolated phage suspensions, which was introduced as the most potentiated agent for phage therapy. In the morphological analysis of the selected phage using an electron microscope, we observed a hexagonal head with a diameter of 95 nm and contractile tail length of 90 nm, which indicated its similarity to the Myoviridae family. In conclusion, results of this study showed that bacteriophages could be appropriate alternatives to combat pathogenic E. coli strains with antibiotic resistance in poultry. Considering the changeable antibacterial effects of bacteriophages against different isolates of extraintestinal avian pathogenic E. coli, it is suggested that future investigations be conducted regarding the efficacy of lytic phages against different bacterial strains for the effective control of the associated infections in this region of Iran.

Keywords

Article Title [French]

Isolation des bactériophages lytiquesagissant contre les souches pathogènes d’Escherichia coli chez les volailles du nord-ouest de l’Iran

Abstract [French]

Dans cette étude, 90 prélèvements d’organes internes de volailles montrant des symptômes de colibacillose ont été obtenus des exploitations avicoles de la ville de Maragheh, située dans l’Est Azerbaïdjan (Iran). Au total, 70 isolats bactériens ont été identifiés comme étant des souches d’Escherichia coli (E. coli) par des tests biochimiques standard. La sensibilité aux antibiotiques des isolats a été ensuite déterminée par la méthode de diffusion sur disque. Les antibiotiques utilisés étaient l’ampicilline, la pénicilline, la nitrofurantoine, la tétracycline, l’amoxicilline, la ciprofloxacine, l’acide nalidixique et le sulfaméthoxazole (n=8). Les isolats étudiés montraient une susceptibilité plus importante vis-à-vis de la ciprofloxacine alors que le penicilline et le amoxicilline représentaient lesdeux antibiotiques les moins performants. Parmi les isolats bactériens, 50% étaient résistants à au moins 5 antibiotiques. Dix isolats multi-résistants ont été sélectionnés pour l’isolation des bactériophages par le biais de test ponctuel (spot test) et par la méthode de double couche de gélose de recouvrement (double-layer agar overlay). Les échantillons d’eau utilisés comme sources de bactériophages provenaient des rivières, des eaux usées d’exploitations avicoles et d’une station d’épuration urbaine. Au total, 8 phages ont été isolés à partir des échantillons récoltés dans la station d’épuration urbaine. Après enrichissement, purification et titration, les phages ont été concentrés davantage par précipitation au polyéthylène glycol. Les titres les plus bas et les plus élevés de bactériophages étaient respectivement de 1.05×106 et 1.9×109 PFU/ml. La spécificité d’hôtes a été déterminée par test ponctuel et a révélé une activité antibactérienne contre 4 isolats d’E. colidans l’une des suspensions de phages analysées, démontrant, de ce fait, l’intérêt thérapeutique de cette échantillon. Nos observations morphologiques par microscopie électronique montraient que le phage d’intérêt exhibait une tête hexagonale de 95 nm de diamètre et une queue contractile d’un diamètre de 90 nm, similaire à la famille des Myoviridae. En conclusion, ces résultats montrent que les bactériophages constituent une alternative thérapeutique appropriée pour combattre les souches pathogènes multi- résistantes d’E. colichez les volailles. Etant donnée la variabilité de l’effet antibactérien des bactériophages selon les différents isolats extra-intestinaux aviaires des souches pathogènes d’E. colianalysés, des études supplémentaires sur l’efficacité des phages lytiques contre différentes souches bactériennes pathogènes provenant d’autres régions d’Iran sont nécessaires et pourraient contribuer à un control plus efficace des infections associées.

Keywords [French]

  • Bactériophage
  • Colibacillose
  • Résistance antibiotique
  • Escherichia coli
  • Volaille

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Archives of Razi Institute
Volume 71, Issue 4
November and December 2016
Pages 235-244

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