Effects of in ovo probiotic administration on the incidence of avian pathogenic Escherichia coli in broilers and an evaluation on its virulence and antimicrobial resistance properties

Abstract

Avian pathogenic Escherichia coli (APEC) causes colibacillosis in poultry, which has been traditionally controlled by the prophylactic in-feed supplementation of antibiotics. However, antibiotics are being removed from poultry diets owing to the emergence of multidrug-resistant (MDR) bacteria. Therefore, alternatives to control APEC are required. This study aimed to evaluate the effects of in ovo inoculation of probiotics on the incidence of APEC in broilers and evaluate the virulence and antimicrobial resistance properties of the APEC isolates. On embryonic day 18, 4 in ovo treatments (T) were applied: T1 (Marek's vaccine [MV]), T2 (MV and Lactobacillus animalis), T3 (MV and Lactobacillus reuteri), and T4 (MV and Lactobacillus rhamnosus). A total of 180 male broilers per treatment were randomly placed in 10 pens. The heart, liver, spleen, and yolk sac were collected on day 0, 14, 28, and 42. Presumptive E. coli isolates were confirmed by real-time PCR. The positive isolates were screened for the APEC-related genes (iroN, ompT, hlyF, iss, and iutA), and E. coli isolates containing one or more of these genes were identified as APEC-like strains. A total of 144 APEC-like isolates were isolated from 548 organ samples. No differences (P > 0.05) among treatments were observed for the incidence of APEC-like strains in all organs when averaged over sampling days. However, when averaged over treatments, the incidence in the heart, liver, and yolk sac was different among sampling days; a significant increase was observed in these organs on day 14 compared with day 0. Twenty-five antimicrobial resistance genes were evaluated for all APEC-like isolates, and 92.4% of the isolates carried at least one antimicrobial resistance gene. Thirty-seven isolates were then selected for antimicrobial susceptibility testing; MDR strains accounted for 37.8% of the isolates. In conclusion, the in ovo inoculation of a single probiotic strain did not confer protection against APEC strains in broilers. The high prevalence of MDR isolates indicates that further research on antibiotic alternatives is required to prevent APEC infections in broilers.

Keywords: Lactobacillus; in ovo inoculation; multidrug-resistant; real-time PCR detection; virulence gene.