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Genomic characterization and therapeutic potential of five broad-spectrum lytic bacteriophages against multidrug-resistant avian pathogenic Escherichia coli (APEC).

UNLABELLED: Colibacillosis, caused by avian pathogenic Escherichia coli (APEC), results in substantial economic losses in global poultry production. The emergence of multidrug-resistant (MDR) APEC poses zoonotic risks through horizontal transfer of antimicrobial resistance (AMR) genes. Bacteriophage therapy emerges as a safe alternative to antibiotherapy; however, comprehensive characterization of phages targeting MDR-APEC from diverse geographical regions remains limited. We isolated five lytic bacteriophages from poultry fecal samples collected from five Indian states and characterized them through morphological analysis, physiological stability testing, whole-genome sequencing, and in vivo efficacy assessment. Host range was determined against APEC isolates, and therapeutic potential was validated in the Galleria mellonella infection model. All phages showed Myovirus-like morphology and stability across physiologically relevant temperatures (up to 55°C-70°C) and pH conditions (3-11). Phages were classified as Escherichia phage vB_EcoM_fRPOT1, vB_EcoM_fDMYT1, vB_EcoM_fBSZT1, vB_EcoM_fUAMT1, and vB_EcoM_fPKPT2. Their genome size ranges from 170 to 356 kb, belonging to three distinct genera: Dhakavirus, Gaprivervirus, and Asteriusvirus. Genomic analysis confirmed the absence of antimicrobial resistance, virulence, toxin, or lysogeny genes. Fifty-one APEC strains were isolated, of which 23 (45.1%) were MDR. Individual phages lysed 37%-51% of tested APEC and 17%-39% of MDR strains. Three phages (fBSZT1, fUAMT1, and fPKPT2) significantly improved larval survival to 60%-80% at an MOI of 10 in G. mellonella infection models compared to the untreated control. This study establishes a well-characterized phage bank targeting MDR-APEC strains, providing a foundation for developing phage-based interventions to reduce antibiotic dependency and mitigate AMR transmission risks under the One Health framework. IMPORTANCE: The overuse of antibiotics in poultry farming has created a crisis. The multidrug-resistant (MDR) bacteria threaten both animal health and human safety through the food chain. When antibiotics fail, farmers face devastating losses, and resistant bacteria can transfer to humans through consumption or environmental contamination. Bacteriophages offer a practical solution as they kill target bacteria without harming beneficial microbes or leaving chemical residues. Our comprehensive characterization confirms that these five phages are safe and effective as they lack any resistance or toxin genes and rescue 60%-80% of infected larvae. This represents a characterized phage bank targeting the specific resistant strains in Indian poultry. By providing a validated alternative to antibiotics, this work supports sustainable food production while reducing the spread of antimicrobial resistance from farms to humans.

Animals

Therapeutic potential of a novel virulent bacteriophage XQ-1 against avian pathogenic Escherichia coli infection in broiler chickens.

Avian Pathogenic Escherichia coli (APEC) represents a significant subgroup within extraintestinal pathogenic Escherichia coli strains and constitutes a substantial threat to the global poultry industry. Although the negative impacts of APEC have been mitigated considerably through antibiotic use, this practice has concurrently facilitated the widespread emergence and dissemination of antibiotic-resistant APEC strains worldwide. Consequently, bacteriophage has emerged as a promising alternative to antibiotics. In the current study, a novel virulent bacteriophage, designated XQ-1, was isolated from a sewage sample collected at a broiler chickens farm in Hubei Province, China. Notably, this bacteriophage exhibited the capability to lyse multiple APEC strains, including O1, O2, and O78 serotypes. The optimal multiplicity of infection (MOI) for bacteriophage XQ-1 was determined to be 0.001, yielding a maximum viral titer of 4.73 ± 0.31 × 1011 plaque-forming units (PFU) per milliliter. This bacteriophage displayed a latent period of 30 min, and a burst period of 80 minutes, corresponding to a burst size of 348 PFU per infected cell. Additionally, bacteriophage XQ-1 retained high lytic activity across a temperature range of 4-50 °C and maintained tolerance with a pH range of 3-11. Further in vitro studies demonstrated that bacteriophage XQ-1 holds potential as an effective disinfectant capable of directly lysing APEC strain JZ-1. In chick models subjected to intraperitoneal injection, exposure to APEC resulted in 100% mortality in chicks. However, treatment with bacteriophage XQ-1 significantly improved the survival rates of infected broiler chickens, reduced organ indexes, decreased bacterial loads in the liver and heart organs, and mitigated intestinal damage caused by APEC infection. Collectively, these findings suggest that bacteriophage XQ-1 represents a promising candidate for the prevention and treatment of APEC infections in poultry.

APEC

Phylogrouping and genotyping of mcr-1 postives avian pathogenic Escherichia coli isolates in Algerian poultry farms.

Colibacillosis is a highly prevalent bacterial disease in poultry, resulting in the widespread use of antibiotics for both curative and preventive purposes. Consequently, avian pathogenic Escherichia coli (APEC) continues to act as a reservoir for antibiotic resistance genes, including the mcr-1 gene, which codes for resistance to colistin, a crucial antibiotic in human medicine. The aim of this study was to evaluate the antibiotic resistance pattern of APEC and to investigate the genotyping, phylogrouping, and virulence of mcr-1-positive isolates. A total of 113 APEC were isolated, of which 92% were multidrug resistant (MDR). The mcr-1 gene was detected in 41 isolates originating from turkeys and broilers. Two isolates carried blaTEM, one of which also harboured blaCTX-M encoding beta-lactamases. The Clermont phylogrouping revealed that 76% of the isolates belonged to phylogroup B1. Concerning the detection of the virulence-associated genes, 88% of isolates carried at least 3 genes. The ERIC-PCR classified our isolates into 6 different clusters. Our study highlights the emergence of colistin resistance and MDR, which pose a real threat to poultry production and public health. Control of antibiotic use in the poultry sector is urgent and mandatory.

Animals

Performance of the IR Biotyper, Nanopore, and Illumina sequencing to discriminate Escherichia coli strains originating from poultry.

UNLABELLED: Escherichia coli is a highly diverse bacterial species that includes avian pathogenic E. coli (APEC), one of the most prevalent causative agents of disease in poultry worldwide. Rapid and accurate discrimination of E. coli strains is essential for outbreak management, antimicrobial resistance surveillance, and vaccine development. In this study, we compared the performance of Fourier Transform Infrared (FTIR) spectroscopy using the IR Biotyper system with Nanopore and Illumina whole-genome sequencing (WGS) for typing 200 E. coli isolates, originating from four poultry rearing farms in the Netherlands. From each farm, we sampled 10 one-day-old meat type rearing chicks, and from every chick, we isolated 5 E. coli strains. FTIR clustering showed strong concordance with WGS-based classifications, particularly serotyping and core-genome similarity determined by PopPUNK analysis (Adjusted Rand Index 0.75-0.92). While Nanopore and Illumina sequencing provided the highest genetic resolution, FTIR offered a faster (max 6 vs 12-28 days for 200 isolates) and more cost-effective alternative for assessing clonality. Across all methods, multiple strains were detected per farm, whereas most birds carried a single dominant E. coli strain. Our findings demonstrate that FTIR provides a reliable and scalable phenotypic method for rapid strain discrimination in E. coli, complementing WGS in diagnostic, surveillance, and epidemiological settings where speed and throughput are critical. IMPORTANCE: Escherichia coli is a major pathogen in poultry and a potential zoonotic risk for humans. Rapid and accurate discrimination of avian pathogenic E. coli (APEC) strains is critical for outbreak management, antimicrobial resistance surveillance, and the design of effective autogenous vaccines. In this study, we compared Fourier Transform Infrared (FTIR) spectroscopy with Nanopore and Illumina whole-genome sequencing for strain typing of E. coli isolates originating from poultry. The results show that FTIR provides comparable clustering accuracy to genomic approaches at a fraction of the time and costs. This work demonstrates that FTIR can serve as a practical, high-throughput alternative for routine monitoring of E. coli in veterinary diagnostics and food safety of poultry meat, enabling faster decision-making and more targeted interventions across the poultry production chain.

Animals