Enterotoxigenic Escherichia coli (ETEC) is one of the most common causative agents of foodborne gastroenteritis globally, spreading through contaminated water and food. The rising incidence of multidrug-resistant (MDR) bacteria has produced an urgent need for alternative antimicrobial treatments. Bacteriophages are potential therapeutic agents against MDR infections; yet, the development of phage resistance remains a critical limitation to their clinical applications. In this study, we isolated and characterized an ETEC-targeting lytic bacteriophage (CU10EC) with potent antibacterial activity. Phage resistance was observed within ~10 h of treatment, making phage therapy ineffective. To address this limitation, we used a novel combinatorial approach that combined cell free probiotic biogenics (PB) produced by the probiotic bacterium Lactobacillus acidophilus with CU10EC-mediated therapy. The PB contains a rich array of biomedically relevant components, including organic acids, antimicrobial peptides, short-chain fatty acids, and proteolytic enzymes, as confirmed by LC-MS analysis. It demonstrated effective antibacterial activity against MDR-ETEC, broad-spectrum activity against other pathogens, and less or no antibacterial activity against commensals. Importantly, in the presence of phage CU10EC, only two-thirds of the PB concentration was required for complete bacterial killing. Additionally, the consecutive administration of PB at 2, 4, and 6 h in a time-kill assay delayed phage resistance by 6–10 h. Overall, the CU10EC-PB combination offers a novel method to enhance antibacterial activity, while conserving gut commensals and delaying phage resistance.
Citation: Abishek Mani, Nashath Kalangadan, Sharayu Magar, Barath Sivaraj, Gospel Gabriel Xavier, Thirumalai Diraviyam, Srivignesh Sundaresan, Sutharsan Govindarajan, Kaushik Rajaram. Probiotic biogenics-mediated augmentation of phage therapy against multidrug-resistant Enterotoxigenic Escherichia coli[J]. AIMS Microbiology, 2026, 12(4): 789-814. doi: 10.3934/microbiol.2026034
Enterotoxigenic Escherichia coli (ETEC) is one of the most common causative agents of foodborne gastroenteritis globally, spreading through contaminated water and food. The rising incidence of multidrug-resistant (MDR) bacteria has produced an urgent need for alternative antimicrobial treatments. Bacteriophages are potential therapeutic agents against MDR infections; yet, the development of phage resistance remains a critical limitation to their clinical applications. In this study, we isolated and characterized an ETEC-targeting lytic bacteriophage (CU10EC) with potent antibacterial activity. Phage resistance was observed within ~10 h of treatment, making phage therapy ineffective. To address this limitation, we used a novel combinatorial approach that combined cell free probiotic biogenics (PB) produced by the probiotic bacterium Lactobacillus acidophilus with CU10EC-mediated therapy. The PB contains a rich array of biomedically relevant components, including organic acids, antimicrobial peptides, short-chain fatty acids, and proteolytic enzymes, as confirmed by LC-MS analysis. It demonstrated effective antibacterial activity against MDR-ETEC, broad-spectrum activity against other pathogens, and less or no antibacterial activity against commensals. Importantly, in the presence of phage CU10EC, only two-thirds of the PB concentration was required for complete bacterial killing. Additionally, the consecutive administration of PB at 2, 4, and 6 h in a time-kill assay delayed phage resistance by 6–10 h. Overall, the CU10EC-PB combination offers a novel method to enhance antibacterial activity, while conserving gut commensals and delaying phage resistance.
AM, NK, GGX, and KR acknowledge support from the Central University of Tamil Nadu for essential laboratory facilities, ICMR small grant (DDP/10/IIRP-SG2024/00365) and HEFA-CSR grant (SANCSR/05/2025-2026). SG acknowledges support from SRM University AP (SRMAP/URG/GENERAL-D/2025-26/007).
None declared.
Abishek Mani: Methodology, data curation, study design, and writing-original draft. Nashath Kalangadan: Methodology, investigation. sharayu magar: methodology, data interpretation, validation. Barath Sivaraj: Methodology, investigation. Gospel Gabriel Xavier: Methodology, investigation. Thirumalai Diraviyam: Resources, visualization. Srivignesh Sundaresan: Formal analysis and visualization. Sutharsan Govindarajan: Resources, supervision, validation, writing-review and editing. Kaushik Rajaram: Conceptualization, supervision, validation, data interpretation, resources, writing-review and editing.
The complete genome sequence of vB_EcoP_CU10EC has been deposited in GenBank under accession number PQ435408.
The supporting data of this study are available from the Zenodo (Data repository) with the following links,
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Figures(8) / Tables(1)
Abishek Mani, Nashath Kalangadan, Sharayu Magar, Barath Sivaraj, Gospel Gabriel Xavier, Thirumalai Diraviyam, Srivignesh Sundaresan, Sutharsan Govindarajan, Kaushik Rajaram. Probiotic biogenics-mediated augmentation of phage therapy against multidrug-resistant Enterotoxigenic Escherichia coli[J]. AIMS Microbiology, 2026, 12(4): 789-814. doi: 10.3934/microbiol.2026034
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