Characterization and genomic analysis of bacteriophages infecting antibiotic-resistant Staphylococcus aureus isolated from hospital surfaces

Project Abstract

This study presents a comprehensive characterization and genomic analysis of bacteriophages that infect antibiotic-resistant Staphylococcus aureus strains isolated from hospital surfaces, aiming to explore phage-based strategies for controlling nosocomial infections. We collected 120 S. aureus isolates from high-touch surfaces in multiple hospital wards over six months, screening for resistance profiles against methicillin, vancomycin, linezolid, and daptomycin. MRSA-like isolates demonstrated multidrug resistance patterns, while phenotypic assays indicated varying biofilm-forming capacities that potentially influence phage susceptibility. Environmental samples were processed to isolate lytic phages, followed by enrichment with host strains and purification to obtain clonal phage populations. Morphological assessment via transmission electron microscopy classified isolates into distinct tailed phage families, with most displaying Siphoviridae and Myoviridae morphologies, hinting at diverse receptor-binding strategies. Whole-genome sequencing of 12 representative phages revealed genome sizes ranging from 38 to 170 kbp, with GC contents consistent with Staphylococcus phages. Bioinformatic analyses identified open reading frames encoding structural proteins, lytic enzymes, tail fibers, and regulatory modules, alongside auxiliary metabolic genes that may modulate host interactions. Importantly, genomes were screened for undesirable traits, including toxin genes, antibiotic resistance determinants, and lysogeny-associated integrases; all selected phages exhibited strictly lytic life cycles with no integrase genes detected, supporting their therapeutic potential. Host range assays demonstrated variable lytic activity across the panel of antibiotic-resistant S. aureus strains, with several phages capable of lysing 60–85% of the tested isolates, including a subset of MDR phenotypes. One-step growth curves characterized adsorption rates, latent periods, and burst sizes, revealing rapid adsorption and robust replication kinetics under physiologic conditions. In vitro bactericidal efficacy was assessed in planktonic cultures and biofilm models, showing substantial reductions in viable counts (up to 4–5 log10 units) within 6–24 hours, and significant disruption of biofilm architecture as observed by confocal microscopy. Phage-encoded depolymerases displayed activity against S. aureus extracellular polysaccharides, correlating with enhanced biofilm penetration. Transcriptomic analyses of phage-host interactions highlighted upregulation of bacterial stress responses and downregulation of virulence-associated pathways during infection. Proteomic profiling of phage particles identified conserved structural proteins and accessory enzymes that may contribute to host cell wall degradation. Additionally, synergy tests combining selected phages with sub-inhibitory antibiotic concentrations indicated potential for phage-antibiotic co-therapy to restore susceptibility in resistant strains. Collectively, the study delineates a diverse pool of lytic bacteriophages with favorable safety profiles and potent anti-staphylococcal activity, establishes genomic determinants of host range and virulence, and provides a foundational framework for deploying phage-based interventions to mitigate hospital-acquired S. aureus infections while addressing the challenge of antibiotic resistance.

Project Overview

What This Project Is About

---

A plain-language overview of the topic and what the project investigates.

---

---

The Problem It Addresses

---

What problem or gap this project tackles and why it matters to the field or society.

---

---

Objectives of the Project

---

1. Identify and isolate bacteriophages that infect antibiotic-resistant Staphylococcus aureus from hospital surfaces.
2. Characterize the phages’ basic traits, such as shape, genome size, and host range.
3. Analyze phage genomes to identify genes related to infection and replication.
4. Assess potential safety concerns and check for undesirable genes that could transfer resistance.
5. Explore how phages could contribute to alternative treatment strategies or infection control.

---

---

What You Will Do Step by Step

---

1. Collect samples from hospital surfaces and culture Staphylococcus aureus strains that show antibiotic resistance.
2. Isolate bacteriophages that can infect these bacteria from the samples.
3. Examine phage morphology using basic imaging methods and determine their host range.
4. Sequence phage genomes and perform basic bioinformatic analyses to annotate genes.
5. Evaluate safety by screening for toxin or antibiotic resistance genes.
6. Compare newly discovered phages to known ones to understand diversity.
7. Discuss how findings could inform future research or practical applications.

---

---

Expected Outcome

Expected outcomes include a set of characterized phages with genome data, a short report on their safety and potential uses, and recommendations for future work in phage therapy or hospital infection control.