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Manley, R.

Publications and source records attributed to Manley, R..

3 recordsLinked to original sources
bioRxiv2026

Lipopolysaccharide truncation and restoration drives a trade-off in resistance to two phages in Pseudomonas aeruginosa

Phage therapy is a promising treatment for multidrug resistant bacterial infections, and for patients no longer able to tolerate antibiotic treatments. A major challenge for phage therapy is emergent phage resistance, which target bacteria acquire by structurally modifying or masking phage receptors to prevent adsorption. Functionally diverse phage cocktails that target a broad range of receptors are less prone to resistance as there is a higher fitness cost associated with modifying multiple receptors. Expanding phage libraries with well-characterised phages that target a broad range of receptors would aid in timely and strategic design of functionally diverse phage cocktails. Here, we aimed to isolate phages targeting novel receptors by enriching wastewater samples on a Pseudomonas aeruginosa PAO1 {Delta}pilA {Delta}galU unmarked deletion mutant lacking O-antigen, outer core lipopolysaccharide (LPS) and type IV pili (T4P) - the three most common Pseudomonas phage receptors. This led to the isolation of a novel phage, named Vale. Vale was predicted to bind the LPS inner core as it could only infect strains with truncated LPS, suggesting that the outer core LPS blocks Vale from accessing the inner core. We identified a trade-oM in resistance to Vale and another phage, Tor, that targets the LPS outer core, mediated by host-derived LPS modifications. The PAO1 host evolved resistance to Tor by 100-200kb genomic deletions, which resulted in LPS truncation and sensitivity to Vale. Complete LPS restoration in the {Delta}pilA {Delta}galU mutant conferred resistance to Vale and sensitivity to Tor in two out of three replicates. Combined treatment with Tor and Vale delayed the emergence of resistance in PAO1 for at least three times longer than individual phage treatments. This study provides an example of how using phage receptors to strategically design phage cocktails can minimise the likelihood of emergent phage resistance. Graphical abstractA summary of the LPS modifications, genomic mutations and phage susceptibilities of Tor and Vale resistant mutants. "Parent strain" refers to PAO1 {Delta}hsdR. {Delta}pilA {Delta}galU refers to PAO1 {Delta}hsdR {Delta}pilA {Delta}galU. The parent strain gains resistance to Tor via LPS truncation associated with 100-200 kb genomic deletions, resulting in sensitivity to Vale. {Delta}pilA {Delta}galU gains resistance to Vale by restoring its LPS, conferring sensitivity to Tor in two out of three repeats. As resistance to one phage sensitises bacteria to the other, combined treatment with both phages suppresses phage resistance for longer than individual treatments. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=193 SRC="FIGDIR/small/700494v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1efee0forg.highwire.dtl.DTLVardef@f64bfeorg.highwire.dtl.DTLVardef@1f71130org.highwire.dtl.DTLVardef@1897bd6_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗
bioRxiv2025

Arbitrium phages can manipulate each other's lysis - lysogeny decisions

Many viruses can switch between lytic replication and dormancy (or lysogeny). It was recently discovered that some viruses that infect bacteria (known as bacteriophage, or phage) employ peptide-based ("arbitrium") communication systems to optimise their lysis/lysogeny switch: high peptide concentrations signal a lack of susceptible hosts and trigger lysogeny, while low peptide concentrations signal an abundance of uninfected hosts and prompt lysis. Here we demonstrate that Arbitrium-phages belonging to different species and genera can influence each others infection dynamics by secreting similar communication peptides, leading to early lysogenisation of the signal-receiving phage, and elevated fitness of the signal-emitting phage. Antagonistic coevolution between signal emitting and signal receiving phages to manipulate each others infection behaviours may explain the rapid diversification of arbitrium systems and their frequent horizontal exchange to escape the noise of cross talk.

ecology↗
bioRxiv2025

Harnessing droplet microfluidics and morphology-based deep learning for the label-free study of polymicrobial-phage interactions

Evaluating the impact of bacteriophages on bacterial communities is required to assess the future utility of phage therapy. Methods able to study bacterial polycultures in the presence of phages are useful to mimic evolutionary pressures found in natural environments and recapitulate complex ecological contexts. Bacteriophages can drive rapid genetic and phenotypic changes in host cells. However, the presence of other bacteria can also impact bacterial densities and community structure, and classical methods remain lengthy and resource intensive. Here, we introduce a microdroplet-based encapsulation method in which bacterial co-cultures are imaged using Z-stack brightfield microscopy. The method relies on automated droplet imaging using a novel AI-based autofocus function, coupled with morphology-based deep learning models for accurate identification of two morphologically distinct bacterial species. We show that we can monitor the relative growth dynamics of P. aeruginosa and S. aureus growing in 11 picolitre droplets for up to 24 hours. We demonstrate quantification of growth rates, bacterial densities and lysis dynamics of the two species without the need for plating. We show that a potent lytic phage of P. aeruginosa can either fully lyse the initial P. aeruginosa population or keep its density low long-term when in the presence of S. aureus.

microbiology↗