Cell-free synthesis and characterization of Salmonella, Escherichia coli, and Shigella-specific bacteriophages.

Irem Iskender, Jaap Bosma, Paul Soudier, Yating Guo, Jinquan Li, Vincent Noireaux, Steven Bowden

Abstract read

In one paragraph

Article in Microbial cell factories, 2026. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.

0numbers the graph read from it

0cells of the map it votes in

0citing papers in PubMed

–field-weighted citation impact

1 · What the graph read from it

What it found

Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.

The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.

2 · The registry

The trial behind it

Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.

Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.

3 · Its place in the literature

Who cites it

0 citing papers in PubMed.

No citing paper in PubMed yet.

4 · The record

Corrections and comments

PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.

5 · Who and what money

Authors and funding

7 authors.

Irem IskenderDepartment of Food Science and Nutrition, University of Minnesota, Saint Paul, MN, 55108, USA.ORCID http://orcid.org/0000-0002-7800-2773

Jaap BosmaDepartment of Food Science and Nutrition, University of Minnesota, Saint Paul, MN, 55108, USA.

Paul SoudierSchool of Physics and Astronomy, University of Minnesota, Minneapolis, MN, 55455, USA.

Yating GuoNational Key Laboratory of Agricultural Microbiology, Key Laboratory of Environment Correlative Dietology, Hubei Hongshan Laboratory, College of Biomedicine and Health, College of Veterinary Medicine, College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China.

Jinquan LiNational Key Laboratory of Agricultural Microbiology, Key Laboratory of Environment Correlative Dietology, Hubei Hongshan Laboratory, College of Biomedicine and Health, College of Veterinary Medicine, College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China.

Vincent NoireauxSchool of Physics and Astronomy, University of Minnesota, Minneapolis, MN, 55455, USA. noireaux@umn.edu.

Steven BowdenDepartment of Food Science and Nutrition, University of Minnesota, Saint Paul, MN, 55108, USA. sbowden@umn.edu.ORCID https://orcid.org/0000-0001-5438-2532

Funding

Human Frontier Science Program LT0007/2024-LNational Science Foundation CBET-FMRG-2228971

6 · The paper itself

Abstract

backgroundCell-free gene expression (CFE) systems provide a rapid and modular platform for synthesizing bacteriophages without the need for living host cells. However, the generalizability of CFE-based phage synthesis across diverse phages, as well as the functional comparability of in vitro-synthesized phages to host-derived counterparts, has not yet been fully explored. In this study, we evaluated the ability of an Escherichia coli-based CFE platform to synthesize diverse bacteriophages relevant to food safety and synthetic biology applications.

resultsUsing an E. coli-based CFE system, we synthesized seven phages from four different families, achieving infectious titers ranging from 10⁶ to 10¹¹ plaque-forming units per milliliter (PFU/mL). Of these seven phages, five phages, vB_SalM-LPST153 (LPST153), vB_SenM-S16 (S16), SP6, vB_Sens_Jbel (Jbel), and vB_EcoM_Alf5 (Alf5), are reported here for the first time as successfully synthesized and characterized in a CFE system. CFE-synthesized phages remained capable of infecting host strains; however, significant differences in efficiency of plating (EOP) were observed across multiple Salmonella, E. coli, and Shigella hosts. Long-term storage tests for LPST153 showed that phage buffer (PB) stably maintained CFE titers above 10⁹ PFU/mL for 12 weeks, while purified CFE-synthesized and host-derived LPST153 lysates exhibited broadly comparable thermal stability profiles across the tested temperatures. Using the PHEIGES workflow, we also demonstrated complete in vitro assembly and expression of LPST153 from PCR fragments. In addition, phages were successfully synthesized using CFE lysates derived from the endotoxin-free E. coli strain ClearColi™, demonstrating the synthesis of infectious phages in an endotoxin-free system.

conclusionsThese findings expand the diversity of bacteriophages compatible with cell-free synthesis (CFS) and demonstrate that CFE-synthesized phages remain infectious and retain several functional properties, although some differences in host-dependent infectivity were observed. The ability to synthesize infectious phages in endotoxin-free systems and assemble phage genomes entirely in vitro highlights the potential of CFE platforms for scalable, decentralized, and cold-chain-independent manufacturing of phage-based therapeutics and food safety interventions.

Indexed as

BacteriophagesEscherichia coliSalmonellaShigellaCell-Free SystemBacteriophageCell-free expression (CFE)Cell-free synthesis (CFS)ClearColi™Endotoxin-free lysateEscherichia coliPhage biomanufacturingSalmonella entericaSynthetic biology

Identifiers

PMID42380887

DOI10.1186/s12934-026-03049-z

PMCPMC13591810

What OpenQuestion holds

Textmetadata

Read underepoch 390

Registered trials

None linked

Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.