Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis

L. Chen; E Wesdorp; Myrthe Jager; Esther Siegers; Mathijs Theelen; Nicolle J M Besselink; C. Vermeulen; Aldert Zomer; Els Broens; Jaap Wagenaar; Jeroen de Ridder

doi:10.1016/j.isci.2025.114005

Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis

L. Chen
, E Wesdorp
, Myrthe Jager
, Esther Siegers
, Mathijs Theelen
, Nicolle J M Besselink
, C. Vermeulen
, Aldert Zomer
, Els Broens
, Jaap Wagenaar
, Jeroen de Ridder^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

Abstract

Sepsis is the leading cause of death in neonatal foals, yet current diagnostics lack sufficient sensitivity and specificity. Here, we present a foal cell-free DNA (cfDNA) sequencing for bacterial identification (cfFBI) workflow that integrates wet-lab and computational protocols, enabling direct bacterial profiling through enrichment of the bacterial cfDNA and minimization of false-positive detections. We applied cfFBI to blood from 25 hospitalized foals and 7 healthy foals (H). Sepsis-associated bacterial genera were elevated in all 11 nSIRS-positive (S+) foals compared to H, and in 8/11 when compared to both nSIRS-negative (nS-) and H, with multiple genera elevated in nearly half (45.5%). While total cfDNA concentration, bacterial fraction, and microbial diversity did not differ between groups, S+ foals showed distinct cfDNA end-motif patterns and reduced mitochondrial cfDNA fractions. These findings indicate that cfDNA sequencing enables the detection of pathogenic bacteria and can help identify additional (host-related) sepsis biomarkers.

Original language	English
Article number	114005
Number of pages	17
Journal	iScience
Volume	28
Issue number	12
Early online date	11 Nov 2025
DOIs	https://doi.org/10.1016/j.isci.2025.114005 https://doi.org/10.1016/j.isci.2025.114005
Publication status	Published - 19 Dec 2025

Bibliographical note

Publisher Copyright:
© 2025 The Author(s)

Funding

This study is supported by the funding of Stichting Vrienden Diergeneeskunde, the Faculty of Veterinary Medicine, Utrecht University, and a Vidi Fellowship (639.072.715) to JdR from the Dutch Research Council (Nederlandse Organisatie voor Wetenschappelijk Onderzoek, NWO) . We thank Rhoxane Korthals from Dierenkliniek Emmeloord for her assistance with sample collection. We thank Joost van Rosmalen for his valuable input on statistical analysis. We acknowledge the Utrecht Sequencing Facility (USEQ) for providing sequencing services and data. USEQ is subsidized by the University Medical Center Utrecht and The Netherlands X-omics Initiative (NWO project 184.034.019) . We thank Marc Pages-Gallego and Dieter Stoker for proofreading the article.

Funders	Funder number
Stichting Vrienden Diergeneeskunde
Faculty of Veterinary Medicine
Utrecht University
Vidi Fellowship from the Dutch Research Council (Nederlandse Organisatie voor Wetenschappelijk Onderzoek, NWO)	639.072.715
Netherlands X-omics Initiative (NWO)	184.034.019

Keywords

Equine microbiology
Equine pediatric medicine
Microbial genomics

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Bacterial cell-free DNA profiling reveals the co-elevation of multiple bacteria in newborn foals with suspected sepsisFinal published version, 2.25 MBLicence: CC BY

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Chen, L., Wesdorp, E., Jager, M., Siegers, E., Theelen, M., Besselink, N. J. M., Vermeulen, C., Zomer, A., Broens, E., Wagenaar, J., & Ridder, J. D. (2025). Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis. iScience, 28(12), Article 114005. https://doi.org/10.1016/j.isci.2025.114005, https://doi.org/10.1016/j.isci.2025.114005

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abstract = "Sepsis is the leading cause of death in neonatal foals, yet current diagnostics lack sufficient sensitivity and specificity. Here, we present a foal cell-free DNA (cfDNA) sequencing for bacterial identification (cfFBI) workflow that integrates wet-lab and computational protocols, enabling direct bacterial profiling through enrichment of the bacterial cfDNA and minimization of false-positive detections. We applied cfFBI to blood from 25 hospitalized foals and 7 healthy foals (H). Sepsis-associated bacterial genera were elevated in all 11 nSIRS-positive (S+) foals compared to H, and in 8/11 when compared to both nSIRS-negative (nS-) and H, with multiple genera elevated in nearly half (45.5\%). While total cfDNA concentration, bacterial fraction, and microbial diversity did not differ between groups, S+ foals showed distinct cfDNA end-motif patterns and reduced mitochondrial cfDNA fractions. These findings indicate that cfDNA sequencing enables the detection of pathogenic bacteria and can help identify additional (host-related) sepsis biomarkers.",

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Chen, L, Wesdorp, E, Jager, M, Siegers, E , Theelen, M, Besselink, NJM, Vermeulen, C, Zomer, A , Broens, E , Wagenaar, J & Ridder, JD 2025, 'Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis', iScience, vol. 28, no. 12, 114005. https://doi.org/10.1016/j.isci.2025.114005, https://doi.org/10.1016/j.isci.2025.114005

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AU - Chen, L.

AU - Wesdorp, E

AU - Jager, Myrthe

AU - Siegers, Esther

AU - Theelen, Mathijs

AU - Besselink, Nicolle J M

AU - Vermeulen, C.

AU - Zomer, Aldert

AU - Broens, Els

AU - Wagenaar, Jaap

AU - Ridder, Jeroen de

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N2 - Sepsis is the leading cause of death in neonatal foals, yet current diagnostics lack sufficient sensitivity and specificity. Here, we present a foal cell-free DNA (cfDNA) sequencing for bacterial identification (cfFBI) workflow that integrates wet-lab and computational protocols, enabling direct bacterial profiling through enrichment of the bacterial cfDNA and minimization of false-positive detections. We applied cfFBI to blood from 25 hospitalized foals and 7 healthy foals (H). Sepsis-associated bacterial genera were elevated in all 11 nSIRS-positive (S+) foals compared to H, and in 8/11 when compared to both nSIRS-negative (nS-) and H, with multiple genera elevated in nearly half (45.5%). While total cfDNA concentration, bacterial fraction, and microbial diversity did not differ between groups, S+ foals showed distinct cfDNA end-motif patterns and reduced mitochondrial cfDNA fractions. These findings indicate that cfDNA sequencing enables the detection of pathogenic bacteria and can help identify additional (host-related) sepsis biomarkers.

AB - Sepsis is the leading cause of death in neonatal foals, yet current diagnostics lack sufficient sensitivity and specificity. Here, we present a foal cell-free DNA (cfDNA) sequencing for bacterial identification (cfFBI) workflow that integrates wet-lab and computational protocols, enabling direct bacterial profiling through enrichment of the bacterial cfDNA and minimization of false-positive detections. We applied cfFBI to blood from 25 hospitalized foals and 7 healthy foals (H). Sepsis-associated bacterial genera were elevated in all 11 nSIRS-positive (S+) foals compared to H, and in 8/11 when compared to both nSIRS-negative (nS-) and H, with multiple genera elevated in nearly half (45.5%). While total cfDNA concentration, bacterial fraction, and microbial diversity did not differ between groups, S+ foals showed distinct cfDNA end-motif patterns and reduced mitochondrial cfDNA fractions. These findings indicate that cfDNA sequencing enables the detection of pathogenic bacteria and can help identify additional (host-related) sepsis biomarkers.

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KW - Equine pediatric medicine

KW - Microbial genomics

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JF - iScience

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Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis

Abstract

Bibliographical note

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Keywords

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