A dual role for PGLYRP1 in host defense and immune regulation during B. pertussis infection

eLife Assessment

Rickert and colleagues demonstrate that the host peptidoglycan-binding protein PGLYRP1 has both beneficial and detrimental effects on Bordetella pertussis infection in mice. Using a solid array of techniques, the study provides useful insights into how the peptidoglycan fragment tracheal cytotoxin alters host immune responses, dampening inflammatory responses later in B. pertussis infection. These studies indicate that release of peptidoglycan fragments with particular structures can be used by bacteria to modulate NOD1 versus NOD2 responses to their advantage.

https://doi.org/10.7554/eLife.108947.3.sa0

Significance of the findings:

Useful: Findings that have focused importance and scope

Landmark
Fundamental
Important
Valuable
Useful

Strength of evidence:

Solid: Methods, data and analyses broadly support the claims with only minor weaknesses

Exceptional
Compelling
Convincing
Solid
Incomplete
Inadequate

During the peer-review process the editor and reviewers write an eLife Assessment that summarises the significance of the findings reported in the article (on a scale ranging from landmark to useful) and the strength of the evidence (on a scale ranging from exceptional to inadequate). Learn more about eLife Assessments

Abstract
Introduction
Results
Discussion
Materials and methods
Data availability
References
Article and author information
Metrics

Abstract

Bordetella pertussis, the etiologic agent of whooping cough, remains a serious public health concern despite widespread vaccination. Improved therapeutics and vaccines are urgently needed. Host recognition of bacterial peptidoglycan (PGN), including B. pertussis extracellular PGN fragment tracheal cytotoxin (TCT) shapes the immune response to infection. Peptidoglycan recognition proteins (PGLYRPs) are a conserved innate immune family that bind bacterial PGN and are primarily known for bactericidal activity in mammals; however, their immune modulatory roles are beginning to gain appreciation. The role of PGLYRPs in mammalian host defenses to Gram-negative pathogens, such as B. pertussis, remains largely unknown. Here, using knockout mice, single-cell and bulk transcriptomics, and functional assays, we identify a dual role for PGLYRP1 in modulating host immune responses to B. pertussis. PGLYRP1 contributes to antibacterial responses and paradoxically dampens inflammatory responses and inhibits bacterial killing later in infection. Mechanistically, PGLYRP1 enhances NOD1 signaling in response to TCT while suppressing NOD2− and triggering receptor expressed on myeloid cells-1 (TREM-1)-mediated inflammatory pathways. TCT-bound PGLYRP1 selectively impairs TREM-1 activation compared to PGNs from other bacteria. These findings demonstrate that B. pertussis co-opts PGLYRP1 to alter immune signaling, revealing a novel immune evasion mechanism with implications for vaccine design and host-directed therapeutics.

Introduction

Bordetella pertussis is a highly contagious respiratory pathogen and the causative agent of whooping cough, a disease characterized by severe, prolonged coughing and airway inflammation (Johnston et al., 1983; Paddock et al., 2008; Birkebaek et al., 1999). Despite widespread vaccination efforts, B. pertussis remains a public health concern due to waning vaccine-induced immunity and the pathogen’s capacity to evade host immune defenses (Wang et al., 2025). Host detection of microbial components, including peptidoglycan (PGN) fragments, is essential for initiating immune responses to infection. Understanding how these host–pathogen interactions shape disease outcomes is critical for developing improved strategies to fight infections.

PGN is a fundamental component of the bacterial cell wall, composed of repeating disaccharide chains cross-linked by short peptides (Schleifer and Kandler, 1972). PGN structure is an important determinant of innate immune recognition and responses (Sanchez et al., 2017; Wolf and Underhill, 2018). During cell wall turnover, some pathogenic bacterial species, including B. pertussis, release PGN fragments into the extracellular environment (Rosenthal et al., 1987; Melly et al., 1984; Jutras et al., 2019). The biological roles of these fragments in infection and immunity are poorly defined.

B. pertussis releases a disaccharide-tetrapeptide monomeric PGN fragment known as tracheal cytotoxin (TCT) (Cookson et al., 1989; Goldman et al., 1982). TCT differs from B. pertussis cell wall PGN by having a 1,6-anhydro group on its MurNAc sugar due to incomplete PGN turnover (Cookson et al., 1989). The functional consequences of this 1,6-anhydro MurNAc modification are poorly understood (Torrens and Cava, 2024). TCT is recognized by the cytosolic pattern recognition receptor (PRR) NOD1 to activate NF-κB-driven immune responses (Magalhaes et al., 2005; Chamaillard et al., 2003). In vitro and ex vivo studies suggest that recognition of TCT by NOD1 in non-ciliated mucus-secreting cells leads to the accumulation of nitric oxide within neighboring ciliated airway epithelial cells and subsequent ciliostasis and extrusion of ciliated cells (Kessie et al., 2020; Flak and Goldman, 1999). We recently demonstrated that TCT has broader immunomodulatory effects, skewing PGN-sensing toward NOD1 and away from NOD2, thereby dampening a stronger NOD2-dependent, pro-inflammatory cytokine production and generation of immune memory (Rickert et al., 2025). However, the broader immunological consequences of TCT release during infection, especially in the context of PGN-sensing by PRRs, remain poorly defined.

Peptidoglycan recognition proteins (PGLYRPs) are a conserved family of soluble extracellular innate immune receptors that bind PGN (Liu et al., 2001). In arthropods, PGRPs act as extracellular PRRs that trigger downstream immune pathways (Dziarski and Gupta, 2006a). In mammals, they have been characterized as bactericidal proteins that target PGN on Gram-positive bacterial surfaces (Kashyap et al., 2011; Dziarski and Gupta, 2006b). However, recent studies have expanded their role beyond antimicrobial defense, suggesting function in regulating host immune responses. PGLYRP1 is a secreted protein stored in neutrophil granules that is directly bactericidal against several bacterial species (Lu et al., 2006). Recent work suggests that mammalian PGLYRPs also regulate adaptive immunity, including CD8+ T cell responses, in autoimmunity and cancer, and act as an accessory protein for the activation of inflammation amplifying receptor TREM-1 and NOD2-dependent signaling (Chen et al., 2025; Schnell et al., 2023; Read et al., 2015).

TREM-1 is a PRR that amplifies inflammatory responses during infection (Arts et al., 2013). Activation of TREM-1 by its ligands, including PGLYRP1–PGN complexes, enhances production of pro-inflammatory cytokines and chemokines (Bouchon et al., 2000). Excessive TREM-1 activation is associated with inflammatory pathology in sepsis and other diseases (Bouchon et al., 2001). Despite growing interest in PGLYRPs, their roles in host–pathogen interactions, particularly in the context of Gram-negative bacteria and secreted PGN fragments, remain poorly defined.

Here, we demonstrate that PGLYRP1 plays a dual role in host responses to B. pertussis infection, contributing to early bactericidal responses and fine-tuning inflammation in response to distinct PGN structures. We demonstrate that exogenous PGLYRP1 enhances NOD1 activation by TCT but fails to enhance muramyl dipeptide (MDP) activation of NOD2. Further, PGLYRP1 directly activates the pro-inflammatory receptor TREM-1, a response that is potentiated by sacculus-derived PGN from Gram-positive bacteria Staphylococcus aureus or Bacillus subtilis. PGLYRP1-mediated activation of TREM-1 was not supported by B. pertussis extracellular PGN monomer TCT. These findings reveal that PGLYRP1 not only contributes to antibacterial responses but suggest it may also be a context-specific immunomodulator whose function is shaped by the PGN it encounters. This work uncovers a previously unrecognized mechanism by which extracellular PGN and innate receptors together shape host inflammation during infection in a structure-specific manner.

Results

Opposing roles of PGLYRP1 in early and late host defenses to B. pertussis

PGLYRP1 has known antibacterial activity in vitro, particularly against Gram-positive bacteria (Lu et al., 2006). To investigate whether PGLYRP1 contributes to host microbial defenses against B. pertussis, we intranasally infected adult BALB/c wild-type (WT) and PGLYRP1-knockout (PGLYRP1 KO) mice and assessed lung bacterial burden at 4 and 7 days post-infection (DPI). Surprisingly, PGLYRP1 had opposing impacts on bacterial burden at these time points (Figure 1A, B). At 4DPI, PGLYRP1 KO mice exhibited significantly higher bacterial burdens compared to WT controls (p < 0.01), indicating that PGLYRP1 contributes to early bacterial control. However, by 7DPI PGLYRP1 KO mice had significantly lower bacterial burden compared to WT mice (p < 0.001), suggesting that PGLYRP1 impairs bacterial clearance later in infection, potentially by regulating or modulating host immune responses.

Figure 1

Download asset Open asset

PGLYRP1 has a dual role in host defense against *Bordetella pertussis*.

Lung bacterial burdens in wild-type BALB/c (WT, black) and PGLYRP1-knockout (PGLYRP1 KO, red) at 4 (A) and 7 (B) days post-infection. Violin-plot visualizing single-cell expression levels of Pglyrp1 across cell types in the lungs of C57/B6 at 4DPI for both infected and sham-challenged mice, assessed via analysis of single-cell RNA sequencing data (C). RNAscope in situ hybridization visualization of *Pglyrp1* transcripts in FFPE sections of *B. pertussis*-infected mouse lung tissue (D). Each dot represents a single Pglyrp1 mRNA molecule. Images are presented at ×20 and ×10 magnification of ex vivo bone marrow-derived neutrophils (E). Neutrophils were isolated from BALB/c (WT) or PGLYRP1 KO mice and incubated with *B. pertussis* for 2 (circles) or 24 (square) hr, and CFU enumerated by serial dilution on Bordet Gengou agar. In vitro bacterial killing assay assessing recombinant murine PGLYRP1 (mPGLYRP1, red) bactericidal activity at 6- and 24-hr post-incubation compared to BSA control against WT (G) or a TCT non-releasing strain (TCT−) at 6 hr (H). Enumeration of CFU following incubation of mPGLYRP1 with wild-type *B. pertussis* (parental) or mutant lacking the *BpsB* gene of the *Bordetella* polysaccharide (Bps) operon for 6 hr (I). Data represent CFU as a percentage of starting inoculum recovered at indicated time point as assessed by serial dilution of culture on BG agar. Data are presented as mean ± SEM; significance determined by unpaired t-test (**A, B, F, G**) or ANOVA (**H, I**). **p-value <0.01, ***p-value <0.005, ****p-value <0.001.

To identify which cell types express PGLYRP1 following infection, we performed single-cell RNA sequencing on lungs from uninfected and B. pertussis-infected C57BL/6 mice at 4DPI. In uninfected lungs, PGLYRP1 expression was most prominent in neutrophils along with heterogeneous expression in some lymphocyte and inflammatory monocyte populations (Figure 1C). Upon infection, the PGLYRP1-expressing cell types expanded to include epithelial cells like ciliated cells, secretory cells, and AT1/AT2s in addition to granulocytes like basophils (Figure 1C). RNAscope in situ hybridization confirmed expression in the airways of infected mice and in neutrophils (Figure 1D, E).

To determine whether PGLYRP1 contributes to antibacterial responses to B. pertussis, we performed in vitro and ex vivo killing assays. Based on transcriptomic data (Figure 1C), we assessed the bactericidal potential of bone marrow-derived neutrophils (BMDNs) from WT and PGLYRP1 KO mice incubated with B. pertussis for 2 or 24 hr. At 24 hr, PGLYRP1 KO neutrophils failed to control viable bacteria compared to WT neutrophils (Figure 1F), indicating that PGLYRP1 contributes to neutrophil-mediated control of B. pertussis. In a cell-free assay, recombinant murine PGLYRP1 (mPGLYRP1) significantly reduced bacterial CFU after 24 hr of incubation compared to BSA control (Figure 1G, p < 0.001), validating its potential antibacterial role in neutrophil-mediated control of B. pertussis growth.

TCT release by B. pertussis is a product of inefficient recycling by the permease AmpG (Cookson et al., 1989). Replacing B. pertussis AmpG with E. coli AmpG resulted in a strain (TCT−) which releases 99% less TCT (TCT(−)), while deletion of AmpG created a strain with 24-fold greater TCT release (TCT(+)) (Lyon, 2001). Since PGLYRP1 binds bacterial PGN and the addition of excess PGN has been shown to prevent PGLYRP bactericidal activity (Lu et al., 2006), we hypothesized that B. pertussis TCT, an extracellular PGN fragment, may act as a decoy to divert PGLYRP1 activity. However, no increase in the bactericidal activity of recombinant mPGLYRP1 was noted between WT or TCT-deficient strains following incubation with PGLYRP1 (Figure 1H), arguing against this hypothesis. Given prior work showing that Bordetella polysaccharides confer resistance to antimicrobial peptides (Fullen et al., 2022), we tested whether extracellular polysaccharides inhibit PGLYRP1 function. The Bps operon is required for effective production of B. pertussis surface polysaccharide (Parise et al., 2007). Using a BpsB-deficient strain, we observed significantly reduced resistance to mPGLYRP1 activity compared to WT bacteria (Figure 1I, p < 0.01), suggesting that endogenous polysaccharides can shield B. pertussis from PGLYRP1 activity.

Together, these results indicate that PGLYRP1 plays a dual role during B. pertussis infection. PGLYRP1 enhances early bacterial control via neutrophil-mediated killing and direct bacteriostatic activity but may also play a deleterious role in controlling pulmonary infection at later stages of infection (Figure 1). While TCT does not appear to interfere with PGLYRP1 activity, Bordetella polysaccharides, previously associated with biofilm production (Parise et al., 2007), do, highlighting a key bacterial evasion strategy against host innate defenses.

PGN fragment TCT enhances PGLYRP1-mediated suppression of host responses in pertussis

Given the opposing effects of PGLYRP1 on bacterial burden at 4 and 7DPI, we hypothesized that PGLYRP1 exerts antibacterial activity at early time points but may suppress protective immune responses or enhance deleterious responses later in infection, impairing bacterial control by the host. Recently, PGLYRP1 has been associated with PGN recognition by innate immune receptors, including NOD2 and TREM-1 (Chen et al., 2025; Read et al., 2015). However, B. pertussis releases a NOD1-activating PGN fragment, TCT (Magalhaes et al., 2005).

To assess the role of PGLYRP1 in B. pertussis-driven inflammation, we performed histological analysis of lungs from WT and PGLYRP1 KO mice at 4 and 7DPI. We observed significantly reduced immunopathology, based on the percentage and severity of bronchovascular bundle formation and infiltration of immune cells into alveolar spaces, in PGLYRP1 KO mice at both time points (Figure 2A, p < 0.05 at 4DPI, p < 0.01 at 7DPI). These findings suggest that PGLYRP1 contributes to inflammatory lung damage during B. pertussis infection.

Figure 2

Download asset Open asset

Assessment of PGLYRP1’s role in pulmonary inflammation and transcriptional responses during *Bordetella pertussis* infection.

Semi-quantitative scoring of hematoxylin and eosin (H&E) stained lung sections from wild-type BALB/c (black dots) and PGLYRP1 knockout (red dots) mice at 4 and 7 days post-infection with *B. pertussis* (A). Data represents average scores of three blinded investigators scoring degree and percentage of peribronchial infiltration and alveolar consolidation. Bulk RNA sequencing was performed on lung homogenates collected from WT and PGLYRP1 KO mice at 4DPI to assess differential gene expression. Volcano plots (**B, D**) and heat-maps (**C, E**) showing total (**B, D**) and immune-related (**C, E**) differentially expressed genes between *B. pertussis* challenged BALB/c (WT) and PGLYRP1 KO mice (**B, C**) or PGLYRP1 KO mice challenged with parental wild-type *B. pertussis* (WT) or TCT-under releasing strain TCT(−). (F) KEGG enrichment pathway analysis was performed on differentially expressed genes from lung tissue isolated from mice challenged with WT or TCT(−) *B. pertussis*. Data represent the mean of biological replicates; statistical analyses were performed by Mann–Whitney U test. *p-value = 0.003, **p-value <0.01.

To investigate PGLYRP1’s role in immune signaling during B. pertussis infection, we performed bulk RNA sequencing on lungs from infected BALB/c (WT) and PGLYRP1 KO mice. PGLYRP1 KO mice exhibited increased expression of pro-inflammatory cytokines (e.g., Il6, Il23a, Il1a, Il1b, and Il36g) and chemokines (e.g., Ccl2, Ccl3, Ccl12, and Ccl28) alongside reduced expression of anti-inflammatory cytokine IL10 (Figure 2B, C). This transcriptomic profile suggests a model in which PGLYRP1 dampens key cytokine responses while paradoxically promoting immunopathology.

PGLYRP1 is a soluble PGN-binding protein. B. pertussis releases an extracellular PGN fragment, TCT. We have recently demonstrated that TCT inhibits immune responses to B. pertussis (Rickert et al., 2025). We hypothesized that interactions with TCT contribute to PGLYRP1’s influence on host inflammatory responses to B. pertussis. To test this, we compared the transcriptional responses of PGLYRP1 KO mice challenged with TCT-producing (parental) or TCT-deficient B. pertussis strains. Both strains induced a similar pattern of cytokine upregulation in the absence of PGLYRP1, but this increased inflammation was more robust in the absence of TCT (Figure 2D, E). Specifically, PGLYRP1 KO mice infected with TCT(−) bacteria exhibited greater expression of Il23a, Il6, Il1b, and Il36g compared to WT infection, suggesting that TCT may potentiate the immune-dampening activity of PGLYRP1.

PGLYRP1 has previously been shown to stimulate inflammatory responses to PGN (Read et al., 2015). These findings indicate that the immunomodulatory function of PGLYRP1 is informed by the PGN it interacts with. TCT release may be a strategy employed by B. pertussis to co-opt PGLYRP1 function and suppress protective inflammation, thereby promoting pathogen persistence. Supporting this, KEGG pathway analysis on differential gene expression of 4DPI WT (parental) and TCT(−) lungs confirmed enrichment of cytokine–cytokine receptor interactions, NOD signaling, and IL-17/TNF pathways in the presence of TCT (Figure 2F).

TCT–PGLYRP1 interactions drive NOD1 signaling

To understand how TCT modulates PGLYRP1-dependent immune signaling, we investigated known mechanisms by which PGLYRP1 influences host PRR signaling. Recent studies suggest that intracellular PGLYRP1 can act as a co-factor in NOD2 signaling (Chen et al., 2025), and TCT is a known NOD1 agonist. Hence, we hypothesized that PGLYRP1 may enhance NOD1 signaling in response to the muropeptide TCT, a NOD1 exclusive agonist. To test this, recombinant PGLYRP1 was incubated with TCT (1 ng/ml) before stimulation of NOD1 and NOD2 reporter cells. Consistent with previous reports, TCT robustly stimulated a NOD1 response (Figure 3A). Notably, both murine (Figure 3A, p < 0.005) and human (Figure 3B, p < 0.001) PGLYRP1 significantly enhanced TCT-mediated NOD1 activation compared to TCT alone. Importantly, PGLYRP1 alone did not induce NOD1 activation (Figure 3A), suggesting the enhanced NOD1 stimulation was not by PGLYRP1 directly activating NOD1 (Figure 3A). Interestingly, this enhanced activation was specific to NOD1 as neither TCT nor TCT + PGLYRP1 activated NOD2 reporter cells (data not shown) since TCT is not a NOD2 agonist. On the other hand, mPGLYRP1 decreased NOD2 recognition of NOD2 agonist MDP (p < 0.01; Figure 3C), suggesting exogenous PGLYRP1 boosts responses to NOD1 stimulating PGN structures like TCT but dampens responses to NOD2 stimulatory PGNs.

Figure 3 with 1 supplement see all

Download asset Open asset

PGLYRP1 selectively modulates NOD1 and NOD2 signaling in response to *Bordetella pertussis* peptidoglycan.

HEK293 reporter cells expressing either mouse (**A, D**) or human (B) NOD1 or (C) NOD2 were stimulated with recombinant mouse (A) or murine or human (**B–D**) PGLYRP1 (12.5 µg/ml), purified tracheal cytotoxin (TCT, 1 ng/ml) (**A, B**), muramyl dipeptide (MDP, NOD2 agonist) (C), or conditioned bacterial growth media (OD = 0.8) (D). NF-κB-driven SEAP reporter activity was measured to assess pathway activation. Reporter activation was quantified after 18–24 hr and normalized to PBS-treated cell controls as a zero baseline. Single-cell RNA sequencing was performed on lung tissue from BALB/c mice infected with *B. pertussis* at 4 days post-infection. Neutrophils were subjected to further sub-clustering and visualized as a UMAP (E). Violin plots visualizing expression of Nod1, Nod2, IL1a, and Pglyrp1 across neutrophil sub-clusters (F). Volcano plot demonstrating differentially expressed genes in Nod1 (blue) or Nod2 (red) expressing neutrophils (G) Single-cell analyses represent combined data from two mice per group. Statistical analyses were performed using Student’s t-test or adjusted p-values as appropriate. **p-value <0.01, ***p-value <0.005, ****p-value <0.001.

To determine if PGLYRP1 modulates NOD responses to other potential PGNs released by B. pertussis, we stimulated reporter cells with mPGLYRP1 incubated with conditioned media from WT or TCT(−), or TCT(+) cultures. Consistent with our results using purified TCT, mPGLYRP1 amplified NOD1 signaling in response to B. pertussis conditioned media, likely due to increased recognition of extracellular TCT (Figure 3D, p < 0.01). In contrast, NOD2 signaling was significantly suppressed by mPGLYRP1 (Figure 3D), consistent with our results with purified PGN (Figure 3A–C). Together, these findings suggest that exogenous PGLYRP1 acts as a selective modulator of NOD signaling, enhancing NOD1 activation while dampening NOD2 responses.

To assess the functional consequence of skewed Nod1 versus Nod2 expression and signaling during infection, we analyzed single-cell RNA sequencing data from lungs of BALB/c mice at 4DPI. We examined NOD1 and NOD2 expression across neutrophil sub-clusters (Figure 3E, F), as neutrophils are the primary expressors of PGLYRP1 during B. pertussis infection (Figure 1C). NOD2-expressing neutrophils showed higher expression of pro-inflammatory genes including Il1a, Ccl3 and Ptgs2 (COX-2) (Figure 3F, G). In contrast, NOD1 expression correlated with genes involved in anti-inflammatory immune responses: Pla2g7 (degrades platelet activating factor), Tmsb4x, which inhibits neutrophil migration, promotes repair, and inhibits NF-κB, and increased expression of PGLYRP1, which may further positively regulate these pathways (Xie et al., 2020). This identifies a correlation between pro-inflammatory cytokine expression and NOD2 signaling in neutrophils.

Interestingly, when we further analyzed the clusters with high Nod2 expression, like clusters 0 and 4, we observed gene expression patterns which correspond to previously identified inflammatory neutrophil populations. Cluster 0 hallmark genes were associated with interferon-stimulated programming (Calhm6, Cxcl9, Tma16, Slamf7, and Baft2) (Figure 3—figure supplement 1), and B. pertussis infection has been shown to cause both deleterious and beneficial interferon responses (Xie et al., 2020; Ardanuy et al., 2020; Ardanuy et al., 2023). Additionally, poor B. pertussis host-responses have been linked to interferon-stimulated neutrophils, providing interesting context to this identified neutrophil cluster (Kumar et al., 2025b; Kumar et al., 2025a). Furthermore, cluster 4 marker (Hilpda, Csf3, Myl4, Odc1, and Spp1) neutrophils have been identified in tumor exhaustion environments and related to hypoxia response, highlighting another inflammatory neutrophil population (Figure 3—figure supplement 1; Veglia et al., 2021).

These results highlight the heightened inflammatory phenotype of NOD2+ neutrophils and suggest that TCT-mediated skewing of NOD responses toward NOD1, amplified by PGLYRP1, may be a mechanism to suppress inflammation and promote B. pertussis persistence.

TCT–PGLYRP1 complexes dampen TREM-1 activation, limiting inflammation during B. pertussis infection

In addition to their role in NOD signaling, PGLYRP1–PGN complexes have been shown to activate TREM-1, driving NF-κB-mediated inflammatory responses (Read et al., 2015). This ability to activate TREM-1 is boosted by the presence of a PGN from the sacculi of S. aureus, B. subtilis, or E. coli (Read et al., 2015). To determine if TREM-1 contributes to host responses to B. pertussis, we first assessed TREM-1 expressing cell types following B. pertussis infection in mice using single-cell RNA sequencing. Following B. pertussis infection, TREM-1 was primarily expressed by neutrophils, with lesser expression noted in monocytes and macrophages (Figure 4A).

Figure 4

Download asset Open asset

TCT inhibits PGLYRP1 signaling via TREM-1.

Violin plot generated from single-cell RNA sequencing (scRNA-seq) displaying TREM-1 expression on neutrophils (green), alveolar macrophages (AM, gold), and inflammatory monocytes (IM) at 4DP1 (A). TREM-1 activation measured via NFAT-driven luminescence (relative luminescence units, RLU) in TREM-1 reporter cells treated with recombinant human PGLYRP1 with or without 5 µg/ml of Lys-type peptidoglycan derived from *Staphylococcus aureus* (5 µg/ml), TCT (5 µg/ml), or a DAP-type PGN from *Bacillus subtilis* (5 µg/ml) (B). Diagram depicting proposed model (C). All TREM-1 activation assays were performed with biological triplicates (n = 3). Statistical analyses were performed using unpaired two-tailed Student’s t-test. *p-value <0.05, **p-value <0.01, ***p-value <0.005, ****p-value <0.001.

To determine if TCT influences PGLYRP1-dependent TREM-1 activation, we assessed the ability of TCT, a DAP-type PGN monomer, to enhance PGLYRP1-mediated TREM-1 activation. As reported previously, preparations of PGNs isolated from S. aureus and B. subtilis enhanced PGLYRP1-mediated activation of a TREM-1 reporter cell line (Figure 4B, p < 0.01) (Tocheva et al., 2013; Sharif et al., 2009). Given the anti-inflammatory role of TCT observed in our data (Rickert et al., 2025 and Figures 2 and 3), and known role for TREM-1 in B. pertussis pathogenesis (Gallop et al., 2021), we proposed that TCT may be a poorer enhancer of PGLYRP-1-mediated TREM-1 activation. Consistent with our hypothesis, the addition of TCT significantly reduced TREM-1 activity (Figure 4C). This suggests that TCT, unlike PGN from the sacculi, fails to support and possibly even antagonizes PGLYRP1’s ability to activate TREM-1 (Figure 4C). Further studies are required to discern the contribution of PGN structure and solubility in this model. Combined with our previous work (Gallop et al., 2021), these findings highlight the importance of PGN in bacterial regulation of TREM-1 and its role in B. pertussis pathogenesis and disease, and the potential impact of skewing TREM-1 responses.

Discussion

Our study reveals a previously unrecognized role for the PRR, PGLYRP1, in orchestrating both antibacterial defenses and inflammatory responses during B. pertussis infection. While traditionally characterized as a bactericidal effector which mediates its effects by binding PGN on the surface of Gram-positive bacteria, emerging work suggests its role may extend to mammalian immune signaling (Schnell et al., 2023). The contribution of PGLYRP1 to PRR-mediated inflammatory responses in mammals, and in particular how PGN structure impacts the outcomes of these interactions, remains largely unexplored. Our findings demonstrate that PGLYRP1 can function as a PGN structure-specific interpreter which translates structure into context and pathogen-specific immune signaling. This dual role expands our understanding of how innate immune receptors fine-tune host–pathogen interactions in the airway based on the structure of PGN encountered. Prior studies have also shown that endogenous PGLYRP1 bolstered NOD2-driven macrophage activation in response to GMTriP-K, but not in response to related PGN fragments such as GMDiP or MDP, highlighting the structural specificity of PGLYRP1-mediated immune recognition (Chen et al., 2025). We expand on these findings to suggest exogenous PGLYRP1 biases NOD1 responses and dampens NOD2 responses. Furthermore, we suggest pathogens like B. pertussis have evolved to release PGN structures, such as TCT, which dampen the ability of PGLYRP1 to signal via TREM-1 (Figure 4C), suggesting an active area of ongoing host–pathogen evolution.

Here, we propose PGLYRP1 as a PGN-structure-specific regulator of inflammatory responses. This is comparable to SIGLECs, which can distinguish self from non-self glycans to ignore or elicit immune responses (Paulson et al., 2012). During B. pertussis infection, PGLYRP1, predominantly expressed by neutrophils, contributes to early bacterial clearance, as PGLYRP1 KO mice exhibit significantly higher bacterial loads at 4DPI. The bactericidal activity of PGLYRP1 is diminished by the presence of extracellular polysaccharide, highlighting the potential for B. pertussis-derived extracellular decoys to prevent antibiotic activity. PGLYRP1 KO mice exhibit elevated inflammatory cytokine expression, suggesting PGLYRP1 also serves an anti-inflammatory function. Transcriptomic analysis of PGLYRP1 KO lungs infected with mutants which fail to release TCT or the parental strain supports the hypothesis that PGLYRP1 limits inflammation in a PGN structure-dependent manner: specifically, the monomeric PGN fragment TCT potentiates the anti-inflammatory activity of PGLYRP1. An alternative explanation for the discrepancy observed in bacterial burden at early and later time points of infection is that PGLYRP1-directed lysis of B. pertussis releases more host-available muropeptides, which PGLYRP1 can then interact with to enhance less protective NOD1-driven immune responses at the expense of more potent NOD2-driven responses (Figure 1; Rickert et al., 2025).

Mechanistically, PGLYRP1 enhances NOD1 signaling in response to TCT while suppressing NOD2 signaling in response to MDP, directing the polarization of immune responses away from a more inflammatory NOD2-driven response. Single-cell RNA sequencing revealed that PGLYRP1 expression is enriched in NOD1+ neutrophils, which have less inflammatory profiles than NOD2+ neutrophils. These data suggest B. pertussis may exploit PGLYRP1 to amplify a NOD1-dominant signaling environment that blunts host inflammation. Recent work from our group highlighted the ability of TCT to polarize NOD responses toward NOD1 as an immune evasion mechanism (Rickert et al., 2025). This work expands on this finding, highlighting the role of PGLYRP1 and PGN structure in these findings, but further work should validate and expand on how PGLYRP1 modulates responses to muropeptide outside the cellular and murine models used in these studies.

These findings shift our understanding of mammalian PGLYRP1 from a passive bactericidal protein to an immunomodulator, whose contribution to inflammation is determined by the structure of PGN it encounters. Additionally, they inform our views of PGN structure and how it impacts host responses, suggesting bacterial release of PGNs as a mechanism to dampen host immunity. These results propose a novel paradigm in host-pathogen interactions; that bacteria may release structurally tailored PGN fragments to bias host immune sensors and signaling in their favor.

Excitingly, we report a novel axis by which PGLYRP1 regulates TREM-1, a known amplifier of inflammation. TREM-1 activation has been reported to strengthen NF-kB signaling pathways which NLRs like NOD1 and NOD2 canonically signal through (Arts et al., 2013). While PGLYRP1 synergized with PGNs from S. aureus and a B. subtilis to enhance TREM-1 activation, the presence of TCT antagonized this effect (Figure 4). There are several differences in the structures of these PGNs: unlike monomeric TCT, PGN tested from other bacteria are derived from the sacculus and are insoluble; additionally, B. subtilis PGN is amidated. Further studies are required to discern if solubility, size, or other factors are responsible for differences in the effect on PGLYRP1-mediated TREM-1 activation. Additionally, TCT contains a 1,6-anhydro MurNAc. Bifidobacterium have been predicted to produce abundant anhydro-PGNs which markedly reduce LPS-stimulated cytokine expression in RAW264.7 cells, underscoring a potential anti-inflammatory effect of 1,6-anhydro-MurNAc containing PGNs, such as TCT.

Previous work from our group demonstrated that inhibition of TREM-1 prevented immunopathology and inhibited cytokine and chemokine responses to infection, highlighting the potential utility of disturbed TREM-1 signaling for B. pertussis (Gallop et al., 2021). TREM-1 inhibitor treated mice showed reduced inflammation and inflammatory cytokine responses following B. pertussis infection, highlighting the importance of this pathway in pertussis pathogenesis and the potential utility of its manipulation by bacteria. Further, our findings suggest that selectively targeting TREM-1 or PGN structure sensing by PGLYRP1 could attenuate disease severity without compromising bacterial clearance, a strategy aligned with emerging host-directed therapeutics in TB and sepsis (Sigalov, 2014; Skerry et al., 2014; Dey et al., 2021). However, the role of these players in generating effective antibody responses still needs to be elucidated so that we can understand the implications on long-term immunity.

We hypothesize that B. pertussis may attenuate host immune responses not merely by evading recognition, but by actively hijacking host regulatory circuits through the release of specific PGN structures. Taken together, our findings support a model in which PGLYRP1 serves dual and temporally distinct roles during infection, by promoting bacterial killing while restraining inflammation in a PGN-structure-specific manner. This duality may reflect temporal or disease stage specific changes in the types of PGN available to host PRRs, as 1,6-anhydro MurNAc containing PGNs are usually internal, unless released following bacterial lysis. The ability of B. pertussis to exploit this shift by releasing TCT to suppress excessive inflammation represents a novel immune evasion strategy.

More broadly, this work demonstrates that the structure of PGN fragments is a key determinant of host immune outcomes. Innate receptors like PGLYRP1 relay this structural information to shape both defense and pathology, providing an avenue for pathogen hijacking by alternating the availability of different PGN structures. Targeting the TCT–PGLYRP1–TREM-1 axis may offer a novel therapeutic or vaccine-mediated approach to mitigate immunopathology in pertussis and potentially other infections involving secreted PGN. These findings reframe PGN as both a pathogen-associated molecular pattern and an immunomodulatory cue that both host and pathogen exploit to regulate the immune-inflammatory environment. Future studies using defined synthetic muropeptides will be necessary to pinpoint structural determinants of the reduced immune activation noted toward TCT. Likewise, how these alterations in PGN structure impact PGLYRP1 interactions with host PRRs remains unclear.

Materials and methods

Key resources table

Reagent type (species) or resource	Designation	Source or reference	Identifiers	Additional information
Cell line (HEK-293)	HEK-Blue mouse NOD1 reporter cells	Invivogen	hkb-mnod1	HEK-293, NF-κB-based SEAP reporter
Cell line (HEK-293)	HEK-Blue human NOD1 reporter cells	Invivogen	hkb-hnod1	HEK-293, NF-κB-based SEAP reporter
Cell line (HEK-293)	HEK-Blue mouse NOD2 reporter cells	Invivogen	hkb-mnod2	HEK-293, NF-κB-based SEAP reporter
Cell line (HEK-293)	HEK-Blue human NOD2 reporter cells	Invivogen	hkb-hnod2v2	HEK-293, NF-κB-based SEAP reporter
Cell line (Jurkat)	TREM-1 reporter cells	Eurofins/DiscoverX	793-1167C19	EFC-based TREM-1 activation reporter
Genetic reagent (Mus musculus)	PGLYRP1 KO	PMID:20709292		Provided generously by Dr. Roman Dziarski (Indiana University SOM)
Strain, strain background (Bordetella pertussis)	TCT(+)	Goldman Lab, UNC (Lyon, 2001)
Strain, strain background (B. pertussis)	TCT(−)	Goldman Lab, UNC (Lyon, 2001)
Strain, strain background (B. pertussis)	ΔbpsB	Deora Lab, OSU		Provided generously by Dr. Rajendar Deora (OSU)
Peptide, recombinant protein	Murine recombinant PGLYRP1	R&D Systems (Bio-Techne)	Cat#: 2696-PGB
Peptide, recombinant protein	Human recombinant PGLYRP1	R&D Systems (Bio-Techne)	Cat#: 2590-PGB
Biological sample (Bordetella pertussis)	Purified TCT	Goldman Lab, UNC	N/A	Isolated from bacterial culture in the Goldman lab
Chemical compound, drug	Purified Staphylococcus aureus peptidoglycan	Sigma-Aldrich	Cat#: SIG-77140-10MG
Chemical compound, drug	Purified Bacillus subtilis peptidoglycan	Sigma-Aldrich	Cat#: SIG-69554-10MG

Mice and ethics statement

Request a detailed protocol

All animal procedures were approved by the Institutional Animal Care and Use Committee at the University of Maryland School of Medicine (protocol 00000108). Adult (6–8 week-old) BALB/c (Charles River, 028) and PGLYRP1 knockout (Pglyrp1^tm1Rdz/Pglyrp1^tm1Rdz in BALB/c background, generously provided by Dr. Roman Dziarski Dziarski et al., 2003), mice were bred and housed under specific pathogen-free conditions. To control for and assess the potential of sex-related differences in pertussis pathogenesis, both male and female mice were used in experiments. For transcriptomic studies, RNA was isolated from the lungs of PBS or B. pertussis challenged female mice.

Bacterial strains and infections

Request a detailed protocol

B. pertussis strains, including wild-type BC36, a streptomycin-resistant Tohama I derivative, or isogenic mutants (TCT−, TCT+, BpsB KO) were cultured on Bordet–Gengou agar supplemented with 10% defibrinated sheep blood for in vivo infections or in Stainer–Scholte (SS) broth for in vitro assays. Strains were grown in SS broth to mid-log stage (OD = 0.8), and the resulting media was centrifuged and filtered to remove any bacteria to make the conditioned media used for cell culture assays. TCT mutants were generously supplied by William Goldman (University of North Carolina at Chapel Hill). TCT− strain was generated by expression of the E. coli AmpG in BC36, resulting in a 50-fold decrease in TCT release. TCT+ was generated by replacing native B. pertussis AmpG with a kanamycin resistance cassette; this resulted in a 24-fold increase in TCT release (Lyon, 2001). Adult mice were anesthetized and inoculated intranasally with 2 × 10⁶ CFU of B. pertussis in 50 µl PBS. Lungs were harvested at 4DPI and 7DPI for bacterial burden analysis by plating of serially diluted lung homogenates.

RNA isolation and processing

Request a detailed protocol

Lung tissue was harvested at 2 or 7DPI for RNA isolation by the Trizol-chloroform method. Briefly, lung tissue was homogenized using the Omni Bead Beader (Omni, Inc), phase separated using chloroform, and precipitated using isopropanol. RNA was then quantified and converted to cDNA (iScript cDNA Synthesis Kit). Quantitative real-time PCR was performed using the Bio-Rad CFX96 real-time PCR instrument. Gene expression was calculated as fold change relative to PBS-inoculated control animals using the 2^−ΔΔCT threshold cycle method and normalized to Hypoxanthine phosphoribosyltransferase (internal housekeeping gene).

Bulk RNA sequencing

Request a detailed protocol

At 4DPI, lungs were harvested from three WT or PGLYRP1 KO BALB/c mice infected with WT or TCT-deficient B. pertussis. Total RNA was extracted from homogenized lungs using Trizol Reagent (Invitrogen) according to the manufacturer’s protocol. RNA integrity was confirmed using a Bioanalyzer (Agilent); alignments were generated by HISAT2 sequencing performed using an Illumina NextSeq 1000. FastQC and RSeQC were used to assess read and alignment quality. Samtools was used to generate alignment statistics. Reads were aligned with the Mus musculus GRCm39 reference genome. Differential gene expression was analyzed using DESeq2, with significance defined as adjusted P-value <0.05.

Single-cell RNA sequencing

Request a detailed protocol

Single-cell suspensions were prepared from lung tissues of duplicate infected and uninfected control C57BL/6 mice at 4DPI, barcoded, and sequenced as individual mice. Following red blood cell lysis, cell viability was assessed by trypan blue staining; viable cells were sorted and loaded onto the 10x Genomics Chromium Controller for barcoding and library construction. Sequencing was performed on an Illumina NovaSeq platform. Data were processed using Cell Ranger and analyzed using the Seurat package in R. Quality control cutoffs of between 1000 and 10000 features and <20% mitochondrial DNA were applied. Data were normalized and scaled in Seurat v5, and groups were integrated via Harmony-based integration. Clusters were identified, determined, and visualized via Seurat library workflow (RunPCA, FindNeighbors, FindClusters, RunUMAP) and then annotated via hallmark markers determined by differential gene expression analysis across clusters using Wilcoxon rank-sum tests. This dataset is now deposited with the Gene Expression Omnibus (GSE324217).

Neutrophil isolation and bactericidal assays

Request a detailed protocol

BMDNs were isolated by density gradient centrifugation from adult C57BL/6 mice and cultured in DMEM supplemented with 10% FBS. For killing assays, BMDNs were incubated with B. pertussis at a multiplicity of infection of 10:1 for 2 or 24 hr. Supernatants were serially diluted and plated to quantify surviving CFU. Recombinant murine PGLYRP1 (R&D Systems) or BSA control was incubated with live B. pertussis or purified PGN fragments (TCT gifted by Prof. William Goldman or S. aureus PGN, Invivogen) to assess direct bactericidal activity.

RNAscope in situ hybridization

Request a detailed protocol

RNA in situ hybridization was performed on 5 µm formalin-fixed paraffin-embedded lung sections with the RNAscope detection kit to confirm RNA sequencing identification of PGLYRP1 transcripts (Advanced Cell Diagnostics). Hybridization, amplification, and detection steps were carried out according to the manufacturer’s protocol. Briefly, slides were deparaffinized in xylene and rehydrated using an ethanol series. Sections were treated with hydrogen peroxide for 10 min at room temperature, followed by treatment with target retrieval reagent (Advanced Cell Diagnostics). Sections were hybridized with target-specific probes (Advanced Cell Diagnostics) followed by amplification and fluorescent detection (Opal 570 and Opal 520). Slides were counterstained with hematoxylin and eosin and imaged using an ECHO Revolve 4 and analyzed with ImageJ.

Reporter cell assays for NOD1, NOD2, and TREM-1 activation

Request a detailed protocol

HEK293-derived reporter cells expressing murine or human NOD1 or NOD2 with an NF-κB-dependent SEAP reporter (InvivoGen) and TREM-1-DAP12 reporter cells in the Jurkat T cell background with an NFAT-dependent luciferase reporter (Eurofins DiscoverX) were stimulated with B. pertussis TCT (1 ng/ml), conditioned media (1:20 ratio of conditioned bacterial media to cell culture media) or PGN fragments (TCT, S. aureus, or B. subtilis PGN (Invivogen)) in the presence or absence of recombinant human PGLYRP1 (R&D Systems) at a concentration of 5 µg per well for 24 hr. Supernatants were analyzed for reporter activity using colorimetric (NOD1 and NOD2 reporter cells; Promega) or luminescent (TREM-1 reporter cells; Invivogen) detection kits (Promega). TCT was quantitated from its peak area elution profile from the final purification step. For each preparation of TCT, this area was compared to the peak area of a purified standard that had been quantitated by amino acid analysis and generously provided by William Goldman (UNC). Cells were obtained directly from the manufacturer and verified as mycoplasma-free using Plasmotest.

Histopathology

Request a detailed protocol

Lungs were fixed in 10% formalin, embedded in paraffin, and sectioned at 5 µm thickness. Slides were stained with hematoxylin and eosin. Scores are given as the average score from three blinded investigators based on percentage and severity of bronchovascular bundle formation and infiltration of immune cells into alveolar spaces.

Statistics

Significant differences in pathology scores were calculated at each time point using a two-tailed Mann–Whitney U test. Each time point was analyzed independently. A p-value <0.05 was considered significant. A p-value <0.05 is denoted by *, <0.01 by **, and <0.005 by ***.

Data availability

Data discussed in this manuscript have been deposited to the NCBI Sequence Read Archive under PRJNA493118 and NIH Gene Expression Omnibus GSE324217.

The following data sets were generated

1. Rickert D
2. Skerry C
(2026) NCBI Gene Expression Omnibus
ID GSE324217. A dual role for PGLYRP1 in host defense and immune regulation during B. pertussis infection.

https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE324217
1. Rickert D
2. Skerry C
(2026) NCBI Sequence Read Archive
ID PRJNA493118. Mus musculus musculus breed:C57BL/6 (eastern European house mouse).

https://www.ncbi.nlm.nih.gov/sra/PRJNA493118

References

(2020) Age-dependent effects of Type I and Type III IFNs in the pathogenesis of Bordetella pertussis infection and disease
Journal of Immunology 204:2192–2202.

https://doi.org/10.4049/jimmunol.1900912
- PubMed
- Google Scholar
(2023) DNA-dependent interferon induction and lung inflammation in Bordetella pertussis infection
Journal of Interferon & Cytokine Research 43:478–486.

https://doi.org/10.1089/jir.2023.0066
- PubMed
- Google Scholar
(2013) TREM-1: Intracellular signaling pathways and interaction with pattern recognition receptors
Journal of Leukocyte Biology 93:209–215.

https://doi.org/10.1189/jlb.0312145
- PubMed
- Google Scholar
1. Birkebaek NH
2. Kristiansen M
3. Seefeldt T
4. Degn J
5. Moller A
6. Heron I
7. Andersen PL
8. Moller JK
9. Ostergård L
(1999) Bordetella pertussis and chronic cough in adults
Clinical Infectious Diseases 29:1239–1242.

https://doi.org/10.1086/313448
- PubMed
- Google Scholar
(2000) Cutting edge: Inflammatory responses can be triggered by TREM-1, a novel receptor expressed on neutrophils and monocytes
Journal of Immunology 164:4991–4995.

https://doi.org/10.4049/jimmunol.164.10.4991
- PubMed
- Google Scholar
(2001) TREM-1 amplifies inflammation and is a crucial mediator of septic shock
Nature 410:1103–1107.

https://doi.org/10.1038/35074114
- PubMed
- Google Scholar
1. Chamaillard M
2. Hashimoto M
3. Horie Y
4. Masumoto J
5. Qiu S
6. Saab L
7. Ogura Y
8. Kawasaki A
9. Fukase K
10. Kusumoto S
11. Valvano MA
12. Foster SJ
13. Mak TW
14. Nuñez G
15. Inohara N
(2003) An essential role for NOD1 in host recognition of bacterial peptidoglycan containing diaminopimelic acid
Nature Immunology 4:702–707.

https://doi.org/10.1038/ni945
- PubMed
- Google Scholar
1. Chen S
2. Putnik R
3. Li X
4. Diwaker A
5. Vasconcelos M
6. Liu S
7. Gondi S
8. Zhou J
9. Guo L
10. Xu L
11. Temme S
12. Bersch K
13. Hyland S
14. Yin J
15. Burstein E
16. Bahnson BJ
17. Gildersleeve JC
18. Grimes CL
19. Reinecker HC
(2025) PGLYRP1-mediated intracellular peptidoglycan detection promotes intestinal mucosal protection
Nature Communications 16:1864.

https://doi.org/10.1038/s41467-025-57126-9
- Google Scholar
(1989) Primary structure of the peptidoglycan-derived tracheal cytotoxin of Bordetella pertussis
Biochemistry 28:1744–1749.

https://doi.org/10.1021/bi00430a048
- PubMed
- Google Scholar
1. Dey RJ
2. Dey B
3. Harriff M
4. Canfield ET
5. Lewinsohn DM
6. Bishai WR
(2021) Augmentation of the riboflavin-biosynthetic pathway enhances Mucosa-Associated Invariant T (MAIT) cell activation and diminishes Mycobacterium tuberculosis virulence
mBio 13:e0386521.

https://doi.org/10.1128/mbio.03865-21
- PubMed
- Google Scholar
1. Dziarski R
2. Platt KA
3. Gelius E
4. Steiner H
5. Gupta D
(2003) Defect in neutrophil killing and increased susceptibility to infection with nonpathogenic gram-positive bacteria in peptidoglycan recognition protein-S (PGRP-S)-deficient mice
Blood 102:689–697.

https://doi.org/10.1182/blood-2002-12-3853
- PubMed
- Google Scholar
1. Dziarski R
2. Gupta D
(2006a) The peptidoglycan recognition proteins (PGRPs)
Genome Biology 7:232.

https://doi.org/10.1186/gb-2006-7-8-232
- PubMed
- Google Scholar
1. Dziarski R
2. Gupta D
(2006b) Mammalian PGRPs: Novel antibacterial proteins
Cellular Microbiology 8:1059–1069.

https://doi.org/10.1111/j.1462-5822.2006.00726.x
- PubMed
- Google Scholar
1. Flak TA
2. Goldman WE
(1999) Signalling and cellular specificity of airway nitric oxide production in pertussis
Cellular Microbiology 1:51–60.

https://doi.org/10.1046/j.1462-5822.1999.00004.x
- PubMed
- Google Scholar
1. Fullen AR
2. Gutierrez-Ferman JL
3. Yount KS
4. Love CF
5. Choi HG
6. Vargas MA
7. Raju D
8. Corps KN
9. Howell PL
10. Dubey P
11. Deora R
(2022) Bps polysaccharide of Bordetella pertussis resists antimicrobial peptides by functioning as a dual surface shield and decoy and converts Escherichia coli into a respiratory pathogen
PLOS Pathogens 18:e1010764.

https://doi.org/10.1371/journal.ppat.1010764
- PubMed
- Google Scholar
(2021) Triggering receptor expressed on myeloid cells-1 (TREM-1) contributes to Bordetella pertussis inflammatory pathology
Infection and Immunity 89:e0012621.

https://doi.org/10.1128/IAI.00126-21
- PubMed
- Google Scholar
(1982) Detection, isolation, and analysis of a released Bordetella pertussis product toxic to cultured tracheal cells
Infection and Immunity 36:782–794.

https://doi.org/10.1128/iai.36.2.782-794.1982
- PubMed
- Google Scholar
(1983) Respiratory morbidity and lung function after whooping-cough
Lancet 2:1104–1108.

https://doi.org/10.1016/s0140-6736(83)90626-8
- PubMed
- Google Scholar
1. Jutras BL
2. Lochhead RB
3. Kloos ZA
4. Biboy J
5. Strle K
6. Booth CJ
7. Govers SK
8. Gray J
9. Schumann P
10. Vollmer W
11. Bockenstedt LK
12. Steere AC
13. Jacobs-Wagner C
(2019) Borrelia burgdorferi peptidoglycan is a persistent antigen in patients with Lyme arthritis
PNAS 116:13498–13507.

https://doi.org/10.1073/pnas.1904170116
- PubMed
- Google Scholar
1. Kashyap DR
2. Wang M
3. Liu LH
4. Boons GJ
5. Gupta D
6. Dziarski R
(2011) Peptidoglycan recognition proteins kill bacteria by activating protein-sensing two-component systems
Nature Medicine 17:676–683.

https://doi.org/10.1038/nm.2357
- PubMed
- Google Scholar
1. Kessie DK
2. Lodes N
3. Oberwinkler H
4. Goldman WE
5. Walles T
6. Steinke M
7. Gross R
(2020) Activity of tracheal cytotoxin of Bordetella pertussis in a human tracheobronchial 3D tissue model
Frontiers in Cellular and Infection Microbiology 10:614994.

https://doi.org/10.3389/fcimb.2020.614994
- PubMed
- Google Scholar
(2025a) Myeloid cell-specific type I interferon signaling mediates age-dependent inflammation and protection in Bordetella pertussis infection
Infection and Immunity 93:e0030625.

https://doi.org/10.1128/iai.00306-25
- PubMed
- Google Scholar
(2025b) Interferon lambda signaling in neutrophils enhances the pathogenesis of Bordetella pertussis infection
Journal of Leukocyte Biology 117:qiae202.

https://doi.org/10.1093/jleuko/qiae202
- PubMed
- Google Scholar
1. Liu C
2. Xu Z
3. Gupta D
4. Dziarski R
(2001) Peptidoglycan recognition proteins: A novel family of four human innate immunity pattern recognition molecules
The Journal of Biological Chemistry 276:34686–34694.

https://doi.org/10.1074/jbc.M105566200
- PubMed
- Google Scholar
1. Lu X
2. Wang M
3. Qi J
4. Wang H
5. Li X
6. Gupta D
7. Dziarski R
(2006) Peptidoglycan recognition proteins are a new class of human bactericidal proteins
The Journal of Biological Chemistry 281:5895–5907.

https://doi.org/10.1074/jbc.M511631200
- PubMed
- Google Scholar
Thesis
1. Lyon RS
(2001)
Tracheal cytotoxin production by the Bordetellae

Washington University, St Louis.
- Google Scholar
1. Magalhaes JG
2. Philpott DJ
3. Nahori MA
4. Jéhanno M
5. Fritz J
6. Le Bourhis L
7. Viala J
8. Hugot JP
9. Giovannini M
10. Bertin J
11. Lepoivre M
12. Mengin-Lecreulx D
13. Sansonetti PJ
14. Girardin SE
(2005) Murine Nod1 but not its human orthologue mediates innate immune detection of tracheal cytotoxin
EMBO Reports 6:1201–1207.

https://doi.org/10.1038/sj.embor.7400552
- PubMed
- Google Scholar
(1984) Ability of monomeric peptidoglycan fragments from Neisseria gonorrhoeae to damage human fallopian-tube mucosa
The Journal of Infectious Diseases 149:378–386.

https://doi.org/10.1093/infdis/149.3.378
- PubMed
- Google Scholar
1. Paddock CD
2. Sanden GN
3. Cherry JD
4. Gal AA
5. Langston C
6. Tatti KM
7. Wu KH
8. Goldsmith CS
9. Greer PW
10. Montague JL
11. Eliason MT
12. Holman RC
13. Guarner J
14. Shieh WJ
15. Zaki SR
(2008) Pathology and pathogenesis of fatal Bordetella pertussis infection in infants
Clinical Infectious Diseases 47:328–338.

https://doi.org/10.1086/589753
- PubMed
- Google Scholar
1. Parise G
2. Mishra M
3. Itoh Y
4. Romeo T
5. Deora R
(2007) Role of a putative polysaccharide locus in Bordetella biofilm development
Journal of Bacteriology 189:750–760.

https://doi.org/10.1128/JB.00953-06
- PubMed
- Google Scholar
(2012) Siglecs as sensors of self in innate and adaptive immune responses
Annals of the New York Academy of Sciences 1253:37–48.

https://doi.org/10.1111/j.1749-6632.2011.06362.x
- PubMed
- Google Scholar
1. Read CB
2. Kuijper JL
3. Hjorth SA
4. Heipel MD
5. Tang X
6. Fleetwood AJ
7. Dantzler JL
8. Grell SN
9. Kastrup J
10. Wang C
11. Brandt CS
12. Hansen AJ
13. Wagtmann NR
14. Xu W
15. Stennicke VW
(2015) Cutting edge: Identification of neutrophil PGLYRP1 as a ligand for TREM-1
Journal of Immunology 194:1417–1421.

https://doi.org/10.4049/jimmunol.1402303
- PubMed
- Google Scholar
Preprint
1. Rickert DM
2. Hill E
3. Himmelberger R
4. Rajbanshi N
5. Cardozo S
6. de Castro TB
7. Bharadwaj R
8. Scott AJ
9. Silverman N
10. Goldman WE
11. Carbonetti NH
12. Scanlon K
13. Skerry C
(2025) B. Pertussis tracheal cytotoxin biases NOD signaling to suppress IL-1 mediated inflammation and evade adaptive immunity
bioRxiv.

https://doi.org/10.1101/2025.08.08.669383
- Google Scholar
(1987) Major fragment of soluble peptidoglycan released from growing Bordetella pertussis is tracheal cytotoxin
Infection and Immunity 55:2117–2120.

https://doi.org/10.1128/iai.55.9.2117-2120.1987
- PubMed
- Google Scholar
1. Sanchez M
2. Kolar SL
3. Müller S
4. Reyes CN
5. Wolf AJ
6. Ogawa C
7. Singhania R
8. De Carvalho DD
9. Arditi M
10. Underhill DM
11. Martins GA
12. Liu GY
(2017) O-Acetylation of peptidoglycan limits helper T cell priming and permits Staphylococcus aureus reinfection
Cell Host & Microbe 22:543–551.

https://doi.org/10.1016/j.chom.2017.08.008
- PubMed
- Google Scholar
1. Schleifer KH
2. Kandler O
(1972) Peptidoglycan types of bacterial cell walls and their taxonomic implications
Bacteriological Reviews 36:407–477.

https://doi.org/10.1128/br.36.4.407-477.1972
- PubMed
- Google Scholar
1. Schnell A
2. Huang L
3. Regan BML
4. Singh V
5. Vonficht D
6. Bollhagen A
7. Wang M
8. Hou Y
9. Bod L
10. Sobel RA
11. Chihara N
12. Madi A
13. Anderson AC
14. Regev A
15. Kuchroo VK
(2023) Targeting PGLYRP1 promotes antitumor immunity while inhibiting autoimmune neuroinflammation
Nature Immunology 24:1908–1920.

https://doi.org/10.1038/s41590-023-01645-4
- PubMed
- Google Scholar
1. Sharif S
2. Singh M
3. Kim SJ
4. Schaefer J
(2009) Staphylococcus aureus peptidoglycan tertiary structure from carbon-13 spin diffusion
Journal of the American Chemical Society 131:7023–7030.

https://doi.org/10.1021/ja808971c
- PubMed
- Google Scholar
1. Sigalov AB
(2014) A novel ligand-independent peptide inhibitor of TREM-1 suppresses tumor growth in human lung cancer xenografts and prolongs survival of mice with lipopolysaccharide-induced septic shock
International Immunopharmacology 21:208–219.

https://doi.org/10.1016/j.intimp.2014.05.001
- PubMed
- Google Scholar
1. Skerry C
2. Pinn ML
3. Bruiners N
4. Pine R
5. Gennaro ML
6. Karakousis PC
(2014) Simvastatin increases the in vivo activity of the first-line tuberculosis regimen
The Journal of Antimicrobial Chemotherapy 69:2453–2457.

https://doi.org/10.1093/jac/dku166
- PubMed
- Google Scholar
(2013) Peptidoglycan transformations during Bacillus subtilis sporulation
Molecular Microbiology 88:673–686.

https://doi.org/10.1111/mmi.12201
- PubMed
- Google Scholar
1. Torrens G
2. Cava F
(2024) Mechanisms conferring bacterial cell wall variability and adaptivity
Biochemical Society Transactions 52:1981–1993.

https://doi.org/10.1042/BST20230027
- PubMed
- Google Scholar
1. Veglia F
2. Hashimoto A
3. Dweep H
4. Sanseviero E
5. De Leo A
6. Tcyganov E
7. Kossenkov A
8. Mulligan C
9. Nam B
10. Masters G
11. Patel J
12. Bhargava V
13. Wilkinson P
14. Smirnov D
15. Sepulveda MA
16. Singhal S
17. Eruslanov EB
18. Cristescu R
19. Loboda A
20. Nefedova Y
21. Gabrilovich DI
(2021) Analysis of classical neutrophils and polymorphonuclear myeloid-derived suppressor cells in cancer patients and tumor-bearing mice
The Journal of Experimental Medicine 218:e20201803.

https://doi.org/10.1084/jem.20201803
- PubMed
- Google Scholar
1. Wang S
2. Zhang S
3. Liu J
(2025) Resurgence of pertussis: Epidemiological trends, contributing factors, challenges, and recommendations for vaccination and surveillance
Human Vaccines & Immunotherapeutics 21:2513729.

https://doi.org/10.1080/21645515.2025.2513729
- PubMed
- Google Scholar
1. Wolf AJ
2. Underhill DM
(2018) Peptidoglycan recognition by the innate immune system
Nature Reviews. Immunology 18:243–254.

https://doi.org/10.1038/nri.2017.136
- PubMed
- Google Scholar
1. Xie X
2. Shi Q
3. Wu P
4. Zhang X
5. Kambara H
6. Su J
7. Yu H
8. Park SY
9. Guo R
10. Ren Q
11. Zhang S
12. Xu Y
13. Silberstein LE
14. Cheng T
15. Ma F
16. Li C
17. Luo HR
(2020) Single-cell transcriptome profiling reveals neutrophil heterogeneity in homeostasis and infection
Nature Immunology 21:1119–1133.

https://doi.org/10.1038/s41590-020-0736-z
- PubMed
- Google Scholar

Article and author information

Author details

David M Rickert

Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, United States

Contribution
Conceptualization, Formal analysis, Investigation, Methodology, Writing – original draft, Writing – review and editing

Competing interests
No competing interests declared

"This ORCID iD identifies the author of this article:" 0009-0002-7229-0046
Sasha Cardozo

Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, United States

Contribution
Data curation, Investigation, Writing – original draft

Competing interests
No competing interests declared

"This ORCID iD identifies the author of this article:" 0000-0003-1608-7086
Nicholas Carbonetti

Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, United States

Contribution
Investigation, Methodology, Writing – original draft

Competing interests
No competing interests declared
William E Goldman

Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, United States

Contribution
Conceptualization, Methodology

Competing interests
No competing interests declared

"This ORCID iD identifies the author of this article:" 0000-0002-1551-6718
Karen M Scanlon

Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, United States

Contribution
Conceptualization, Data curation

Competing interests
No competing interests declared

"This ORCID iD identifies the author of this article:" 0000-0002-5300-8749
Ciaran Skerry

Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, United States

Contribution
Conceptualization, Resources, Data curation, Formal analysis, Supervision, Funding acquisition, Investigation, Methodology, Writing – original draft, Project administration, Writing – review and editing

For correspondence
cskerry@som.umaryland.edu

Competing interests
No competing interests declared

"This ORCID iD identifies the author of this article:" 0009-0009-3511-7472

Funding

National Institute of Allergy and Infectious Diseases (R01AI 167947)

Ciaran Skerry

National Institute of Allergy and Infectious Diseases (T32 AI095190)

David M Rickert

The funders had no role in study design, data collection, and interpretation, or the decision to submit the work for publication.

Acknowledgements

This work was supported by NIH award R01AI167947 and by the NIAID Institutional Training Grant T32 AI095190. Prof. Roman Dziarski (Indiana University School of Medicine) generously provided PGLYRP1 KO mice. Prof. Rajendar Deora (Ohio State College of Medicine) generously provided the BpsB deletion mutant strain of B. pertussis. This work was supported by funding from the National Institute for Allergy and Infectious Diseases grant R01 AI167947 (CS).

Ethics

This study was performed in accordance with the recommendations in the Guide for Care and Use of Laboratory Animals of the National Institutes of Health. All animals were handled according to the approved Institutional Animal Care and Use Committee (IACUC) (protocol 00000108) of the University of Maryland Baltimore.

Version history

Sent for peer review: September 26, 2025
Preprint posted: September 27, 2025
Reviewed Preprint version 1: January 2, 2026
Reviewed Preprint version 2: June 9, 2026
Version of Record published: August 11, 2026

Cite all versions

You can cite all versions using the DOI https://doi.org/10.7554/eLife.108947. This DOI represents all versions, and will always resolve to the latest one.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.