bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dysregulated neutrophils failed to control bacteria in an intradermal murine model of Orientia tsutsugamushi infection
Authors: Liang Yuejin , Card Galen , Fisher James Robert , Sun Jiaren , Walker David Hughes , Soong Lynn 
Published: February 2023

Abstract
Scrub typhus is a vector-borne febrile illness caused by the obligately intracellular bacterium, Orientia tsutsugamushi that are transmitted by the larvae of trombiculid mites. Approximately 1 million people are infected every year in an Asia-Pacific region. Patients without appropriate treatment may develop severe damage in various organs and lead to death. However, the host protective immunity against this infection is still not understood. Using an intradermal infection mouse model and CFSE-labeled bacteria, we found that neutrophils and monocytes were the majority of infected cells in the skin infection site at 4 and 8 hours post-infection (hpi). However, very few of them can migrate to local lymph node at 24 hpi. Instead, the infected langerhans cells were observed in local lymph nodes, but not in other organs including lung, bone marrow and spleen, as well as blood. To investigate the role of neutrophils in Orientia infection, we treated the infected mice with Anti-Ly6G antibody daily from −1 to 3 dpi. However, the bodyweight loss was comparable between control and neutrophil-depleted mice until 14 dpi. No significant difference of bacterial loads were found in organs including lung and brain. Our in vitro study showed that Orientia infection induced an immunosuppressive neutrophil phenotype and increased neutrophil death. Collectively, our data suggest that infected-neutrophil may undergo the functional dysregulation and not be able to control bacteria efficiently, while antigen-presenting cells may contribute to bacterial dissemination from skin to systemic infection in scrub typhus.
The corruption of innate immunity by Streptococcus anginosus group
Authors: Budziaszek Joanna , Pilarczyk-Zurek Magdalena , Sitkiewicz Izabela , Koziel Joanna 
Published: February 2023

Abstract
Streptococcus anginosus group (SAG), formerly known as Streptococcus milleri, consists of three distinct streptococcal species: Streptococcus anginosus, Streptococcus constellatus, and Streptococcus intermedius. SAG was considered as commensal bacteria of colon, oral cavity and vagina. Recent observation reported those bacteria as potent pathogens found in brain or liver abscess. Mechanism of SAG pathogenesis is still unknown, despite the strong set of clinical data. In our study, we determined the virulence of different SAG clinical isolates by examining the activity of hemolysins, DNAses and proteases. By using Dictyostelium discoideum as model for screening SAG virulence we selected a few high and low virulence strains out of 41 SAG isolates. Finally, we established and characterized the infection process of selected SAG strains by wax worm Galleria mellonella. Therefore, we aimed to characterize how those bacteria can interact with human innate immunity. In our experiments, using both human serum and whole blood we demonstrated, that these bacteria are resistant to killing by non-specific mechanisms of the immune response. Our studies have demonstrated that SAG have numerous virulence factors which differ among examined strains, and which allow bacteria to effectively colonize the human body and avoid elimination by the immune system.
Supported by National Science Centre, Poland 2018/29/B/NZ6/00624
Ineffective CD4 T cell polarization reveals a novel immune evasion target of Borrelia burgdorferi
Authors: Hammond Elizabeth M , Olsen Kimberly J. , Baumgarth Nicole 
Published: February 2023

Abstract
Appropriate polarization of effector CD4 T cells is essential for eliminating invading pathogens. Borrelia burgdorferi (Bb), the spirochetal agent of Lyme disease, evades immune clearance in infected mice and humans, establishing bacterial persistence. Here we aimed to explore the functionality of CD4 T cell as a possible target of Borrelia-induced immune evasion. In response to Bb infection, CD4 T cell polarization is considered to be Th1 dominant. However, when we stimulated CD4 T cells from Bb infected mice ex vivo with Bb surface Ag Arthritis Related Protein (Arp), we were unable to detect IFNγ, prompting us to re-examine the polarization state of lymph node and blood effector CD4 T cells throughout Bb infection by quantitative RT-PCR and flow cytometry. In contrast to prevailing views, we found only modest numbers of Th1-polarized activated CD4 T cells, as measured by expression of Tbet (Tbx21), and little to no induction of Ifng, including in Bb-specific CD4 T cells, identified by a novel I-Ab tetramer to Arp. This could not be explained by enhanced polarization to Th2 or Th17, as GATA-3 and RORγt expression were also largely absent. Nor could it be explained by polarization at a T cell effector site, the skin, as skin-resident T cells also showed no polarization even 4 months after infection. Furthermore, Th1 polarization was not necessary for controlling bacterial dissemination, as Bb infection of Tbet−/− mice showed significant reductions in Borrelia numbers in the tibiotarsal joints, which correlated with enhanced Th17 polarization as measured by RORγt expression. Together, our data demonstrate a failure of CD4 T cell polarization after Bb infection, indicating effector CD4 T cell polarization as a novel target of Bb immune evasion.
Dual transcriptomics to decipher the interferon-gamma independent responses to intestinal Cryptosporidium parvum infection.
Authors: GALLEGO-LOPEZ GINA MARCELA , Mendoza-Cavazos Carolina , Tibabuzo-Perdomo Andres Mauricio , OConnor Roberta , Knoll Laura Jean 
Published: February 2023

Abstract

Background:
This study characterizes the IFNγ independent host immune response against C. parvum to discover new targets to halt the disease. We used transcriptomic analysis of ileum and cecum intestinal sections of IFNγ-deleted mice that were treated with Soluble Toxoplasma gondii antigen (STAg) or PBS upon C. parvum infection.

Results:
STAg treatment reduced the oocyst shedding in C. parvum infected IFNγ deleted mice. To understand the IFNγ independent mechanism of STAg treatment against C. parvum, we performed 12 RNAseq comparisons between tissues, treatments, and infection status for Mus musculus and C. parvum. In the ileum, regardless of STAg or PBS treatment, more than 100 genes had a significant differential expression. Within STAg treated mice, a total of 129 genes for ileum and 78 for the cecum were classified as differentially expressed. We found in high abundance at least 28 genes related to IFN type I response in infected mice treated with STAg, among them, the oligoadenylate synthetase and Schlafen family gene members. For C. parvum transcriptome analysis, we observed in the ileum high abundance of the oocyst wall protein 1 and some mucins; while in the cecum, the RRM domain containing-protein and Tryptophan synthase beta chain transcripts were the more abundant transcripts.

Conclusions:
STAg treatment and C. parvum infection induced a type I interferon response that would be the cause of the reduction of oocyst shedding in STAG treated mice in an IFNy independent manner. While the depth of sequence did not allow us to observe in detail differential expression of genes in C. parvum, we found differences in parasite gene expression between the ileum and the cecum.
Effector function prior to establishment of the phagosomal pathogen niche is required for protective CD4+ T cell-mediated immunity against Leishmania
Authors: Hohman Leah S , Carneiro Matheus B.H. , Kratofil Rachel M. , Peters Nathan C. 
Published: February 2023

Abstract
Leishmania is an appealing model organism to study CD4+ T cell-mediated protective immunity against phagosomal pathogens which features localized primary and secondary infection sites with defined innate and adaptive responses. Upon secondary challenge of chronic L. major-infected C57Bl/6 mice, rapid CD4+T effector (TEFF) function via IFN-g-mediated activation of infected monocytes is associated with optimal immunity. However, the requirement for immediate effector function has yet to be demonstrated. Thus, we isolated time as a variable in the delivery of CD4+ TEFF effector function. We adoptively transferred (AT) chronic mouse-derived Ly6C+CD4+TEFFs into naïve recipients immediately (D0) or 4 days (D4) post-L. major challenge. In this time window Leishmania establishes an intracellular niche but does not proliferate. At day 21 post-challenge, D4 AT resulted in a total loss of the parasite control mediated by D0 transfer. To address whether parasite niche establishment modulated Th1 TEFF cell recruitment, we employed intravital imaging. Ag-sp T cells were present in the infected dermis at significantly lower numbers following D4 vs D0 transfer at 4 days post-T cell transfer, indicating a recruitment deficit. Rapid CD4+ Th1 effector function was required for circulating Th1 TEFF cells to capitalize on an early recruitment window associated with cxcl9 and cxcl10 expression and to prevent parasite niche establishment through altered cell recruitment, gene expression, and functional capacity of both innate and adaptive immune cells. Thus, acute TEFF cell availability is required to prevent host immunomodulation by Leishmania and is a key consideration for vaccination against phagosomal pathogens reliant on Th1 immunity.
Murine iNOS expression is essential for antifungal defenses in kidneys during disseminated cryptococcosis
Authors: Goughenour Kristie D , Zhao Jessica , Xu Jintao , Zhao Peter , Freeman Christine , Ganguly Anutosh , Olszewski Michal A 
Published: February 2023

Abstract
C. neoformans, is one of the top 3 fungal pathogens, causing disseminated disease with high mortality in the absence of immunoprotecion. The protective Th1 immune response leads to M1 macrophage polarization that upregulates inducible nitric oxide synthase (iNOS) generating microbicidal nitric oxide. The role of iNOS has been studied in pulmonary defenses to C. neoformans, while its role in disseminated infection remains unknown. Here we study the effect of iNOS deletion in mouse model of disseminated cryptococcosis. The iNOS−/− and the WT, C57BL/6 mice were infected with 106 CFU of C. neoformans 52D via retro-orbital intravenous inoculation. Infected iNOS−/− mice showed significantly reduced survival times, however, they lacked significant increases in fungal burdens in the common infection sites (lung, spleen and CNS). Interestingly, iNOS deletion resulted in nearly 100-fold increase of the kidney fungal burden. Kidney histology revealed extensive lesions resulting in almost complete destruction of the kidney cortical area in iNOS−/− mice and a corresponding loss of kidney function was observed. The lack of fungal control was not due to a failure to recruit immune cells as iNOS−/− mice had increased kidney leukocyte recruitment as well as increases in monocytes and neutrophils. BMDMs from iNOS−/− mice were able to kill Cryptococcus despite a slight M2 bias seen in vivo. iNOS−/− mice had no defect in T-cell polarization, but a slight decrease in surface markers of activation. We conclude that iNOS expression is particularly important for local fungal control in kidneys, although its deletion does not induce a shift in a systemic immunophenotype in the iNOS−/− mice; rather it serves as an effector molecule in a local kidney environment.
Heterogeneity of human intestinal intraepithelial T cells and their abnormal distribution in Crohn’s disease revealed at high resolution
Authors: Jaeger Natalia , Gamini Ramya , Cella Marina , Schettini Jorge L. , Bugatti Mattia , Zhao Shanrong , Rosadini Charles V. , Esaulova Ekaterina , Di Luccia Blanda , Kinnett Baylee , Vermi Willam , Artyomov Maxim N. , Wynn Thomas A , Xavier Ramnik J , Jelinsky Scott A. , Colonna Marco 
Published: February 2023

Abstract
Crohn’s disease (CD) is a chronic transmural inflammation of intestinal segments caused by dysregulated interaction between microbiome and gut immune system. Recurrent/relapsing CD and resistance to medical treatments result in complications requiring surgery. High-dimensional single-cell profiling approaches, such as scRNA-seq and mass cytometry, have been recently performed on intestinal specimens from patients with IBD and controls. However, most of these studies have analyzed whole mucosal biopsies or the lamina propria (LP) compartment, while few have addressed the intraepithelial lymphocytes (IEL) compartment. Here, we profiled T cells purified from the IEL and LP from terminal ileum resections of adult severe CD cases by single cell technologies. Our study defined a vast heterogeneity of T cell lineages in the IEL compartment. IEL included, among others, unique γδT cell subsets: NKp30+γδ T cells expressing RORγt, which produced IL-26 upon NKp30 engagement and a subset expressing PDGFD and CSF1, which may act on epithelial cells, IEL ILC1s, and macrophages, respectively. We have also observed long-lived memory TCF7+CD8+ T cells expressing DC chemoattractants and TFH subsets that may respond to distinct glutathione-conjugated lipids. CD IEL showed a significant increase of activated TH17, coupled with decreased CD8+ T cells, γδT cells, TFH, and Treg. Conversely, the LP showed increased CD8+ T cells and reduced CD4+ T cells with a relative increase of TH17 over Treg/TFH. Results provide an unbiased view of diversity of cell lineages and their functional states in the intestinal mucosa of controls and CD and identify an altered spatial distribution of T cell subsets between the IEL and the LP compartments.
Th17 immunity in the colon is controlled by two novel subsets of colon-specific mononuclear phagocytes
Authors: Huang Hsin-I , Jewell Mark , Hammer Gianna 
Published: February 2023

Abstract
Intestinal immunity is coordinated by specialized populations of mononuclear phagocytes. Although the cell subsets that constitute these populations are thought uniform throughout the intestine, small and large intestine have distinct anatomy, microbial composition, and immunological demands. Whether these distinctions demand organ-specific mononuclear phagocyte populations with dedicated organ-specific roles in immunity is unknown. Here we implement a new strategy to subset murine intestinal mononuclear phagocytes and identify two novel subsets which are colon-specific: a macrophage subset and a Th17-inducing dendritic cell (DC) subset. Colon-specific DCs and macrophages co-expressed CD24 and CD14, and surprisingly, both were dependent on the transcription factor IRF4. Novel IRF4-dependent CD14+CD24+ macrophages were markedly distinct from conventional macrophages and failed to express classical markers including CX3CR1, CD64 and CD88, and surprisingly expressed little IL-10, which was otherwise robustly expressed by all other intestinal macrophages. We further found that colon-specific CD14+CD24+ mononuclear phagocytes were essential for Th17 immunity in the colon, and provide definitive evidence that colon and small intestine have distinct antigen presenting cell requirements for Th17 immunity. Our findings reveal unappreciated organ-specific diversity of intestine-resident mononuclear phagocytes and organ-specific requirements for Th17 immunity.
Role of membrane mucin MUC17 in enterocytic barrier dysfunction associated with Crohn’s disease
Authors: Hernandez Elena Layunta , Jäverfelt Sofia , Pelaseyed Thaher 
Published: February 2023

Abstract
The aim of this study is to determine the role of membrane mucin MUC17 in Crohn’s disease (CD). The incidence of CD is increasing in Western countries. However, the epithelial defects that cause disease are unclear. CD is characterized by the disruption of intestinal barriers leading to detrimental bacterial challenges that result in chronic intestinal inflammation. Our recent studies show that enterocytes are coated by the membrane mucin MUC17 that creates a glycocalyx barrier that protects enterocytes against luminal bacteria (Layunta et al. 2021 Accepted). Despite recent advances in defining intestinal barriers, the role of MUC17 and glycocalyx in CD remains unknown.
In order to determine the role of MUC17 and glycocalyx in CD, we studied MUC17 and glycocalyx in ileum biopsies from CD (n=10) and non-CD (n=29) patients collected at Sahlgrenska University Hospital (Gothenburg). Our preliminary data show that MUC17 is downregulated in CD. In mouse ileum, MUC17 forms the glycocalyx barrier. Then, we asked if decrement of MUC17 levels could affect glycocalyx barrier function in CD. To address this question we designed a glycocalyx permeability assay in which we can measure the barrier function of MUC17 in glycocalyx by incubating patient biopsies with live GFP-expression E.coli (GFP+ E.Coli) and exogenous molecules such as fluorescent dextran. Our results showed that the glycocalyx in ileum of CD patients was more penetrable to GFP+ E.Coli and dextran compared to non-CD patients.
In summary, we propose that changes in MUC17 levels in the glycocalyx contributes to disruption of epithelial barrier function during CD. In addition, our novel glycocalyx permeability assay can be used to diagnose CD, and to evaluate treatment response in patients.
The contributions of NAIP/NLRC4 inflammasome activation in Tuft cells to host barrier defense and immunity.
Authors: Churchill Madeline J , Rauch Isabella 
Published: February 2023

Abstract
The small intestine is a key interface between host cells and potentially pathogenic agents, and as such has developed an array of host defense mechanisms to mitigate disease. The role of the epithelium in preventing systemic disease is becoming more appreciated, however the function of a subset of epithelial cells, the tuft cells, outside of parasitic sensing remains unclear.
Tuft cells express all of the NAIP/NLRC4 inflammasome components, suggesting that tuft cells have the ability to recognize bacterial pathogens. The NAIP/NLRC4 inflammasome is a multi-protein complex that forms when bacterial flagellin or type III secretion systems are detected in the cytosol of the host cell, leading to induction of pyroptosis. The exact contributions of NAIP/NLRC4 inflammasome activation in tuft cells towards host barrier defense and immunity are not known.
Data generated by my lab demonstrates that activation of the NAIP/NLRC4 inflammasome in tuft cells facilitates robust release of prostaglandin D2. The specific function of prostaglandin D2 release after NAIP/NLRC4 inflammasome activation in not currently known. However, it has been demonstrated that a subset of innate lymphoid cells (ILCs), ILC3s express the receptor for prostaglandin D2 and after activation, secrete IL-17 and IL-22, key cytokines that promote host defense against bacterial infections. We find that after specific activation of NAIP/NLRC4 inflammasome in tuft cells, IL-22 protein is increased in the small intestine. These findings support our hypothesis that tuft cells are not simply parasitic sensors but contribute to host barrier defense and immunity against bacterial pathogens.
Genetically Engineering ocular probiotics to manipulate ocular immunity and disease
Authors: Rigas Yannis Elias , Treat Benjamin R , Shane Jackie , Shanks Robert QM , Leger Anthony J St. 
Published: February 2023

Abstract
Recently, our lab discovered that the eye harbors a microbiome that includes Corynebacterium mastitidis, which induces the recruitment of gdT cells and the production of interleukin 17 (IL-17). This local immune response to C. mast can protect the eye from more serious infections, such as Candida albicans and Pseudomonas aeruginosa. Because of C. mast’s ability to stably colonize the ocular surface while inducing a beneficial immune response, we believe that this bacterium has the potential to act as a natural vehicle to deliver therapeutics locally at the ocular surface to alleviate ocular surface disease. Before this can become a reality, a reliable method to genetically modify C. mast is required. Therefore, we took the initial steps toward achieving this goal by genetically modifying C. mast to express genes of interest using transposon mutagenesis. This allowed us to create a mutant library with 1,000+ distinct candidates that expressed the fluorescent protein, mCherry, and kanamycin resistance. Using FACS, we were able to select the brightest C. mast mutants. The brightest four mutants were used to inoculate the eyes of C57BL/6 mice, so that we could: 1) track C. mast in vivo and 2) ensure that the promoters upstream of the genes of interest remain active when C. mast has colonized the ocular mucosa. We were able to image recombinant bacteria directly ex vivo and also confirmed that there was no decrease in bacterial fitness or the protective immune response elicited by C. mast. We concluded from these studies that C. mast can be engineered to express proteins of interest without a reduction in bacterial fitness or protective immunity.
Myeloid cells direct recruitment of colon intraepithelial lymphocytes to the epithelium, where they undergo functional maturation through MHC class II interactions
Authors: Lefferts Adam R , Mann Sarah , Van Der Kraak Lauren A , Canna Scott , Kuhn Kristine A 
Published: February 2023

Abstract
Intraepithelial lymphocytes (IELs) are T cells defined by direct contact with epithelial surfaces. While important for barrier integrity, the mechanisms of recruitment and functional maturation remain unknown. We identified sphingosine-1-phosphate (S1P) receptor 1 (S1PR1) as highly enriched in the IEL population compared to lamina propria T cells by microarray, suggesting importance in IEL recruitment. Blockade of S1P signaling using FTY720 significantly reduced recruitment of TCRab+ IELs, but not TCRgd+ IELs, following bacterial recolonization of antibiotic-treated mice, which is known to recruit IELs. Since mice lacking either Myd88 or an intact microbiome lack IELs, we hypothesized that either the epithelium or mononuclear phagocytes responded to microbial TLR ligands and produced S1P. Myeloid lineage cells were deemed the cells responsible for Myd88/microbe dependent IEL recruitment. In vitro, cecal contents stimulated LysM-Cre+ Myd88fl/fl bone-marrow derived macrophages do not upregulate sphk1, the enzyme responsible for most S1P production, and fail to induce S1P dependent T cell migration in a transwell assay. To understand how the IEL-IEC synapse affects IEL function, we utilized Villin-Cre+ MHCIIfl/fl mice. We observed altered maturation in both CD4+ and CD4− CD8− IEL compartments, associated with increased CD44− CD62L+ naive and CD44+ CD62L+ central memory IELs, and decreased CD44+ CD62L− effector, and CD44− CD62L− “revertant” IELs, suggesting a block in T cell maturation. Together these data show a coordinated, multi-system process by which IELs are first recruited to, and then functionally matured by the IECs. Further work will investigate how inflammation of the mucosal barrier alters these processes.