The potency of hematopoietic stem cell reprogramming for changing immune tone.

Innate immune memory endows innate immune cells with antigen independent heightened responsiveness to subsequent challenges. The durability of this response can be mediated by inflammation induced epigenetic and metabolic reprogramming in hematopoietic stem and progenitor cells (HSPCs) that are maintained through differentiation to mature immune progeny. Understanding the mechanisms and extent of trained immunity induction by pathogens and vaccines, such as BCG, in HSPC remains a critical area of exploration with important implications for health and disease. Here we review these concepts and present new analysis to highlight how inflammatory reprogramming of HSPC can potently alter immune tone, including to enhance specific anti-tumor responses. New findings in the field pave the way for novel HSPC targeting therapeutic strategies in cancer and other contexts of immune modulation. Future studies are expected to unravel diverse and extensive effects of infections, vaccines, microbiota, and sterile inflammation on hematopoietic progenitor cells and begin to illuminate the broad spectrum of immunologic tuning that can be established through altering HSPC phenotypes. The purpose of this review is to draw attention to emerging and speculative topics in this field where we posit that focused study of HSPC in the framework of trained immunity holds significant promise.

A bacterial effector protein promotes nuclear translocation of Stat3 to induce IL-10.

The multifunctional Yersinia effector YopM inhibits effector triggered immunity and increases production of the anti-inflammatory cytokine Interleukin-10 (IL-10) to suppress the host immune response. Previously it was shown that YopM induces IL-10 gene expression by elevating phosphorylation of the serine-threonine kinase RSK1 in the nucleus of human macrophages. Using transcriptomics, we found that YopM strongly affects expression of genes belonging to the JAK-STAT signaling pathway. Further analysis revealed that YopM mediates nuclear translocation of the transcription factor Stat3 in Y. enterocolitica infected macrophages and that knockdown of Stat3 inhibited YopM-induced IL-10 gene expression. YopM-induced Stat3 translocation did not depend on autocrine IL-10, activation of RSK1 or tyrosine phosphorylation of Stat3. Thus, besides activation of RSK1, stimulation of nuclear translocation of Stat3 is another mechanism by which YopM increases IL-10 gene expression in macrophages.

Rapid development of cefiderocol resistance in a carbapenem-resistant Pseudomonas aeruginosa isolate associated with mutations in the pyoverdine biosynthesis pathway.

Here we report the in vivo development of cefiderocol resistance within 11 days after therapy initiation in a critically ill patient with bloodstream infection, infection of peri-anal fistula, and pneumonia caused by a VIM-2 harbouring, carbapenem-resistant Pseudomonas aeruginosa. Compared to a cefiderocol-naïve P. aeruginosa blood culture isolate, agar diffusion susceptibility testing found a reduced cefiderocol inhibition zone diameter in a P. aeruginosa recovered from peri-anal abscess tissue cultures after initiation of cefiderocol therapy. Subsequent whole-genome sequencing suggested that both isolates were of clonal origin. Comparison of genomes found an accumulation of missense mutations within pvdP, pvdE, pvdJ, and pvdD (i.e. genes associated with biosynthesis of pyoverdine), the main siderophore produced by P. aeruginosa. Quantification of pyoverdine production under iron-depleted conditions showed a significantly (P = 0.0003) higher pyoverdine production by the cefiderocol-resistant isolate. While pyoverdine quantity alone appears not to be decisive for cefiderocol resistance, the reported case highlights the potentially rapid emergence of cefiderocol resistance in P. aeruginosa and points towards a potential involvement of iron up-take systems in this process.

Detection of human herpesvirus 6 DNA and chromosomal integration after allogeneic hematopoietic stem cell transplantation: A retrospective single center analysis.

INTRODUCTION: Human herpesvirus 6 (HHV-6) can reactivate after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and may be associated with significant morbidity and mortality. METHODS: The epidemiology of HHV-6 infections and their impact on outcome after allo-HSCT were retrospectively analyzed in 689 adult allo-HSCT recipients (January 2015-December 2018). Chromosomal integration of HHV-6 (ciHHV-6) in the donor was retrospectively investigated to critically evaluate antiviral treatment strategies. RESULTS: HHV-6 DNA in any specimen was found in 89 patients. HHV-6 infections (encephalitis (one), gastroenteritis (44), dermatitis (two), hepatitis (one), or pneumonitis (five)) were diagnosed in 53/689 patients (7.7%). Elevated levels of HHV-6 DNA were found in 38 patients (5.5%). ciHHV-6, analyzed in patients with HHV-6 viral loads ≥104  copies/ml, was identified in four patients (10/38 patients; 10.5%). Two of those displayed copy numbers of HHV-6 ranging from ≥2 × 105 to 2.5 × 106  copies/ml (HHV-6A). Here, ciHHV-6 was integrated into donor and not into the patients' cells. In this series of allo-HSCT recipients, 10.5% of patients with blood viral loads of HHV-6 showed ciHHV-6. CONCLUSION: Screening of the donor for ciHHV-6 before initiation of antiviral therapy is recommended.

Yersinia remodels epigenetic histone modifications in human macrophages.

Various pathogens systematically reprogram gene expression in macrophages, but the underlying mechanisms are largely unknown. We investigated whether the enteropathogen Yersinia enterocolitica alters chromatin states to reprogram gene expression in primary human macrophages. Genome-wide chromatin immunoprecipitation (ChIP) seq analyses showed that pathogen-associated molecular patterns (PAMPs) induced up- or down-regulation of histone modifications (HMod) at approximately 14500 loci in promoters and enhancers. Effectors of Y. enterocolitica reorganized about half of these dynamic HMod, with the effector YopP being responsible for about half of these modulatory activities. The reorganized HMod were associated with genes involved in immune response and metabolism. Remarkably, the altered HMod also associated with 61% of all 534 known Rho GTPase pathway genes, revealing a new level in Rho GTPase regulation and a new aspect of bacterial pathogenicity. Changes in HMod were associated to varying degrees with corresponding gene expression, e. g. depending on chromatin localization and cooperation of the HMod. In summary, infection with Y. enterocolitica remodels HMod in human macrophages to modulate key gene expression programs of the innate immune response.

Clinical evaluation of a laboratory-developed quantitative BK virus-PCR assay using the cobas® omni Utility Channel.

BACKGROUND: In immunocompromised patients, BK Virus (BKV) reactivation may cause serious disease with high morbidity. Particularly for patient management after solid organ transplantation, monitoring of viral load in different clinical specimens is crucial to ensure early diagnosis and response to reactivation. In this study, we evaluated the clinical performance of a custom designed primer /probe set for detection of BKV on the cobas® 6800, a high-throughput platform, employing the open channel of the system for integration of a lab-developed test (LDT). MATERIALS/METHODS: A primer/probe set was optimized for the use on a high-throughput platform. Clinical performance was assessed in EDTA-plasma, serum and urine samples. Limit-of-detection (LOD) was determined by using a dilution series of BKV WHO standard. A CE-labeled PCR test (Altona Diagnostics) was used as a comparison to the assay. RESULTS: The LOD for the LDT BKV assay was 6.7 IU/mL. Inter-and intra-run variability (at 5 x LOD) was low (<1.5 Ct in all specimens). All quality control panel specimens (Instand Germany n = 19) were correctly identified. Of 290 clinical samples tested, results were concordant for 280 samples. Sensitivity and specificity of the assay were 96 % and 98 % respectively. The quantitative analysis revealed a strong correlation (linear regression) between the CE-labelled comparator assay and the new BKV LDT assay with r2 = 0.96 for n = 123 urine samples and r2 = 0.98 for n = 167 plasma/serum samples. CONCLUSION: Compared to a CE-IVD assay, the adapted LDT showed good analytical and clinical sensitivity and specificity for the detection and quantification of BKV in different clinical specimens. It represents a convenient solution to automate the LDT workflow with low hands-on time and thus facilitates high-throughput screening for BKV reactivation in immunocompromised patients.

Major central nervous system complications after allogeneic stem cell transplantation: A large retrospective study on 888 consecutive adult patients.

OBJECTIVES: Major complications affecting the central nervous system (CNS) present a challenge after allogeneic stem cell transplantation (allo-SCT). METHODS: Incidence, risk factors, and outcome were retrospectively analyzed in 888 patients in a monocentric study. RESULTS: Cumulative incidence (CI) of major CNS complications at 1 year was 14.8% (95%CI 12.3%-17.2%). Median follow-up is 11 months. CNS complications were documented in 132 patients: in 36 cases, classified metabolic; 26, drug-related neurotoxicity (14 attributed to cyclosporine A, 4 to antilymphocyte globulin); 11, cerebrovascular (ischemic n = 8, bleeding n = 3); 9, infections; 9, psychiatric; and 9, malignant. The cause of CNS symptoms remained unclear for 37 patients (28%). Multivariate analysis demonstrated an association of CNS complication with patient age (P < .001). The estimated OS of patients with any CNS complication was significantly lower than in patients without neurological complications (P < .001), and the CI of non-relapse mortality (NRM) was higher for patients with CNS complication (P < .001). A significant negative impact on survival can only be demonstrated for metabolic CNS complications and CNS infections (NRM, P < .0001 and P = .0003, respectively), and relapse (P < .0001). CONCLUSION: CNS complications after allo-SCT are frequent events with a major contribution to morbidity and mortality. In particular, the situations of unclear neurological complications need to be clarified by intensive research.

Detection of SARS-CoV-2 in Human Retinal Biopsies of Deceased COVID-19 Patients.

PURPOSE: To report the presence of viral ribonucleic acid (RNA) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in human retina in deceased patients with confirmed novel coronavirus disease 2019 (COVID-19). PATIENTS AND METHODS: Fourteen eyes of 14 deceased patients with confirmed COVID-19 disease were enucleated during autopsy. A sample of human retina was secured and fixed in RNAlater™. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to detect three different viral RNA sequences (RdRp-gene, E-gene and Orf1 gene) of SARS-CoV-2. RESULTS: In three out of 14 eyes SARS-CoV-2 viral RNA was detected in the retina of deceased COVID-19 patients. As analysis for three different sequences (RdRp-gene, E-gene and Orf1 gene) revealed positive results in RT-PCR, the existence of SARS-CoV-2 viral RNA in human retina is proven according to the standards of the World-Health-Organization. CONCLUSION: Viral RNA of SARS-CoV-2 is detectable in the retina of COVID-19 patients.

Monocenter study on epidemiology, outcomes, and risk factors of infections in recipients of 166 allogeneic stem cell transplantations during 1 year.

OBJECTIVES: During allogeneic hematopoietic stem cell transplantation (allo-SCT), infections significantly contribute to morbidity and mortality. A monocentric prospective analysis was performed to assess epidemiology, risk factors, and outcomes of infections during the peri-transplant period. METHODS: Data were recorded prospectively using a predefined questionnaire. RESULTS: In 2015, 163 consecutive patients, 37.4% female, median age 59 (range 18-79) years received 166 allo-SCT. Median duration of leukopenia <109 /L was 14.5 days (range 4-43 days). Fever of unknown origin (FUO) occurred in 118/166 patients (71.1%). Severe sepsis developed in 95, and septic shock developed in 26 patients. Intensive diagnostic workup helped to identify causative microorganisms only in a small number of infectious courses. All but 13 patients needed antibiotic therapy, each according to the standard operating procedures of the department. Cumulative incidence of death by infection after 1 year was 16.6% (95% CI: 11.3-22.7). The only risk factor for FUO in neutropenia was duration of neutropenia

Immunosuppressive Yersinia Effector YopM Binds DEAD Box Helicase DDX3 to Control Ribosomal S6 Kinase in the Nucleus of Host Cells.

Yersinia outer protein M (YopM) is a crucial immunosuppressive effector of the plaque agent Yersinia pestis and other pathogenic Yersinia species. YopM enters the nucleus of host cells but neither the mechanisms governing its nucleocytoplasmic shuttling nor its intranuclear activities are known. Here we identify the DEAD-box helicase 3 (DDX3) as a novel interaction partner of Y. enterocolitica YopM and present the three-dimensional structure of a YopM:DDX3 complex. Knockdown of DDX3 or inhibition of the exportin chromosomal maintenance 1 (CRM1) increased the nuclear level of YopM suggesting that YopM exploits DDX3 to exit the nucleus via the CRM1 export pathway. Increased nuclear YopM levels caused enhanced phosphorylation of Ribosomal S6 Kinase 1 (RSK1) in the nucleus. In Y. enterocolitica infected primary human macrophages YopM increased the level of Interleukin-10 (IL-10) mRNA and this effect required interaction of YopM with RSK and was enhanced by blocking YopM's nuclear export. We propose that the DDX3/CRM1 mediated nucleocytoplasmic shuttling of YopM determines the extent of phosphorylation of RSK in the nucleus to control transcription of immunosuppressive cytokines.