Gut Microbiota in Diagnosis, Therapy and Prognosis of Cholangiocarcinoma and Gallbladder Carcinoma-A Scoping Review.

Cancers of the biliary tract are more common in Asia than in Europe, but are highly lethal due to delayed diagnosis and aggressive tumor biology. Since the biliary tract is in direct contact with the gut via the enterohepatic circulation, this suggests a potential role of gut microbiota, but to date, the role of gut microbiota in biliary tract cancers has not been elucidated. This scoping review compiles recent data on the associations between the gut microbiota and diagnosis, progression and prognosis of biliary tract cancer patients. Systematic review of the literature yielded 154 results, of which 12 studies and one systematic review were eligible for evaluation. The analyses of microbiota diversity indices were inconsistent across the included studies. In-depth analyses revealed differences between gut microbiota of biliary tract cancer patients and healthy controls, but without a clear tendency towards particular species in the studies. Additionally, most of the studies showed methodological flaws, for example non-controlling of factors that affect gut microbiota. At the current stage, there is a lack of evidence to support a general utility of gut microbiota diagnostics in biliary tract cancers. Therefore, no recommendation can be made at this time to include gut microbiota analyses in the management of biliary tract cancer patients.

Longitudinal dynamics of gut bacteriome and mycobiome interactions pre- and post-visceral surgery in Crohn's disease.

Introduction: Alterations of the gut microbiome are involved in the pathogenesis of Crohn's disease (CD). The role of fungi in this context is unclear. This study aimed to determine postoperative changes in the bacterial and fungal gut communities of CD patients undergoing intestinal resection, and to evaluate interactions between the bacteriome and mycobiome and their impact on the patients' outcome. Methods: We report a subgroup analysis of a prospective cohort study, focusing on 10 CD patients whose fecal samples were collected for bacterial 16S rRNA and fungal ITS2 genes next-generation sequencing the day before surgery and on the 5th or 6th postoperative day. Results: No significant differences in bacterial and fungal diversity were observed between preoperative and postoperative stool samples. By in-depth analysis, significant postoperative abundance changes of bacteria and fungi and 17 interkingdom correlations were detected. Network analysis identified 13 microbial clusters in the perioperative gut communities, revealing symbiotic and competitive interactions. Relevant factors were gender, age, BMI, lifestyle habits (smoking, alcohol consumption) and surgical technique. Postoperative abundance changes and identified clusters were associated with clinical outcomes (length of hospital stay, complications) and levels of inflammatory markers. Conclusions: Our findings highlight the importance of dissecting the interactions of gut bacterial and fungal communities in CD patients and their potential influence on postoperative and disease outcomes.

Mitochondria supply sub-lethal signals for cytokine secretion and DNA-damage in H. pylori infection.

The bacterium Helicobacter pylori induces gastric inflammation and predisposes to cancer. H. pylori-infected epithelial cells secrete cytokines and chemokines and undergo DNA-damage. We show that the host cell's mitochondrial apoptosis system contributes to cytokine secretion and DNA-damage in the absence of cell death. H. pylori induced secretion of cytokines/chemokines from epithelial cells, dependent on the mitochondrial apoptosis machinery. A signalling step was identified in the release of mitochondrial Smac/DIABLO, which was required for alternative NF-κB-activation and contributed to chemokine secretion. The bacterial cag-pathogenicity island and bacterial muropeptide triggered mitochondrial host cell signals through the pattern recognition receptor NOD1. H. pylori-induced DNA-damage depended on mitochondrial apoptosis signals and the caspase-activated DNAse. In biopsies from H. pylori-positive patients, we observed a correlation of Smac-levels and inflammation. Non-apoptotic cells in these samples showed evidence of caspase-3-activation, correlating with phosphorylation of the DNA-damage response kinase ATM. Thus, H. pylori activates the mitochondrial apoptosis pathway to a sub-lethal level. During infection, Smac has a cytosolic, pro-inflammatory role in the absence of apoptosis. Further, DNA-damage through sub-lethal mitochondrial signals is likely to contribute to mutagenesis and cancer development.

Spontaneous activity of the mitochondrial apoptosis pathway drives chromosomal defects, the appearance of micronuclei and cancer metastasis through the Caspase-Activated DNAse.

Micronuclei are DNA-containing structures separate from the nucleus found in cancer cells. Micronuclei are recognized by the immune sensor axis cGAS/STING, driving cancer metastasis. The mitochondrial apoptosis apparatus can be experimentally triggered to a non-apoptotic level, and this can drive the appearance of micronuclei through the Caspase-activated DNAse (CAD). We tested whether spontaneously appearing micronuclei in cancer cells are linked to sub-lethal apoptotic signals. Inhibition of mitochondrial apoptosis or of CAD reduced the number of micronuclei in tumor cell lines as well as the number of chromosomal misalignments in tumor cells and intestinal organoids. Blockade of mitochondrial apoptosis or deletion of CAD reduced, while experimental activation CAD, STING-dependently, enhanced aggressive growth of tumor cells in vitro. Deletion of CAD from human cancer cells reduced metastasis in xenograft models. CAD-deficient cells displayed a substantially altered gene-expression profile, and a CAD-associated gene expression 'signature' strongly predicted survival in cancer patients. Thus, low-level activity in the mitochondrial apoptosis apparatus operates through CAD-dependent gene-induction and STING-activation and has substantial impact on metastasis in cancer.

Host Blood Gene Signatures Can Detect the Progression to Severe and Cerebral Malaria.

Malaria is a major international public health problem that affects millions of patients worldwide especially in sub-Saharan Africa. Although many tests have been developed to diagnose malaria infections, we still lack reliable diagnostic biomarkers for the identification of disease severity, especially in endemic areas where the diagnosis of cerebral malaria is very difficult and requires the exclusion of all other possible causes. Previous host and pathogen transcriptomic studies have not yielded homogenous results that can be harnessed into a reliable diagnostic tool. Here we utilized a multi-cohort analysis approach using machine-learning algorithms to identify blood gene signatures that can distinguish severe and cerebral malaria from moderate and non-cerebral cases. Using a Regularized Random Forest model, we identified 28-gene and 32-gene signatures that can reliably distinguish severe and cerebral malaria, respectively. We tested the specificity of both signatures against other common infectious diseases to ensure the signatures reliability and suitability as diagnostic markers. The severe and cerebral malaria gene-signatures were further integrated through k-top scoring pairs classifiers into ten and nine gene pairs that could distinguish severe and cerebral malaria, respectively. These signatures have various implications that can be utilized as blood diagnostic tools for malaria severity in endemic countries.

Next- and Third-Generation Sequencing Outperforms Culture-Based Methods in the Diagnosis of Ascitic Fluid Bacterial Infections of ICU Patients.

OBJECTIVES: Infections of the ascitic fluid are serious conditions that require rapid diagnosis and treatment. Ascites is often accompanied by other critical pathologies such as gastrointestinal bleeding and bowel perforation, and infection increases the risk of mortality in intensive care patients. Owing to a relatively low success rate of conventional culture methods in identifying the responsible pathogens, new methods may be helpful to guide antimicrobial therapy and to refine empirical regimens. Here, we aim to assess outcomes and to identify responsible pathogens in ascitic fluid infections, in order to improve patients' care and to guide empirical therapy. METHODS: Between October 2019 and March 2021, we prospectively collected 50 ascitic fluid samples from ICU patients with suspected infection. Beside standard culture-based microbiology methods, excess fluid underwent DNA isolation and was analyzed by next- and third-generation sequencing (NGS) methods. RESULTS: NGS-based methods had higher sensitivity in detecting additional pathogenic bacteria such as E. faecalis and Klebsiella in 33 out of 50 (66%) ascitic fluid samples compared with culture-based methods (26%). Anaerobic bacteria were especially identified by sequencing-based methods in 28 samples (56%), in comparison with only three samples in culture. Analysis of clinical data showed a correlation between sequencing results and various clinical parameters such as peritonitis and hospitalization outcomes. CONCLUSIONS: Our results show that, in ascitic fluid infections, NGS-based methods have a higher sensitivity for the identification of clinically relevant pathogens than standard microbiological culture diagnostics, especially in detecting hard-to-culture anaerobic bacteria. Patients with such infections may benefit from the use of NGS methods by the possibility of earlier and better targeted antimicrobial therapy, which has the potential to lower the high morbidity and mortality in critically ill patients with ascitic bacterial infection.

The AML-associated K313 mutation enhances C/EBPα activity by leading to C/EBPα overexpression.

Mutations in the transcription factor C/EBPα are found in ~10% of all acute myeloid leukaemia (AML) cases but the contribution of these mutations to leukemogenesis is incompletely understood. We here use a mouse model of granulocyte progenitors expressing conditionally active HoxB8 to assess the cell biological and molecular activity of C/EBPα-mutations associated with human AML. Both N-terminal truncation and C-terminal AML-associated mutations of C/EBPα substantially altered differentiation of progenitors into mature neutrophils in cell culture. Closer analysis of the C/EBPα-K313-duplication showed expansion and prolonged survival of mutant C/EBPα-expressing granulocytes following adoptive transfer into mice. C/EBPα-protein containing the K313-mutation further showed strongly enhanced transcriptional activity compared with the wild-type protein at certain promoters. Analysis of differentially regulated genes in cells overexpressing C/EBPα-K313 indicates a strong correlation with genes regulated by C/EBPα. Analysis of transcription factor enrichment in the differentially regulated genes indicated a strong reliance of SPI1/PU.1, suggesting that despite reduced DNA binding, C/EBPα-K313 is active in regulating target gene expression and acts largely through a network of other transcription factors. Strikingly, the K313 mutation caused strongly elevated expression of C/EBPα-protein, which could also be seen in primary K313 mutated AML blasts, explaining the enhanced C/EBPα activity in K313-expressing cells.

Draft Genome Sequences and Antimicrobial Profiles of Three Staphylococcus epidermidis Strains from Neonatal Blood Samples.

Data on molecular characterization of coagulase-negative staphylococci causing neonatal sepsis in low-income countries are highly limited. This report highlights the isolation of three Staphylococcus epidermidis non-genome assembly strains (NGASs) from blood samples from neonates with unknown transmission sources. Pathogenic factors and sources of transmission of these strains warrant further investigation.

Comprehensive Integration of Genome-Wide Association and Gene Expression Studies Reveals Novel Gene Signatures and Potential Therapeutic Targets for Helicobacter pylori-Induced Gastric Disease.

Helicobacter pylori is a gram-negative bacterium that colonizes the human gastric mucosa and can lead to gastric inflammation, ulcers, and stomach cancer. Due to the increase in H. pylori antimicrobial resistance new methods to identify the molecular mechanisms of H. pylori-induced pathology are urgently needed. Here we utilized a computational biology approach, harnessing genome-wide association and gene expression studies to identify genes and pathways determining disease development. We mined gene expression data related to H. pylori-infection and its complications from publicly available databases to identify four human datasets as discovery datasets and used two different multi-cohort analysis pipelines to define a H. pylori-induced gene signature. An initial Helicobacter-signature was curated using the MetaIntegrator pipeline and validated in cell line model datasets. With this approach we identified cell line models that best match gene regulation in human pathology. A second analysis pipeline through NetworkAnalyst was used to refine our initial signature. This approach defined a 55-gene signature that is stably deregulated in disease conditions. The 55-gene signature was validated in datasets from human gastric adenocarcinomas and could separate tumor from normal tissue. As only a small number of H. pylori patients develop cancer, this gene-signature must interact with other host and environmental factors to initiate tumorigenesis. We tested for possible interactions between our curated gene signature and host genomic background mutations and polymorphisms by integrating genome-wide association studies (GWAS) and known oncogenes. We analyzed public databases to identify genes harboring single nucleotide polymorphisms (SNPs) associated with gastric pathologies and driver genes in gastric cancers. Using this approach, we identified 37 genes from GWA studies and 61 oncogenes, which were used with our 55-gene signature to map gene-gene interaction networks. In conclusion, our analysis defines a unique gene signature driven by H. pylori-infection at early phases and that remains relevant through different stages of pathology up to gastric cancer, a stage where H. pylori itself is rarely detectable. Furthermore, this signature elucidates many factors of host gene and pathway regulation in infection and can be used as a target for drug repurposing and testing of infection models suitability to investigate human infection.

Antimicrobial Susceptibility Patterns and Wild-Type MIC Distributions of Anaerobic Bacteria at a German University Hospital: A Five-Year Retrospective Study (2015-2019).

Local antimicrobial susceptibility surveys are crucial for optimal empirical therapy guidelines and for aiding in antibiotic stewardship and treatment decisions. For many laboratories, a comprehensive overview of local antimicrobial susceptibility patterns of anaerobic bacteria is still lacking due to the long incubation time and effort involved. The present study investigates the antimicrobial susceptibility patterns and related clinical and demographic data of 2856 clinical isolates of anaerobic bacteria that were submitted for analysis to the Institute for Medical Microbiology and Hygiene of the Freiburg University Medical Center (a tertiary university medical center in Southern Germany) between 2015 and 2019. Antimicrobial susceptibility testing has been carried out according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guideline. Minimum inhibitory concentration (MIC)50 and MIC90 for penicillin, metronidazole, moxifloxacin, and clindamycin were established for Gram-positive anaerobes and for ampicillin-sulbactam, meropenem, metronidazole, moxifloxacin, and clindamycin for Gram-negative anaerobes. The distribution of MIC-values for various antibiotics against anaerobic bacteria was also established, especially for those having no specific breakpoints according to EUCAST guidelines. Most clinically relevant anaerobic bacteria originated from general surgery, neurological, and orthopedic wards. A high proportion of isolates were resistant to moxifloxacin and clindamycin indicating the importance of their susceptibility testing before administration. Based on our study metronidazole and other ß-lactam/ß-lactamase inhibitor combinations such as ampicillin-sulbactam remain suitable for empirical treatment of infections with anaerobic bacteria.