Human papilloma virus (HPV) integration signature in Cervical Cancer: identification of MACROD2 gene as HPV hot spot integration site

Background Cervical cancer (CC) remains a leading cause of gynaecological cancer-related mortality with infection by human papilloma virus (HPV) being the most important risk factor. We analysed the association between different viral integration signatures, clinical parameters and outcome in pre-treated CCs.Methods Different integration signatures were identified using HPV double capture followed by next-generation sequencing (NGS) in 272 CC patients from the BioRAIDs study [{"type":"clinical-trial","attrs":{"text":"NCT02428842","term_id":"NCT02428842"}}NCT02428842]. Correlations between HPV integration signatures and clinical, biological and molecular features were assessed.Results Episomal HPV was much less frequent in CC as compared to anal carcinoma (p < 0.0001). We identified >300 different HPV-chromosomal junctions (inter- or intra-genic). The most frequent integration site in CC was in MACROD2 gene followed by MIPOL1/TTC6 and TP63. HPV integration signatures were not associated with histological subtype, FIGO staging, treatment or PFS. HPVs were more frequently episomal in PIK3CA mutated tumours (p = 0.023). Viral integration type was dependent on HPV genotype (p < 0.0001); HPV18 and HPV45 being always integrated. High HPV copy number was associated with longer PFS (p = 0.011).Conclusions This is to our knowledge the first study assessing the prognostic value of HPV integration in a prospectively annotated CC cohort, which detects a hotspot of HPV integration at MACROD2; involved in impaired PARP1 activity and chromosome instability.Subject terms: Oncology, Molecular medicine, Biomarkers, Molecular biology     

Effect of residual doxycycline concentrations on resistance selection and transfer in porcine commensal Escherichia coli

Pig feed may contain various levels of antimicrobial residues due to cross-contamination. A previous study showed that a 3% carry-over level of doxycycline (DOX) in the feed results in porcine faecal concentrations of approximately 4?mg/L.The aim of this study was to determine the effect of residual DOX concentrations (1 and 4?mg/L) in vitro on selection of DOX–resistant porcine commensal Escherichia coli and transfer of their resistance plasmids.Three different DOX–resistant porcine commensal E. coli strains and their plasmids were characterised. These strains were each brought in competition with a susceptible strain in a medium containing 0, 1 and 4?mg/L DOX. Resistant bacteria, susceptible bacteria and transconjugants were enumerated after 24?h and 48?h.The tet(A)–carrying plasmids showed genetic backbones that are also present among human E. coli isolates. Ratios of resistant to susceptible bacteria were significantly higher at 1 and 4?mg/L DOX compared with the blank control, but there was no significant difference between 1 and 4?mg/L. Plasmid transfer frequencies were affected by 1 or 4?mg/L DOX in the medium for only one of the resistance plasmids.In conclusion, DOX concentrations of 1 and 4?mg/L can select for resistant E. coli in vitro. Further research is needed to determine the effect of these concentrations in the complex environment of the porcine intestinal microbiota.     

Association between ADHD drug use and injuries among children and adolescents

To study the association between attention deficit hyperactivity disorder (ADHD) drug use and the incidence of hospitalization due to injuries. A random sample of 150,000 persons (0–18 years) was obtained from the Dutch PHARMO record linkage system. An ADHD medication cohort as well as an up to six age/sex/index date sampled control cohort with no history of ADHD drug use was formed. Differences in incidence of hospitalization due to injuries were stratified for age and sex and compared prior, during and after exposure on ADHD drugs. The overall incidence of hospital admissions for injuries was two times higher in the ADHD medication cohort [incidence rate ratios (IRR) 2.2 (95 % CI 1.6–2.9)]. The incidence rate for injuries during exposure to ADHD drugs was lower in the exposed period compared to the period prior to ADHD drug use, although the difference was not statistically significant [IRR 0.68 (95 % CI 0.29–1.60)]. The relative risk for injuries was almost five times higher in the ADHD medication cohort among those who concomitantly used other psychotropics [IRR 4.8 (95 % CI 1.4–16.9)]. Risk for injuries was highest in 12–18 years olds. Children and adolescents using ADHD medication showed a twofold risk for hospital admissions for injuries. ADHD drug use might diminish the increased injury risk, but still overall risk is higher than in age/sex sampled children and adolescents without treatment with ADHD drugs. Use of ADHD and concomitant psychotropics increases the risk for injuries compared to only ADHD drug use.     

Superior Sensitivity of Ex Vivo IFN‐γ Release Assays as Compared to Skin Testing in Immunocompromised Patients

Comparative assessment of the tuberculin skin testing (TST) and commercial IFN‐γ release‐assays (IGRAs) is hampered by the use of different antigens (tuberculin PPD in TST vs. ESAT‐6/CFP‐10 in IGRAs). Thus, PPD was used as a common stimulus to compare performance of the TST and three IGRAs in 72 controls, 101 hemodialysis patients and 100 renal transplant recipients. Results of the TST were compared with PPD‐induced IFN‐γ induction in vitro detected by ELISPOT, ELISA or a flow‐cytometric FACS assay. Percentages of positive tests were significantly lower in TST (9.2%) compared to ELISA (55.3%), ELISPOT (45.3%) and FACS (44.9%, p < 0.0001). Agreement between TST and IGRAs was highest for controls (κ = 0.19–0.32) and poor in immunocompromised patients (κ = 0 for transplant patients, κ = 0.06–0.13 for hemodialysis patients). Discrepant results were largely TST negative and IGRA positive. Among IGRAs, agreement was highest between ELISPOT and FACS (κ = 0.61). Unlike TST, all IGRAs were associated with variables of mycobacterial exposure. Among IGRAs, the FACS assay was least affected by the level of immunosuppression. In conclusion, both the percentage of positive results and between‐test‐agreement were higher with IGRAs as compared to TST. This indicates superiority of IGRAs in detecting a PPD‐specific immune response which may also apply for immunity toward Mycobacterium tuberculosis–specific antigens.     

Jagged2 acts as a Delta-like Notch ligand during early hematopoietic cell fate decisions

Notch signaling critically mediates various hematopoietic lineage decisions and is induced in mammals by Notch ligands that are classified into 2 families, Delta-like (Delta-like-1, -3 and -4) and Jagged (Jagged1 and Jagged2), based on structural homology with both Drosophila ligands Delta and Serrate, respectively. Because the functional differences between mammalian Notch ligands were still unclear, we have investigated their influence on early human hematopoiesis and show that Jagged2 affects hematopoietic lineage decisions very similarly as Delta-like-1 and -4, but very different from Jagged1. OP9 coculture experiments revealed that Jagged2, like Delta-like ligands, induces T-lineage differentiation and inhibits B-cell and myeloid development. However, dose-dependent Notch activation studies, gene expression analysis, and promoter activation assays indicated that Jagged2 is a weaker Notch1-activator compared with the Delta-like ligands, revealing a Notch1 specific signal strength hierarchy for mammalian Notch ligands. Strikingly, Lunatic-Fringe- mediated glycosylation of Notch1 potentiated Notch signaling through Delta-like ligands and also Jagged2, in contrast to Jagged1. Thus, our results reveal a unique role for Jagged1 in preventing the induction of T-lineage differentiation in hematopoietic stem cells and show an unexpected functional similarity between Jagged2 and the Delta-like ligands.