Human papillomavirus vaccination of girls in the German model region Saarland: Insurance data-based analysis and identification of starting points for improving vaccination rates

In Germany, the incidence of cervical cancer, a disease caused by human papillomaviruses (HPV), is higher than in neighboring European countries. HPV vaccination has been recommended for girls since 2007. However, it continues to be significantly less well received than other childhood vaccines, so its potential for cancer prevention is not fully realized. To find new starting points for improving vaccination rates, we analyzed pseudonymized routine billing data from statutory health insurers in the PRÄZIS study (prevention of cervical carcinoma and its precursors in women in Saarland) in the federal state Saarland serving as a model region. We show that lowering the HPV vaccination age to 9 years led to more completed HPV vaccinations already in 2015. Since then, HPV vaccination rates and the proportion of 9- to 11-year-old girls among HPV-vaccinated females have steadily increased. However, HPV vaccination rates among 15-year-old girls in Saarland remained well below 50% in 2019. Pediatricians vaccinated the most girls overall, with a particularly high proportion at the recommended vaccination age of 9–14 years, while gynecologists provided more HPV catch-up vaccinations among 15-17-year-old girls, and general practitioners compensated for HPV vaccination in Saarland communities with fewer pediatricians or gynecologists. We also provide evidence for a significant association between attendance at the children´s medical check-ups “U11” or “J1” and HPV vaccination. In particular, participation in HPV vaccination is high on the day of U11. However, obstacles are that U11 is currently not financed by all statutory health insurers and there is a lack of invitation procedures for both U11 and J1, resulting in significantly lower participation rates than for the earlier U8 or U9 screenings, which are conducted exclusively with invitations and reminders. Based on our data, we propose to restructure U11 and J1 screening in Germany, with mandatory funding for U11 and organized invitations for HPV vaccination at U11 or J1 for both boys and girls

German federal-state-wide seroprevalence study of 1st SARS-CoV-2 pandemic wave shows importance of long-term antibody test performance

Background Reliable data on the adult SARS-CoV-2 infection fatality rate in Germany are still scarce. We performed a federal state-wide cross-sectional seroprevalence study named SaarCoPS, that is representative for the adult population including elderly individuals and nursing home residents in the Saarland. Methods Serum was collected from 2940 adults via stationary or mobile teams during the 1st pandemic wave steady state period. We selected an antibody test system with maximal specificity, also excluding seroreversion effects due to a high longitudinal test performance. For the calculations of infection and fatality rates, we accounted for the delays of seroconversion and death after infection. Results Using a highly specific total antibody test detecting anti-SARS-CoV-2 responses over more than 180 days, we estimate an adult infection rate of 1.02% (95% CI: [0.64; 1.44]), an underreporting rate of 2.68-fold (95% CI: [1.68; 3.79]) and infection fatality rates of 2.09% (95% CI: (1.48; 3.32]) or 0.36% (95% CI: [0.25; 0.59]) in all adults including elderly individuals, or adults younger than 70 years, respectively. Conclusion The study highlights the importance of study design and test performance for seroprevalence studies, particularly when seroprevalences are low. Our results provide a valuable baseline for evaluation of future pandemic dynamics and impact of public health measures on virus spread and human health in comparison to neighbouring countries such as Luxembourg or France

The Slicer Activity of ARGONAUTE1 is Required Specifically for the Phasing, Not Production, of Trans-Acting Short Interfering RNAs in Arabidopsis

ARGONAUTE1 (AGO1) mediates posttranscriptional silencing by microRNAs (miRNAs) and short interfering RNAS (siRNAs). AGO1-catalyzed RNA cleavage (slicing) represses miRNA targets, but current models also highlight the roles of slicing in formation of siRNAs and siRNA-AGO1 complexes. miRNA-guided slicing is required for biogenesis of phased, trans-acting siRNAs (tasiRNAs), whose cleaved precursor fragments are converted to double-stranded RNA by RNA-dependent RNA polymerase 6 (RDR6). In addition, unwinding of duplex siRNA bound to AGO1 requires passenger strand cleavage in vitro. In this study, we analyze how mutation of four metal ion-coordinating residues of Arabidopsis thaliana AGO1 affects slicer activity in vitro and siRNA function in vivo. We show that while all four residues are required for slicer activity, they do not contribute equally to catalysis. Moreover, passenger strand cleavage is required for assembly of active AGO1-siRNA complexes in vivo, and many AGO1-bound siRNAs are trimmed in the absence of slicer activity. Remarkably, seedlings defective in AGO1 slicer activity produce abundant siRNAs from tasiRNA loci in vivo. These siRNAs depend on RDR6 and SUPPRESSOR OF GENE SILENCING3, but unlike wild-type tasiRNAs, they are unphased. These results demonstrate that slicing is solely required for phase definition of tasiRNAs, and they strongly support recruitment of RDR6 by AGO1 rather than by cleavage fragments

Argonaute Family Protein Expression in Normal Tissue and Cancer Entities.

The members of the Argonaute (AGO) protein family are key players in miRNA-guided gene silencing. They enable the interaction between small RNAs and their respective target mRNA(s) and support the catalytic destruction of the gene transcript or recruit additional proteins for downstream gene silencing. The human AGO family consists of four AGO proteins (AGO1-AGO4), but only AGO2 harbors nuclease activity. In this study, we characterized the expression of the four AGO proteins in cancer cell lines and normal tissues with a new mass spectrometry approach called AGO-APP (AGO Affinity Purification by Peptides). In all analyzed normal tissues, AGO1 and AGO2 were most prominent, but marked tissue-specific differences were identified. Furthermore, considerable changes during development were observed by comparing fetal and adult tissues. We also identified decreased overall AGO expression in melanoma derived cell lines compared to other tumor cell lines and normal tissues, with the largest differences in AGO2 expression. The experiments described in this study suggest that reduced amounts of AGO proteins, as key players in miRNA processing, have impact on several cellular processes. Deregulated miRNA expression has been attributed to chromosomal aberrations, promoter regulation and it is known to have a major impact on tumor development and progression. Our findings will further increase our basic understanding of the molecular basis of miRNA processing and its relevance for disease

Argonaute protein distribution in melanoma and non-melanoma cell lines.

<p>(A) AGO protein percentage distribution of each AGO to total AGO protein amount in non-melanoma cell lines CaCo2, HepG2, SW1353, MCF7, HeLa and melanoma cell lines Mel Ju, Mel Im, Mel Wei, Mel Ei and Mel Ho derived from AGO-APP. (B) Total protein amount determination of each AGO enriched by AGO-APP in CaCo2, HepG2, SW1353, MCF7, HeLa, Mel Ju, Mel Im, Mel Wei, Mel Ei and Mel Ho. (C) Average AGO protein amount of each AGO in non-melanoma compared to melanoma cell lines. The reduced AGO concentration in melanoma cell lines compared to non-melanoma cell lines is only significant for AGO2. ** = p<0.01 (D) AGO1 western blot analysis and corresponding (E) Western blot quantification of melanoma cell lines (Mel Ju, Mel Im, Mel Wei, Mel Ei, Mel Ho) and non-melanoma cell lines (CaCo2, SW1353, MCF7, HeLa, HepG2). The two additional values in the AGO1 western blot quantification illustrate the average AGO1 concentration in melanoma and non-melanoma cell lines. Quantification was done relative to Actin in the respective blot.</p

Argonaute gene expression in different human healthy tissues compared to NHEMs.

<p>Relative (A) AGO1, (B) AGO2, (C) AGO3 and (D) AGO4 mRNA expression in NHEMs (derived from different cultivation passages (P4-P6) from three different donors respectively), skin (derived from three different tissue samples) and heart, kidney, bone marrow, fetal brain, bowel, thymus, uterus, trachea, brain, muscle, bone marrow, liver, fetal liver, brain and fetal brain (derived from a total RNA bank and shown as technical replicates). (E) The compilation of percentage distribution of each AGO to the total AGO amount in the respective cell line or tissue.</p

Oligonucleotide sequences and qRT-PCR conditions.

<p>Oligonucleotide sequences and qRT-PCR conditions.</p

Argonaute gene expression in different melanoma and non-melanoma cell lines.

<p>Relative (A) AGO1, (B) AGO2, (C) AGO3 and (D) AGO4 mRNA expression in melanoma cell lines derived from primary tumors (Mel Ei, Mel Juso, Mel Ho and Mel Wei) and metastases (Mel Ju, Mel Im, SkMel28 and Hmb2) and other non-melanoma cell lines (HeLa, CaCo2, PLC, Jurkat, Hep3b, SW1353 and MCF7). Each point shows the measurement of one independently derived cDNA sample. Bars show mean and S.D. (E) The compilation of percentage distribution of each AGO to the aggregate AGO amount. (F) Entire mRNA expression of all four AGOs compared to actin in melanoma cell lines derived from primary tumors or metastases and in other non-melanoma cell lines.</p