Identification of Naturally Processed Hepatitis C Virus-Derived Major Histocompatibility Complex Class I Ligands

Fine mapping of human cytotoxic T lymphocyte (CTL) responses against hepatitis C virus (HCV) is based on external loading of target cells with synthetic peptides which are either derived from prediction algorithms or from overlapping peptide libraries. These strategies do not address putative host and viral mechanisms which may alter processing as well as presentation of CTL epitopes. Therefore, the aim of this proof-of-concept study was to identify naturally processed HCV-derived major histocompatibility complex (MHC) class I ligands. To this end, continuous human cell lines were engineered to inducibly express HCV proteins and to constitutively express high levels of functional HLA-A2. These cell lines were recognized in an HLA-A2-restricted manner by HCV-specific CTLs. Ligands eluted from HLA-A2 molecules isolated from large-scale cultures of these cell lines were separated by high performance liquid chromatography and further analyzed by electrospray ionization quadrupole time of flight mass spectrometry (MS)/tandem MS. These analyses allowed the identification of two HLA-A2-restricted epitopes derived from HCV nonstructural proteins (NS) 3 and 5B (NS31406–1415 and NS5B2594–2602). In conclusion, we describe a general strategy that may be useful to investigate HCV pathogenesis and may contribute to the development of preventive and therapeutic vaccines in the future

Item Level Cataloguing in the Austrian Library Network: Results of a Working Group of the Austrian Cataloguing Committee

Die Bedeutung der exemplarspezifischen Erschließung – also der Darstellung von besonderen Merkmalen und der Vorgeschichte eines bestimmten Buches im Katalog – ist in den vergangenen Jahren im Zusammenhang mit der Erforschung und Rekonstruktion alter und wertvoller Bestände, der NS-Provenienzforschung und der Digitalisierung gewachsen. Um gemeinsame Lösungen für die Erschließung, vor allem aber für die verbundweite Darstellung und Suchbarkeit von Exemplarspezifika zu finden, wurde eine Arbeitsgruppe der Zentralen Redaktion des Österreichischen Bibliothekenverbundes eingerichtet. Der Beitrag präsentiert die Ergebnisse der Arbeitsgruppe, die Arbeitsunterlagen für die Erschließung ebenso umfassen wie das Setup der Verbundsuchmaschine, das spezifische Suchen, exemplarspezifische Links und nicht zuletzt die provisorische Nutzung von Normdaten erlaubt.The importance of cataloguing on an item level – i.e., the presentation of special features as well as data on the history of a particular book in the catalogue – increased in recent years due to research on and reconstruction of old and valuable holdings, research concerning holdings of Nazi-provenance, and digitization. In order to find common solutions for cataloguing, but especially for the network-wide presentation and searchability of item level information, a working group of the Austrian Cataloguing Committee was established. The article presents the results of the working group, including working documents for indexing as well as the necessary set-up of the Consortium Search Interface, which allows for specific searches, item-specific links and, last but not least, the provisional use of authority data

Recommendations for the Visibility of Open Access Publications in the Search Engine of the Austrian Library Network: Report of the OBV Working Group “Repositories in the Network”

Im vorliegenden Beitrag werden die Ergebnisse der OBV-Arbeitsgruppe „Repositorien im Verbund“ präsentiert. Die AG verfolgte das Ziel, einen Leitfaden für die Erfassung von Metadaten für Objekte in Repositorien, der dazu beiträgt, einheitliche Standards in dieser Hinsicht zu entwickeln, mit dessen Hilfe es in weiterer Folge ermöglicht werden soll, Repositorienbestände ohne Erzeugung von Dubletten in Alma bzw. im Verbundkatalog nachzuweisen. Weitere Ziele waren die Erarbeitung von Empfehlungen für eine zentrale Bereitstellung von Metadaten von Open Access-Publikationen zur Vereinfachung der lokalen Workflows (analog zum DFG-geförderten Projekt DeepGreen) mittels Teilautomatisierung sowie von Empfehlungen für eine Etablierung eines Reiters für Open Access-Materialien in der Suchmaschine des Österreichischen Bibliothekenverbundes (analog zu den Reitern „Fachliteratur“, „Hochschulschriften“ und „Nachlässe / Handschriften“).This paper presents the results of the OBV working group “Repositories in the Austrian Library Network”. The aim of the working group was to develop a guideline for the registration of metadata for objects in repositories, which would contribute to the development of uniform standards in this regard, and with the help of which it should subsequently be possible to identify repository holdings in Alma or in the Austrian Union Catalogue without creating duplicates. Further goals were the development of recommendations for a central provision of metadata of open access publications to simplify local workflows (analogous to the DFG-funded project DeepGreen) by means of partial automation, as well as recommendations for establishing a tab for open access materials in the search engine of the Austrian Library Network (analogous to the tabs “Literature”, “Theses and Dissertations” and “Bequests / Autographs”)

Studie zur Evaluation der Reproduzierbarkeit und Objektivität zweier Schmerzskalen bei der Katze

Die mangelhafte Schmerztherapie von Katzen in der tierärztlichen Praxis ist unter anderem in der Schwierigkeit der Schmerzerkennung begründet. Um diesem Problem zu begegnen, braucht es den Einsatz von möglichst objektiven Schmerzerkennungsskalen als Hilfsmittel. Im Rahmen der vorliegenden Studie wurden eine multidimensionale Schmerzevaluationsskala „Composite Pain Scale“, CPS, und eine interaktive visuelle analoge Skala „Dynamic and Interactive Visual Analogue Scale“, DIVAS, auf ihre Reproduzierbarkeit und Variabilität zwischen verschiedenen Beobachtern überprüft. Anhand von Videoaufnahmen wurden 21 Katzen nach einer Ovariohysterektomie von 18 Beobachtern (8 Veterinäranästhesisten, 5 Privattierärzte und 5 Studenten der Veterinärmedizin) zu zwölf Zeitpunkten mit der CPS nach Al-Gizawiy und Rudé (2004) und einer DIVAS bezüglich dem Schmerzempfinden evaluiert. Es wurde eine Varianzanalyse durchgeführt, welche eine grosse Variabilität (p<0.05) zwischen den Beobachtern zeigte. Sowohl die Totalwerte der CPS und die DIVAS, als auch die einzelnen Kategorien der CPS variierten zu fast allen Zeitpunkten. Bei beiden Skalen lag die grösste Streuung der Resultate innerhalb der ersten vier Stunden nach der Operation. Die Kappa-Test-Analyse ergab für beide Skalen eine schlechte Reproduzierbarkeit. Somit konnte gezeigt werden, dass weder die angewendete CPS noch die DIVAS dazu geeignet sind, Schmerz bei Katzen durch verschiedene Beobachter objektiv zu beurteilen Summary Pain in cats is often undertreated. Among other reasons this is due to difficulties in recognition of pain in this species. To solve this problem, objective pain assessment scales are needed. In the present study, a multidimensional pain scale (Composite Pain Scale, CPS) and a dynamic and interactive visual analogue scale (DIVAS) have been tested regarding their inter-observer variability and reproducibility for pain assessment in cats. Based on video recordings, 18 observers (8 veterinary anesthetists, 5 private veterinarians and 5 veterinary students) evaluated 21 cats at twelve different time points pre- and postoperatively after ovariohysterectomy with a CPS from Al-Gizawiy and Rudé (2004) and a DIVAS. An analysis of variance was performed. The results show a large inter-observer variability (p<0.05), for all observers as well as between groups. They show for total CPS and DIVAS as well as in separate categories of the CPS significant differences between observers at most of the time points. Both scales showed the widest variation during the first four hours after surgery. The kappa test analysis revealed a poor reproducibility. Neither the CPS used in this study nor the DIVAS are reliable scales in cats for an impartial pain assessment by different observers

Detection and characterization of peptide NS3_1406–1415 in MHC class I ligands isolated from UNS3-4A/A2-27.35 cells.

<p>Naturally processed MHC class I ligands of UNS3-4A/A2-27.35 cells were separated by high performance liquid chromatography (HPLC) and analyzed by mass spectrometry. (A) A chromatogram of the total ion current (TIC) is shown. (B) The mass chromatogram of ionized peptides with m/z = 499.3 reveals a peak at retention time t = 49.5 min that correlates with (C) a signal of the synthetic peptide NS3_1406–1415 (KLVALGINAV) which elutes after 51.3 min. In all graphs, HPLC retention times are shown on the abscissa and relative signal intensity on the ordinate. Small insets in panels B and C show mass spectra of peptides eluted at the indicated time points. (D) MSMS spectrum of the synthetic peptide NS3_1406–1415 (KLVALGINAV). (E) MSMS spectrum of the natural peptide isolated from UNS3-4A/A2-27.35 cells, revealing amino acid sequence KLVALGINAV identical to the synthetic peptide shown in panel A. Identified amino acid sequences of peptide fragments are indicated on the top of the peaks. Due to the protonated N-terminal lysine residue, the b series of peptide fragments is dominating the MSMS spectra.</p

Cell lines UNS3-4A/A2-27.35 and UHCV/A2-27.

<p>(A) Indirect immunofluorescence microscopy of UNS3-4A/A2-27.35 and UHCV/A2-27 cells cultured for 48 h in the presence (+ tet) or absence (− tet) of tetracycline. Monoclonal antibody 1B6 against HCV NS3 was used as primary antibody. (B) Immunoblot analysis of UNS3-4A/A2-27.35 and UHCV/A2-27 cells were cultured for 48 h in the presence (+ tet) or absence (− tet) of tetracycline. Monoclonal antibodies 1B6 against HCV NS3 or 11H against NS5A were used as primary antibodies. (C) Left panel: HLA-A2 surface expression of UNS3-4A/A2-27.35 (blue histogram) compared to the founder cell line UNS3-4A-24 (red histogram). Right panel: HLA-A2 surface expression of UHCV/A2-27 (blue histogram) compared to the founder cell line UHCVcon-57.3 (red histogram).</p

Identification of naturally processed HLA-A2 ligands.

<p>Following large-scale expansion, cell pellets were lysed and MHC class I molecules with bound ligands were purified by immunoprecipitation. Subsequently, ligands were eluted and separated by high performance liquid chromatography (HPLC). Fractions of interest were sequenced by mass spectrometry.</p

Assessment of the situation in our study areas regarding risk factors for bTB maintenance.

*<p>Due to organizational reasons, sampling took place only during the 2010/11 hunting season. Apparent prevalence was calculated based on combined results of PCR investigations on tissue samples and of isolation attempts of <i>M. bovis</i> and/or <i>M. caprae</i>. Methods for estimating population densities were: capture-resight (Geneva); calculations by the first author using data obtained by direct counts conducted by game wardens in the Swiss National Park, where hunting is prohibited (Grisons; for this canton, no data from the exact location of sampling regions were available); head-light counts (Saint Gall and Liechtenstein). The situation regarding risk factors within the study areas was assessed by a telephone survey among hunting officials. The following questions were asked: (1) Do aggregation sites exist, where wild boar and/or red deer frequently gather in high numbers?; (2) Is supplemental feeding of wild boar and/or red deer permitted and/or commonly practiced?; (3) Is offal of hunted wild boar and red deer regularly left in the fields?; (4) Are there areas of private property where wild boar and red deer are intensively managed and hunted? (similar to the hunting industry abroad, e.g. in Spain).</p

Increase of population numbers and hunting bags of red deer and wild boar in Switzerland.

<p>Red deer counts (black line); red deer hunting bag (dashed line); wild boar hunting bag (dotted line). No wild boar counts available. (Source: Swiss hunting statistics: <a href="http://www.wild.uzh.ch/jagdst/" target="_blank">http://www.wild.uzh.ch/jagdst/</a>).</p

Macroscopic and microbiological results, red deer.

<p>Real-time PCR from tissue material (PCR_T) and real-time PCR from culture (PCR_C) for the detection of mycobacteria of the Mycobacterium tuberculosis-complex (MTBC). Positive mycobacterial cultures negative by PCR_C were classified as atypical mycobacteria. Cultures were considered positive if typical growth occurred and acid-fast bacilli were subsequently detected with Ziehl-Neelsen staining.</p