Plasma Cell-Free DNA Levels Are Elevated in Acute Puumala Hantavirus Infection

Peer reviewe

Fatal Outcome in Bacteremia is Characterized by High Plasma Cell Free DNA Concentration and Apoptotic DNA Fragmentation: A Prospective Cohort Study

INTRODUCTION: Recent studies have shown that apoptosis plays a critical role in the pathogenesis of sepsis. High plasma cell free DNA (cf-DNA) concentrations have been shown to be associated with sepsis outcome. The origin of cf-DNA is unclear. METHODS: Total plasma cf-DNA was quantified directly in plasma and the amplifiable cf-DNA assessed using quantitative PCR in 132 patients with bacteremia caused by Staphylococcus aureus, Streptococcus pneumoniae, ß-hemolytic streptococcae or Escherichia coli. The quality of cf-DNA was analyzed with a DNA Chip assay performed on 8 survivors and 8 nonsurvivors. Values were measured on days 1-4 after positive blood culture, on day 5-17 and on recovery. RESULTS: The maximum cf-DNA values on days 1-4 (n = 132) were markedly higher in nonsurvivors compared to survivors (2.03 vs 1.26 ug/ml, p<0.001) and the AUCROC in the prediction of case fatality was 0.81 (95% CI 0.69-0.94). cf-DNA at a cut-off level of 1.52 ug/ml showed 83% sensitivity and 79% specificity for fatal disease. High cf-DNA (>1.52 ug/ml) remained an independent risk factor for case fatality in a logistic regression model. Qualitative analysis of cf-DNA showed that cf-DNA displayed a predominating low-molecular-weight cf-DNA band (150-200 bp) in nonsurvivors, corresponding to the size of the apoptotic nucleosomal DNA. cf-DNA concentration showed a significant positive correlation with visually graded apoptotic band intensity (R = 0.822, p<0.001). CONCLUSIONS: Plasma cf-DNA concentration proved to be a specific independent prognostic biomarker in bacteremia. cf-DNA displayed a predominating low-molecular-weight cf-DNA band in nonsurvivors corresponding to the size of apoptotic nucleosomal DNA

Pienten näytteiden dissekointiohjeistus : Työohjeet bioanalytiikan opiskelijoille

Histologia on patologian eli tautiopin osa-alue, jossa tutkitaan kudosnäytteitä. Kliinisessä histologiassa lääkäri tutkii kudosnäytteen mikroskooppisesti ja antaa havainnoistaan lausunnon ja diagnoosin, joiden avulla päätetään potilaan hoidosta. Jotta kudosnäytteen voi tutkia mikroskooppisesti, tarvitsee se ensin käsitellä laboratoriossa prosessissa, jonka vaiheita ovat esikäsittely, kudoskuljetus, leikkaus ja värjäys. Esikäsittelyvaihe pitää sisällään makroleikkelyn eli dissekoinnin, jonka tarkoituksena on varmistaa, että mikroskooppitutkimukseen päätyvä kudosleike on näytteen oikeasta kohdasta otettu. Patologian laboratoriossa bioanalyytikon työnkuvaan kuuluu tiettyjen pienten kudosnäytteiden dissekointi. Tämän opinnäytetyön tarkoituksena oli laatia pienten kudosnäytteiden dissekointiohjeistus Tampereen ammattikorkeakoulun bioanalyytikon tutkinto-ohjelman opiskelijoiden käyttöön. Ohjeistuksen kudokset on valittu Fimlab Laboratoriot Oy:n työntekijöiden avustuksella ja niiden on tarkoitus edustaa yleisimpiä bioanalyytikon dissektioon tulevia pieniä kudosnäytteitä. Tavoitteena oli luoda selkeä, yksinkertainen ja helppokäyttöinen oppimateriaali. Opinnäytetyömme on toiminnallinen opinnäytetyö eli se koostuu raporttiosuudesta ja produktina syntyneistä työohjeista. Kirjallisessa osuudessa käsittelemme oppimista, hyvän oppimateriaalin kriteerejä ja toiminnallisen opinnäytetyön prosessia. Lisäksi esittelemme ohjeistuksemme kudokset ja sen, miten niiden dissekoinnin voi laboratoriossa suorittaa. Työohjeissa (5kpl) ohjeistamme pienten ihonäytteiden, sappirakon, umpilisäkkeen, vasektomianäytteen sekä paksu- ja peräsuolen polyypin oikeaoppisen dissekoinnin. Ohjeistuksessa on kirjallisen ilmaisun tukena käytetty kuvia (piirroksia ja valokuvia). Työohjeet edesauttavat bioanalytiikan opiskelijan histologian oppimista ja valmistavat työelämään. Ohjeita voi hyödyntää histologian opintojaksoilla ja työelämäharjoittelussa. Opettajat voivat käyttää ohjeistusta opetusmateriaalina.Part of the studies in the Degree Programme of Biomedical Laboratory Science studies is histology. Histology is the study of microscopic structure of cells and tissues. One part of biomedical laboratory scientists’ job at the clinical pathology laboratory is to dissect small tissue samples. The purpose of dissection is to obtain a representative sample of tissue for a microscopic examination performed by a pathologist. The purpose of this study was to prepare working instructions on dissecting small histological samples for Biomedical Laboratory Science students at Tampere University of Applied Sciences. The tissues for the instructions have been selected with the assistance of clinical pathology employees of Fimlab Laboratories. These tissues represent the most common small tissue samples in clinical pathology laboratory. The aim of this study was to create clear, simple and userfriendly study material for Biomedical Laboratory Science students to use in their professional studies. This is practice-based thesis that consists of a report and five dissection instructions. The instructions are for small skin samples, gall bladder, vermiform appendix, vasectomy samples and polyps of the colon and rectum. Instruction consist of instructions, photos and illustrations. These instructions can be used as teaching material in histology courses. Working instructions help students in their histology studies and prepare them for working life

Characteristics of the study population.

a<p>BMI data available on 101 patients.</p

Plasma cell free DNA (cf-DNA) values in patients with bacteremia stratified by various demographic features, underlying conditions and causative organism.

a<p>BMI data available on 101 patients,</p>b<p>smoking data available on 120 patients,</p>c<p>ultimately or rapidly fatal disease,</p>d<p>value indicates the difference in cf-DNA values between groups of patients stratified by different causative organism.</p

Spearman′s test correlation between cf-DNA level and C-reactive protein (CRP), soluble Fas (sFas), Fas ligand (FasL), kynurenine to tryptophan ratio (kyn/trp) and visually graded band intensity in gel electrophoresis.

<p>Acute phase values (during days 1 to 4 after blood culture) were studied.</p

cf-DNA ROC curve.

<p>Receiver operating characteristic (ROC) curve for maximal plasma cell free DNA (cf-DNA) concentration detected on days 1–4 after positive blood culture in relation to case fatality in bacteremia patients.</p

cf-DNA and survival curve.

<p>Cumulative 30-d survival in bacteremia patients with maximum plasma cell free DNA (cf-DNA) >1.52 ug/ml compared to those with cf-DNA ≤1.52 ug/ml. The cut off value of 1.52 ug/ml was chosen using ROC curve. The survival curve was calculated using the Kaplan-Meier method, and survival differences between groups were compared by log-rank test.</p

Clinical disease severity of patients stratified by maximum plasma cell free DNA (cf-DNA) value (1 to 4 days after blood culture).

a<p>intensive care unit,</p>b<p>sequential organ failure assessment,</p>c<p>mean arterial pressure, $continuous variable (OR and CI cannot be applied).</p