Plasma free DNA: Evaluation of temperature-associated storage effects observed for Roche Cell-Free DNA collection tubes

Introduction: Standardized pre-analytical blood sample procedures for the analysis of circulating cell-free DNA (ccfDNA) are still not available. Therefore, the present study aimed at evaluating the impact of storage conditions related to different times (24 and 48 h) and temperatures (room temperature (RT) and 4 - 8 °C) on the plasma ccfDNA concentration of blood samples drawn into Cell-Free DNA collection tubes (Roche Diagnostics GmbH, Mannheim, Germany). Materials and methods: Venous blood from 30 healthy individuals was collected into five 8.5 mL Cell-Free DNA Collection Tubes (Roche Diagnostics GmbH) each. Plasma samples were processed at time point of blood collection (tube 1), and after storage under the following conditions: 24 h at RT (tube 2) or 4-8 °C (tube 3), and 48 h at RT (tube 4) or 4 - 8 °C (tube 5). Circulating cell-free DNA concentrations were determined by EvaGreen chemistry-based droplet digital PCR (ddPCR). Results: No statistically significant differences between median (interquartile range) plasma ccfDNA concentrations (ng/mL) at time point of blood collection (3.17 (2.13 – 3.76)) and after storage for 24 h (RT: 3.02 (2.41 – 3.68); 4-8 °C: 3.21 (2.19 – 3.46)) and 48 h (RT: 3.13 (2.10 – 3.76); 4-8 °C: 3.09 (2.19 – 3.50)) were observed (P values from 0.102 – 0.975). Conclusions: No unwanted release of genomic DNA from white blood cells could be detected in plasma samples after tube storage for 24 and 48 h regardless of storage temperature

Long time blood-transfusion trend in a European general hospital

Reports about long-time transfusion trends in Austrian hospitals are rare. In our hospital, we implemented an algorithm of preoperative anemia management as part of a patient blood management (PBM) program in October 2011. Anemic individuals with elective surgery underwent an adequate preoperative anemia classification and treatment with erythropoietin and intravenous iron. The aim of this study was to assess red blood cell (RBC), platelet and plasma transfusions before and after implementation of an anemia management program in a general hospital in Austria. This retrospective study evaluated a 12-year trend (2006 – 2017) of RBC, platelet and plasma transfusions in an Austrian general hospital comprising a 6-year period before (2006 – 2011) and a 6-year period after (2012 – 2017) the implementation of an algorithm-guided anemia management. From overall 49,142 transfused RBC units between 2006 - 2017, 22,745 units were transfused in the post-implementation period compared to 26,397 units before PBM initiation (-13.8 %). The plasma unit use decreased also distinctly (787 vs. 1065 units, - 26.1 %) in the period after PBM implementation, whereas a slight decrease of platelet concentration use (807 vs. 843 units, - 4.3 %) was observed, only. This study demonstrates a 12-year pattern of blood use in an Austrian hospital with a distinct decreasing trend of transfused RBC and plasma units during this period. The implementation of PBM activities decreased the need of blood utilization at our institution. Further initiatives are needed to continue this trend in the next years

Method evaluation study of a new generation of vitamin D assays

Introduction: Recently several diagnostic manufacturers have launched new 25-hydroxy-vitamin D (25[OH]D) assays, which are aligned to the National Institute of Standards and Technology (NIST) Standard Reference Materials (SRM) (NIST, Gaithersburg, Maryland). The aim of this study was to compare the performance of one liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, one enzyme linked immunosorbent assay (ELISA), and one recalibrated and previous version of a chemiluminescence immunoassay (CLIA). Material and methods: Serum-aliquots of 198 patient samples from routine 25(OH)D analysis were measured by the ClinMass® LC-MS/MS Complete Kit (RECIPE Chemicals + Instruments GmbH, Munich, Germany), the ORGENTEC 25(OH)D3/D2 ELISA (ORGENTEC Diagnostika GmbH, Mainz, Germany), the recalibrated Immunodiagnostic Systems (IDS)-iSYS 25(OH)DS and the previous used IDS-iSYS 25(OH)D CLIA (Immunodiagnostic Systems Ltd, Boldon, United Kingdom). Bland-Altman and Deming regression analyses were calculated for methods comparison of all tested 25(OH)D assays. The LC-MS/MS method was defined as the reference method. Within-run and between-run precision measurements were performed for all methods with three different concentration levels. Results: Compared to the LC-MS/MS method, the new IDS-iSYS 25(OH)DS and ORGENTEC 25(OH)D3/D2 assay demonstrated mean relative biases of 16.3% and 17.8%. The IDS-iSYS 25(OH)D assay showed the lowest mean bias of 1.5%. Deming regression analyses of the recalibrated IDS-iSYS 25(OH)DS and the ORGENTEC 25(OH)D3/D2 assay showed proportional differences, when compared to the reference method. All assays showed a within-run and between-run imprecision of ≤ 20% at each of the evaluated concentration levels. Conclusions: The evaluated standardized immunoassays and LC-MS/MS are useful methods for measuring 25(OH)D serum-levels in clinical laboratories

CONGENITAL CYTOMEGALOVIRUS INFECTION

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Lactose Malabsorption Testing in Daily Clinical Practice: A Critical Retrospective Analysis and Comparison of the Hydrogen/Methane Breath Test and Genetic Test ( C

The aim of this study was to establish a retrospective evaluation and comparison of the hydrogen/methane (H2/CH4) breath test and genetic test (C/T−13910 polymorphism) results in lactose malabsorption testing. In total 263 consecutive patients with suspected lactose malabsorption were included in this study. They underwent the H2/CH4 breath test following the ingestion of 50 g lactose and were tested for the C/T−13910 polymorphism. In total 51 patients (19.4%) had a C/C−13910 genotype, indicating primary lactose malabsorption. Only 19 patients (7.2%) also had a positive H2/CH4 breath test. All in all 136 patients (51.69%) had a C/T−13910 and 76 patients (28.91%) a T/T−13910 genotype, indicating lactase persistence. Four patients (1.5%) with the C/T−13910 genotype and one patient (0.4%) with the T/T−13910 genotype had a positive H2/CH4 breath test result, indicating secondary lactose malabsorption. Cohen's Kappa measuring agreement between the two methods was 0.44. Twenty patients (7.6%) with a positive H2/CH4 peak within 60 minutes after lactose ingestion were classified as patients with lactose-dependent small intestinal bacterial overgrowth (SIBO). In conclusion, only moderate agreement between the breath test and the genetic test was shown. Secondary lactose malabsorption as well as preanalytical limitations of the combined H2/CH4 breath test procedure can cause discrepant results. This trial is registered with K-42-13

Refining small intestinal bacterial overgrowth diagnosis by means of carbohydrate specificity: a proof-of-concept study

Background: Diagnosis of small intestinal bacterial overgrowth (SIBO) remains challenging. This study aimed at proving the diagnostic concept of carbohydrate-specific SIBO (cs-SIBO) using glucose, fructose and sorbitol hydrogen (H 2 )/methane (CH 4 ) breath testing (HMBT). Methods: In this study 125 patients referred to our outpatient clinic for SIBO testing were included. All individuals underwent glucose (50 g), fructose (25 g) and sorbitol (12.5 g) HMBT at 3 consecutive days. Patients with cs-SIBO (i.e. early H 2 /CH 4 peak) were given rifaximin (600 mg/day) in a 10-day treatment. HMBT results were reassessed in a subset of patients 3–6 months after antibiotic therapy. In view of cs-SIBO diagnosis, agreements between HMBT results obtained for different sugars were calculated using Cohen’s kappa (κ) with 95% confidence intervals (CIs). Results: A total of 59 (47.2%) patients presented an early H 2 /CH 4 peak with one or more sugars. Among these, 21 (16.8%), 10 (8.0%) and 7 (5.6%) individuals had a positive HMBT result with either glucose, fructose or sorbitol, respectively. Another 21 (16.8%) patients with a positive glucose HMBT result were also found positive with an early H 2 /CH 4 peak obtained after ingestion of fructose and/or sorbitol. Fair agreement was observed between glucose and fructose (κ = 0.26, p = 0.0018) and between glucose and sorbitol (κ = 0.18, p = 0.0178) HMBT results. Slight agreement was observed between fructose and sorbitol (κ = 0.03, p = 0.6955) HMBT results only. Successful antibiotic therapy with rifaximin could be demonstrated in 26/30 (86.7%) of patients as indicated by normal HMBT results and symptom remission. Conclusions: Combined glucose, fructose and sorbitol HMBT has the potential to optimize cs-SIBO diagnosis. Furthermore, the majority of patients with cs-SIBO seem to benefit from rifaximin therapy regardless of its carbohydrate specificity

Clinical Study The Impact of an Algorithm-Guided Management of Preoperative Anemia in Perioperative Hemoglobin Level and Transfusion of Major Orthopedic Surgery Patients

The aim of this study was to evaluate a laboratory-guided therapeutic algorithm of preoperative anemia. 335 patients with elective hip or knee arthroplasty were included in this retrospective before-after study. Group I ( = 101) underwent conventional preoperative procedures before algorithm implementation. Group II ( = 234) underwent algorithm-guided preoperative anemia management. A hemoglobin-level of 13 g/dL was the therapeutic cut-off for men and women. Reticulocyte hemoglobin content (CHr) and soluble transferrin receptor (sTfR)/log ferritin ratio were used in the form of the Thomas plot. Iron deficiency (ID) was substituted with 1000 mg iron intravenous (i.v.) and 10000 international units (I.U.) of erythropoiesis-stimulating agent (ESA) subcutaneous (s.c.) or i.v., anemia of chronic disease (ACD) (without functional ID) with 40000 I.U. ESA s.c. or i.v and additionally 200 mg iron i.v. Substituted anemic patients in Group II ( = 32) showed a distinctly higher preoperative (Hb-median 13 versus 11.95 g/dL) ( < 0.01) and postoperative (Hb-median 9.75 versus 9.0 g/dL) ( < 0.05) Hb level compared with untreated anemic patients in Group I ( = 24). In Group II red blood cell (RBC) units (35 units/234 patients) were reduced by 44% compared with Group I (27 units/101 patients). Algorithm-guided preoperative anemia management raises perioperative Hb-level and reduces blood use

Association between increased plasma levels of homocysteine and depression observed in individuals with primary lactose malabsorption.

BACKGROUND:Current literature proposes associations between homocysteine (HCY), folic acid (FA), vitamin B12 metabolism and depression. However, the exact underlying biological mechanisms remain unclear. This study aimed at evaluating a possible link between primary adult-type lactose malabsorption (PALM), HCY, FA and vitamin B12 metabolism and depressive disorder. METHODS:Plasma levels of HCY, FA and vitamin B12 were determined in 78 patients with PALM and 160 individuals with lactase persistence sub-grouped by the presence or absence of major depression. RESULTS:In 78 patients with PALM, the subgroup of 22 individuals with major depression showed significantly higher median (interquartile range) HCY (10.10 [8.46-12.03] vs. 8.9 [7.54-9.86] μmol/L, p = 0.029) and lower plasma FA levels (5.7 [4.68-9.14] vs. 6.95 [5.24-10.56] μmol/L, p = 0.272) compared to the subgroup of 56 individuals without depression, respectively. No such associations could be observed for those 160 individuals without PALM (i.e., lactase persistence) Plasma HCY levels were positively correlated with depressive symptoms (p = 0.052), and showed negative correlations with FA (p = < 0.001) and vitamin B12 (p = 0.029), respectively. CONCLUSION:Depressed individuals with PALM were found with significantly higher HCY and lower FA levels compared to non-depressed individuals with PALM, however, this association was absent in the subgroup of lactase persistent individuals. These findings suggest an association between increased HCY levels, lactose malabsorption and depression