A simple genotyping procedure without DNA extraction to identify rare blood donors.

BACKGROUND: Transfusion-induced alloimmunization has severe clinical consequences including haemolytic transfusion reactions, impaired transfused RBCs longevity and greater difficulty in finding compatible blood. Molecular analysis of genomic DNA now permits prediction of blood group phenotypes based on identification of single nucleotide polymorphisms. Implementation of molecular technologies in donor centres would be helpful in finding RBC units for special patient populations, but DNA extraction remains an obstacle to donor genotyping. MATERIALS AND METHODS: We propose a simple method compatible with high throughput that allows blood group genotyping using a multiplex commercial kit without the need for DNA extraction. The principle relies on pre-PCR treatment of whole blood using heating/cooling procedure in association with a recombinant hotstart polymerase. RESULTS: In a prospective analysis, we yielded 5628 alleles identification and designated 63 donors with rare blood, that is either negative for a high-frequency antigen or with a rare combination of common antigens. CONCLUSION: The procedure was optimized for simplicity of use in genotyping platform and would allow not only to supply antigen-matched products to recipients but also to find rare phenotypes. This methodology could also be useful for establishing a donor repository for human platelet antigens (HPA)-matched platelets since the same issues are involved for patients with neonatal alloimmune thrombocytopenia or post-transfusion purpura.

Candidate gene acting as a suppressor of the RH locus in most cases of Rh-deficiency.

The Rh antigen is a multi-subunit complex composed of Rh polypeptides and associated glycoproteins (Rh50, CD47, LW and glycophorin B); these interact in the red cell membrane and are lacking or severely reduced in Rhnull cells. As a result, individuals with Rhnull suffer chronic haemolytic anaemia known as the Rh-deficiency syndrome. Most frequently, Rhnull phenotypes are caused by homozygosity of an autosomal suppressor gene unlinked to the RH locus (Rhnull regulator or Rhmod types). We have analysed the genes and transcripts encoding Rh, CD47 and Rh50 proteins in five such unrelated Rhnull cases. In all patients, we identified alteration of Rh50--frameshift, nucleotide mutations, or failure of amplification--which correlated with Rhnull phenotype. We propose that mutant alleles of Rh50, which map to chromosome 6p11-21.1, are likely candidates for suppressors of the RH locus accounting for most cases of Rh-deficiency.

Molecular characterization of a new D- - haplotype in a Comorian man.

The D- - phenotype is a genetic variant of the Rh blood group system. It expresses D antigen but lacks C, c, E and e antigens. In D- - phenotype, the RHCE coding region is extensively modified by RHD sequence replacement, nucleotide deletion or splice-site changes. This article reports the identification of a new D- - haplotype in a Comorian man. It exhibits a hybrid gene in which RHCE gene exons 3-8 have been replaced by RHD sequences on the RHCE * C allele background. This allele is associated with no expression of c/C and e/E antigens and overexpression of RhD antigen.

Molecular background of D-negative phenotype in the Tunisian population.

BACKGROUND: Most studies of the molecular basis of Rhesus D-negative phenotype have been conducted in Caucasian and African populations. A comprehensive survey of RHD alleles was lacking in people from North Africa (Tunisians, Moroccans and Algerians) which could be very efficient for managing donors and patients carrying an RHD molecular variant. We analyse the molecular background of D-negative population in Tunisia in the present study. MATERIALS AND METHODS: Blood samples were collected from native Tunisians. A total of 448 D-negative donors from different regions of Tunisia were analysed by RHD genotyping according to an adopted strategy using real-time PCR, ASP-PCR and sequencing. RESULTS: Among the 448 D-negative samples, 443 were phenotyped unequivocally as true D-negative including three molecular backgrounds which were RHD gene deletion (n = 437), RHDψ pseudogene (n = 2) and RHD-CE-D hybrid gene (n = 4) with the respective frequencies of 0·9900, 0·0023 and 0·0046. The remaining five samples, in discordance with the serological results, were identified as two weak D type 11, one weak D type 29, one weak D type 4·0 and one DBT-1 partial D. CONCLUSION: This study showed that the Tunisian population gets closer to Caucasians, given that the RHD gene deletion is the most prevalent cause of D-negative phenotype, but it is slightly different by the presence of the RHDψ pseudogene which was found with a very low frequency compared with that described in the African population. Nevertheless, the relative occurrence of weak D variants among studied serologically D-negative samples make necessary the adaptation of RHD genotyping strategy to the spectrum of prevalent alleles.

Molecular characterization of the human macrophage mannose receptor: demonstration of multiple carbohydrate recognition-like domains and phagocytosis of yeasts in Cos-1 cells.

The macrophage mannose receptor is an integral membrane protein expressed on the surface of tissue macrophages. After ligation of mannose-rich glycoconjugates or pathogens, the receptor mediates endocytosis and phagocytosis of the bound ligands by macrophages. The cDNA-derived primary structure of the mannose receptor predicts a cysteine-rich NH2-terminal domain, followed by a fibronectin type II region. The remainder of the ectodomain is comprised of eight carbohydrate recognition-like domains, followed by a transmembrane region, and a cytoplasmic tail. Transfection of the mannose receptor cDNA into Cos-I cells is necessary for receptor-mediated endocytosis of mannose-rich glycoconjugate as well as phagocytosis of yeasts. Deletion of the cytoplasmic tail results in a mutant receptor that is able to bind but not ingest the ligated pathogens, suggesting that the signal for phagocytosis is contained in the cytoplasmic tail.

Respiratory-gated positron emission tomography and breath-hold computed tomography coupling to reduce the influence of respiratory motion: methodology and feasibility.

BACKGROUND: Respiratory motion causes uptake in positron emission tomography (PET) images of chest and abdominal structures to be blurred and reduced in intensity. PURPOSE: To compare two respiratory-gated PET binning methods (based on frequency and amplitude analyses of the respiratory signal) and to propose a "BH-based" method based on an additional breath-hold computed tomography (CT) acquisition. MATERIAL AND METHODS: Respiratory-gated PET consists in list-mode (LM) acquisition with simultaneous respiratory signal recording. A phantom study featured rectilinear movement of a 0.5-ml sphere filled with (18)F-fluorodeoxyglucose ((18)F-FDG) solution, placed in a radioactive background (sphere-to-background contrast 6:1). Two patients were also examined. Three figures of merit were calculated: the target-to-background ratio profile (TBRP) in the axial direction through the uptake (i.e., the sphere or lesion), full-width-at-half-maximum (FWHM) values, and maximized standard uptake values (SUV(max)). RESULTS: In the phantom study, the peak TBRP was 0.9 for non-gated volume, 1.83 for BH-based volume, and varied between 1.13 and 1.73 for Freq-based volumes and between 1.34 and 1.66 for Amp-based volumes. A reference volume (REF-static) was also acquired for the phantom (in a static, "expiratory" state), with a peak TBRP at 1.88. TBRPs were computed for patient data, with higher peak values for all gated volumes than for non-gated volumes. CONCLUSION: Respiratory-gated PET acquisition reduces the blurring effect and increases image contrast. However, Freq-based and Amp-based volumes are still influenced by inappropriate attenuation correction and misregistration of mobile lesions on CT images. The proposed BH-based method both reduces motion artifacts and improves PET-CT registration.

[Evolution of the early respiratory function after lung resection for cancer]. / Evolution de la fonction respiratoire postopératoire immédiate après résection pulmonaire pour cancer.

The assessment of the postoperative risk in lung resection is a major challenge for pneumologists and thoracic surgeons. Restrictive syndromes have been observed along with a disproportionate decrease of FEV1 in lobectomies. The purpose of the present study is to describe the early response of pulmonary function after thoracotomy and resection for lung cancer. In a prospective study, the authors included 31 patients (19 lobectomy patients: mean age 59+/-10 years and 12 pneumonectomy patients: mean age 56+/-9 years) without postoperative complications. Pulmonary function tests were performed before and after surgery on Days 1, 5 (D5), 10 and within the fourth month. The main aspect of the ventilation was an unexpected similarity in subgroups during the early perioperative period up to D5. When compared with the preoperative value, about a 50% decrease in the vital capacity and total lung capacity was observed. In both subgroups about a 40% decrease was noted in the inspiratory and expiratory reserve volume. In the lobectomy sub-group, the change in the forced expiratory volume in one second over forced vital capacity (FEV/FVC) ratio was found to be higher than predicted (52+/-16% at D5 versus 67+/-14% predicted). However, the FEV/FVC ratio did not change, attesting to major restrictive ventilation. Partial recovery of the FEV was dependant on the mobile volume and especially the inspiratory volume. These findings should have implications in patient management.

Correction to: Functional outcomes in adult patients with herpes simplex encephalitis admitted to the ICU: a multicenter cohort study.

The original article unfortunately contained a mistake. Due to technical problems the study group was not tagged correctly. Please find the correct tagging down below. We apologize for the mistake.

Functional outcomes in adult patients with herpes simplex encephalitis admitted to the ICU: a multicenter cohort study.

PURPOSE: We aimed to study the association of body temperature and other admission factors with outcomes of herpes simplex encephalitis (HSE) adult patients requiring ICU admission. METHODS: We conducted a retrospective multicenter study on patients diagnosed with HSE in 47 ICUs in France, between 2007 and 2017. Fever was defined as a body temperature higher or equal to 38.3 °C. Multivariate logistic regression analysis was used to identify factors associated with poor outcome at 90 days, defined by a score of 3-6 (indicating moderate-to-severe disability or death) on the modified Rankin scale. RESULTS: Overall, 259 patients with a score on the Glasgow coma scale of 9 (6-12) and a body temperature of 38.7 (38.1-39.2) °C at admission were studied. At 90 days, 185 (71%) patients had a poor outcome, including 44 (17%) deaths. After adjusting for age, fever (OR = 2.21; 95% CI 1.18-4.16), mechanical ventilation (OR = 2.21; 95% CI 1.21-4.03), and MRI brain lesions > 3 lobes (OR = 3.04; 95% CI 1.35-6.81) were independently associated with poor outcome. By contrast, a direct ICU admission, as compared to initial admission to the hospital wards (i.e., indirect ICU admission), was protective (OR = 0.52; 95% CI 0.28-0.95). Sensitivity analyses performed after adjustment for functional status before admission and reason for ICU admission yielded similar results. CONCLUSIONS: In HSE adult patients requiring ICU admission, several admission factors are associated with an increased risk of poor functional outcome. The identification of potentially modifiable factors, namely, elevated admission body temperature and indirect ICU admission, provides an opportunity for testing further intervention strategies.

Results from a four-year study on the prevalence of nosocomial infections in Franche-Comté: attempt to rank the risk of nosocomial infection.

The aim of this study was to rank the risk of nosocomial infection (NI) according to patient type by analysing the results of annual prevalence studies carried out in Franche-Comté from 2001 to 2004. Patients (N=14,905) were divided into four categories according to the number of endogenous risk factors (age, immunodepression, MacCabe score). The overall prevalence of infection was 6.1% and varied according to the category of patient from 1.93% (no risk factors) to 15.2% (three risk factors). The frequencies of NI related to an invasive procedure and to cross-contamination with multi-drug-resistant (MDR) bacteria were 30.9% and 12.3%, respectively; these percentages did not depend on the type of patient. The prevalence of NI decreased over time for patients with two or three risk factors, but was stable for patients with no risk factors. More than 40% of NIs were potentially avoidable (related to invasive procedures or involving cross-transmission of an MDR bacterium) regardless of the category of patient. This study suggests that at least 30% of NIs could be avoided.

[Influenza vaccination among health-care workers. Vaccination rates in universitary hospital of Besançon, winter 2003-2004]. / La vaccination antigrippale du personnel hospitalier. Enquête de prévalence au CHU de Besançon, hiver 2003-2004.

BACKGROUND: Influenza vaccination reduces the mortality of the patients when the vaccination rates of healthcare workers is important. PURPOSE AND METHOD OF THE STUDY: To investigate the vaccination rates at the Universitary Hospital of Besançon by anonymous questionnaire. RESULTS: Three thousand hundred seventy-seven answers were analyzed (228 men and 1145 women). Two hundred seventy-seven persons declared receiving the vaccine (20.1%) corresponding to sixty-three men (27.6%) and two hundred thirteen women (18.6%) (P = 0.001). The average age of the healthcare workers vaccined was of 38.9+/-11 years. Among most than 50 years, 34% was vaccined. Among the doctors, 40.5% were vaccined against 20.6% of the nurses. In the services of geriatrics, 78.5% of the staff was vaccined. CONCLUSIONS: Our results indicate a weak rate of influenza vaccination in our establishment and a misunderstanding of the character nosocomial of the influenza among the nurse.

Prevalence of aortic valve dystrophy and insufficiency in a cohort of 255 HIV-positive patients followed-up in a cardiology department between 2012 and 2014.

OBJECTIVES: To study valve appearance and the presence of valve disease in a cohort of people living with HIV (PLHIV). DESIGN: A prospective study of PLHIV examined at the cardiology department of the Clermont Ferrand university hospital group (CHU) between January 1, 2012, and December 31, 2014. Were excluded those with a history of infection associated with a possible endocarditis. METHODS: Demographic, medical characteristics and cardiovascular disease risk factors at time of cardiovascular examination and Doppler-echocardiography were recorded and analyzed. RESULTS: In total, 903 PLHIV were examined in the infectious diseases department, 255 of whom were included. These consisted of 67 women (26.3%) and 188 men, of a mean age of 51.2±9.7years, in whom coronary artery disease was diagnosed in 18 patients (7.0%), two women and 16 men, representing a prevalence of 3.0% in females and 8.5% in males. The appearance of the aortic cusps was considered dystrophic in 14.1% of cases (36/255), dysplastic in two cases (0.8%), exhibiting a bicuspid deformity in one case. The prevalence of aortic valve abnormality was therefore 6.0% in the women (4/67) and 17.0% in the men (32/188). On facing off this data with the Kora Monica study findings, an increase in prevalence appears only to truly manifest after 50years of age. We registered 35 aortic insufficiency cases (13.7%), representing a higher incidence than that of the Framingham cohort, with age and masculine gender being the determining factors. CONCLUSION: Valve disease, along with coronary artery disease, should be closely monitored in PLHIV.

Cryptogenic mechanism in ischaemic stroke patients is a predictor of 5-year survival: A population-based study.

INTRODUCTION: The present study sought to identify factors affecting mortality beyond 28 days in ischaemic stroke patients with whatever ischaemic mechanism. PATIENTS AND METHODS: A prospective population-based registry was set up in Brest County, Brittany, France. Demographic data, clinical presentation, vascular risk factors and mortality were collected from January 2008 to December 2012. At "home without help" was used as a surrogate marker for low Rankin (0-1) at discharge from the hospital. IS was classified on the TOAST classification. Overall mortality was calculated using the Kaplan-Meier method. Multivariate analysis of mortality beyond 28 days was implemented, using a Cox model, on significant risk factors identified on univariate analysis. RESULTS: About 3024 IS cases were followed up beyond 28 days. Overall mortality beyond 28 days was 38.49% at 60 months. On multivariate analysis, age (10 years: HR = 1.84; [1.66-2.02]), coronary artery disease (HR = 1.28; [1.05-1.56]), cardiac arrhythmia (HR = 1.36; [1.11-1.67]), peripheral artery disease (HR = 1.66 [1.29-2.13]) and incomplete assessment (HR = 1.39; [1.12-1.74]) were associated with higher mortality risk, whereas female gender (HR = 0.80; [0.68-0.94]), high Glasgow Coma Scale score (GCS > 12) (HR = 0.58; [0.45-0.76]), lacunar syndrome (HR = 0.82; [0.68-0.99], being 'at home without help' (HR = 0.50; [0.41-0.59]) and negative assessment (HR = 0.75; [0.58-0.97], compared to cardioembolism) were associated with better survival probability. DISCUSSION: Initial clinical status, prior cardiovascular diseases and age was associated with more risk of death: an increment of 10 years almost doubled mortality. Women had more survival probability than men, controlling for age. Ischaemic stroke mechanisms were predictors of late 5-year mortality. CONCLUSION: Patients with negative assessment, i.e. representing truly cryptogenic ischaemic stroke, had the best survival probability probably due to fewer atherosclerotic markers.

Molecular and functional characterization of an amphibian urea transporter.

We report the characterization of a frog (Rana esculenta) urea transporter (fUT). The cloned cDNA is 1.4 kb long and contains a putative open reading frame of 1203 bp. In frog urinary bladder, the gene is expressed as two mRNAs of 4.3 and 1.6 kb. The fUT protein is 63.1 and 56.3% identical to rat UT-A2 and UT-B1, respectively. The internal duplication of UT-A2 and UT-B, as well as the double LP box urea transporter signature sequence were found in this amphibian urea transporter. When expressed in Xenopus oocytes, fUT induced a 10-fold increase in urea permeability, which was blocked by both phloretin and mercurial reagents. The fUT protein did not transport thiourea, but the fUT-mediated urea transport was strongly inhibited by this compound. Thus, this amphibian urea transporter displays transport characteristics in between those of UT-A2 and UT-B.

Characterization of a murine monoclonal antibody specific for the human P1 blood group antigen.

Four monoclonal antibodies (MAbs) directed against the P1 blood group antigen were produced by hybridomas obtained from mouse immunized with turtle-dove avomucoid. One of the MAb (154 IX B6) selected as a blood typing reagent agglutinated native P1 and Pk1 red cells with a high titer but was inactive against native P2, Pk2 and p erythrocytes. After papain treatment the reactivity towards P1 and Pk1 erythrocytes was enhanced whereas p erythrocytes remained unreactive. A weak cross-reactivity of the MAb with the Pk antigen was suspected since enzyme-treated Pk2 erythrocytes became significantly agglutinated. Further analysis of the antibody specificity was established by binding studies using neutral glycolipids prepared from P1 and P2 erythrocytes, affinity immunoabsorbents carrying known oligosaccharide structures and hapten inhibition with synthetic oligosaccharides. The MAb bound weakly to the Gal alpha 1-4Gal structure common to P1 and Pk antigens but had a marked preference for the P1 determinant (Gal alpha 1-4 Gal beta 1-4 GlcNAc) and the binding was abolished by prior treatment of oligosaccharide antigens by alpha(not beta)-galactosidase, which supports evidence that a terminal alpha-galactose residue is involved in the blood group P1 and Pk specificities. The MAb has a slightly broader specificity than the human anti-P1 counterpart but can be used safely for routine blood typing.

Molecular characterization of a new urea transporter in the human kidney.

A cDNA clone (HUT2) sharing 61.1% and 89.9% sequence identity with the human erythroid (HUT11) and the rabbit (UT2) urea transporters, respectively, was isolated by homology cloning from a human kidney library. HUT2 transcripts were restricted to the kidney and the HUT2 polypeptide was not immunoprecipitated with blood group Kidd-related antibodies (anti-Jk3) in coupled transcription-translation assays. Functional expression studies in Xenopus oocytes demonstrated that HUT2-mediated urea transport was not inhibited by p-chloromercuribenzene sulfonate (pCMBS) which, however, inhibited the urea flux mediated by HUT11. These findings demonstrate that at least two distinct urea transporters are present in human tissues. By in situ hybridization, the gene encoding HUT2 has been assigned to chromosome 18q12.1-q21-1, as found previously for the Kidd/urea transporter HUT11, suggesting that both genes evolved from duplication of a common ancestor.

Intracardiac ectopic thyroid.

The case of a patient with an intracardiac ectopic thyroid is reported. A lesion was found in a 25-year-old man and was diagnosed by two-dimensional echocardiography as a right intraventricular tumor. An operation was performed. Histologic and ultrastructural studies showed that the tumor was a thyroid mass. The origin of intracardiac ectopic thyroids is probably to be found in disturbances occurring early in embryogenesis.

Association of private isolation rooms with ventilator-associated Acinetobacter baumanii pneumonia in a surgical intensive-care unit.

OBJECTIVE: To determine the rates and routes of Acinetobacter baumanii colonization and pneumonia among ventilated patients in a surgical intensive-care unit (SICU) before and after architectural modifications. DESIGN: A nonsequential study comparing two groups of patients. All isolates from systematic and clinical samples were genotyped by pulsed-field gel electrophoresis (PFGE). Records of patients hospitalized during the first and second periods were reviewed and findings were compared. Between the two periods, the SICU was remodeled from enclosed isolation rooms and open rooms to only enclosed isolation rooms with handwashing facilities in each room. SETTING AND PATIENTS: All patients hospitalized and mechanically ventilated for more than 48 hours in the 15-bed SICU of the University Hospital of Besançon (France). RESULTS: For the first and second periods, the rates of colonization were, respectively, 28.1% and 5.0% of patients (P < 10(-7); relative risk [RR], 2.23; 95% confidence interval [CI95], 1.8-2.75) and the specific rates of bronchopulmonary (BP) colonization were, respectively, 9.1 and 0.5 per 1,000 days of mechanical ventilation (P < 10(-5). Seven major PFGE isolate types were identified, 4 of which were isolated from 44 of the 47 colonized or infected patients. Logistic regression analysis showed that colonization was not associated with patient characteristics. CONCLUSION: Conversion from open rooms to isolation rooms may help control nosocomial BP tract acquisition of A baumanii in mechanically ventilated patients hospitalized in an SICU.

Systemic-pulmonary shunt with a right retroesophageal subclavian artery.

A 19-day-old child suffering from cyanosis due to tetralogy of Fallot was palliated by using his right retroesophageal subclavian artery. It was anastomosed side-to-side onto the ascending aorta and end-to-side onto the right pulmonary artery. The palliation obtained with this systemic-pulmonary shunt was satisfying. The right brachial vascular flow was normal.