Mutations in CRLF1 cause familial achalasia.

We here report a family from Libya with three siblings suffering from early onset achalasia born to healthy parents. We analyzed roughly 5000 disease-associated genes by a next-generation sequencing (NGS) approach. In the analyzed sibling we identified two heterozygous variants in CRLF1 (cytokine receptor-like factor 1). Mutations in CRLF1 have been associated with autosomal recessive Crisponi or cold-induced sweating syndrome type 1 (CS/CISS1), which among other symptoms also manifests with early onset feeding difficulties. Segregation analysis revealed compound heterozygosity for all affected siblings, while the unaffected mother carried the c.713dupC (p.Pro239Alafs*91) and the unaffected father carried the c.178T>A (p.Cys60Ser) variant. The c.713dupC variant has already been reported in affected CS/CISS1 patients, the pathogenicity of the c.178T>A variant was unclear. As reported previously for pathogenic CRLF1 variants, cytokine receptor-like factor 1 protein secretion from cells transfected with the c.178T>A variant was severely impaired. From these results we conclude that one should consider a CRLF1-related disorder in early onset achalasia even if other CS/CISS1 related symptoms are missing.

[Diagnostics and Eradication Therapy for MRSA Carriers in the Outpatient Sector: an Analysis of the Reimbursement Situation in the Light of Current Reimbursement Changes]. / Diagnostik und ambulante Eradikationstherapie von Trägern mit MRSA in der vertragsärztlichen Versorgung ­ Eine Analyse der Erstattungssituation im Kontext aktueller Vergütungsveränderungen.

Infection with methicillin-resistant Staphylococcus aureus (MRSA) occurs in both the inpatient and outpatient sector. The reimbursement for diagnostic services and eradication therapy in the outpatient sector was regulated for the first time on 01.04.2012 and after a 2-year test period, has been adopted into the standard range of care services. The aim of this retrospective study was to give an overview of the current situation in services and reimbursement in Germany and describe MRSA patients and their treatment in the outpatient sector. Secondary data, namely reimbursement data of the National Association of Statutory Health Insurance Physicians (KBV) und the Physicians' Association (KV) Mecklenburg-West Pomerania for the period 01/04/2012-31/03/2014 were analyzed. Results show that on the federal level, MRSA services amounting to € 3,235,870.18 have been reimbursed and that diagnostic costs exceed treatment costs. In Germany, 5,627 doctors invoiced services related to MRSA; 51,56% of these were general practitioners and 21,25% specialists in internal medicine working in general practice. In the KV Mecklenburg-Western Pomerania, patients were elderly (average age 69,13), cost for services were on average 27,76 €, and 76,85% of the patients were treated within one quarter. On the whole, there were regional differences in the identification and eradication of MRSA in the outpatient setting. In order to provide an extended base for a more efficient resource allocation in the health care sector, in addition to analysis of MRSA eradication from the medical point of view, attention needs to be paid to patient flow between the out- and inpatient sectors, as well as economic aspects.

[Additional Costs for Care of Patients with Multi-Resistant Pathogens--An Analysis from the Perspective of a Statutory Health Insurance]. / Mehrkosten bei der Versorgung von Patienten mit multiresistenten Erregern--Eine Analyse aus Sicht einer gesetzlichen Krankenversicherung.

Aim of this study was to determine the additional expenditures for a German statutory health insurance which are induced by patients with multi-resistant bacteria. Therefore a nationwide cross-sectional data analysis using routine data of the health insurance "Techniker Krankenkasse" was conducted. In the consideration of costs we included expenditures for inpatient and outpatient care and on drugs in a time period of 12 months. A control group was matched by age, gender, basic disease, quarterly period and region. On average additional costs of 17,500 Euro per insured were calculated due to the presence of multi-resistant bacteria. The hypothesis was corroborated in that the level of these costs differ widely by age, gender and basic disease.

Analysis of MRSA-attributed costs of hospitalized patients in Germany.

Infections with methicillin-resistant Staphylococcus aureus (MRSA) are assumed to have a high economic impact due to increased hygienic measures and prolonged hospital length of stay. However, surveys on the real expenditure for the prevention and treatment of MRSA are scarce, in particular with regard to the German Diagnosis-Related Groups (G-DRG) payment system. The aim of our study is to empirically assess the additional cost for MRSA management measures and to identify the main cost drivers in the whole process from the hospital's point of view. We conducted a one-year retrospective analysis of MRSA-positive cases in a German university hospital and determined the cost of hygienic measures, laboratory costs, and opportunity costs due to isolation time and extended lengths of stay. A total of 182 cases were included in the analysis. The mean length of hospital stay was 22.75 days and the mean time in isolation was 17.08 days, respectively. Overall, the calculated MRSA-attributable costs were 8,673.04 per case, with opportunity costs making up, by far, the largest share (77.45 %). Our study provides a detailed up-to-date analysis of MRSA-attributed costs in a hospital. It allows a current comparison to previous studies worldwide. Moreover, it offers the prerequisites to investigate the adequate reimbursement of MRSA burden in the DRG payment system and to assess the efficiency of targeted hygienic measures in the prevention of MRSA.

Mutations in the gene encoding immunoglobulin mu-binding protein 2 cause spinal muscular atrophy with respiratory distress type 1.

Classic spinal muscular atrophy (SMA) is caused by mutations in the telomeric copy of SMN1. Its product is involved in various cellular processes, including cytoplasmic assembly of spliceosomal small nuclear ribonucleoproteins, pre-mRNA processing and activation of transcription. Spinal muscular atrophy with respiratory distress (SMARD) is clinically and genetically distinct from SMA. Here we demonstrate that SMARD type 1 (SMARD1) results from mutations in the gene encoding immunoglobulin micro-binding protein 2 (IGHMBP2; on chromosome 11q13.2-q13.4). In six SMARD1 families, we detected three recessive missense mutations (exons 5, 11 and 12), two nonsense mutations (exons 2 and 5), one frameshift deletion (exon 5) and one splice donor-site mutation (intron 13). Mutations in mouse Ighmbp2 (ref. 14) have been shown to be responsible for spinal muscular atrophy in the neuromuscular degeneration (nmd) mouse, whose phenotype resembles the SMARD1 phenotype. Like the SMN1 product, IGHMBP2 colocalizes with the RNA-processing machinery in both the cytoplasm and the nucleus. Our results show that IGHMBP2 is the second gene found to be defective in spinal muscular atrophy, and indicate that IGHMBP2 and SMN share common functions important for motor neuron maintenance and integrity in mammals.

Mutations in the skeletal muscle alpha-actin gene in patients with actin myopathy and nemaline myopathy.

Muscle contraction results from the force generated between the thin filament protein actin and the thick filament protein myosin, which causes the thick and thin muscle filaments to slide past each other. There are skeletal muscle, cardiac muscle, smooth muscle and non-muscle isoforms of both actin and myosin. Inherited diseases in humans have been associated with defects in cardiac actin (dilated cardiomyopathy and hypertrophic cardiomyopathy), cardiac myosin (hypertrophic cardiomyopathy) and non-muscle myosin (deafness). Here we report that mutations in the human skeletal muscle alpha-actin gene (ACTA1) are associated with two different muscle diseases, 'congenital myopathy with excess of thin myofilaments' (actin myopathy) and nemaline myopathy. Both diseases are characterized by structural abnormalities of the muscle fibres and variable degrees of muscle weakness. We have identified 15 different missense mutations resulting in 14 different amino acid changes. The missense mutations in ACTA1 are distributed throughout all six coding exons, and some involve known functional domains of actin. Approximately half of the patients died within their first year, but two female patients have survived into their thirties and have children. We identified dominant mutations in all but 1 of 14 families, with the missense mutations being single and heterozygous. The only family showing dominant inheritance comprised a 33-year-old affected mother and her two affected and two unaffected children. In another family, the clinically unaffected father is a somatic mosaic for the mutation seen in both of his affected children. We identified recessive mutations in one family in which the two affected siblings had heterozygous mutations in two different exons, one paternally and the other maternally inherited. We also identified de novo mutations in seven sporadic probands for which it was possible to analyse parental DNA.

Poly(vinyl alcohol) freeze casts with nano-additives as potential thermal insulators.

Freeze-casting consists of freezing a liquid suspension (aqueous or other), followed by sublimation of the solidified state to the gas state under reduced pressure, and subsequent sintering of the remaining scaffold to consolidate and densify the struts and walls. The structure is very porous with the pores being a replica of the solvent crystals. The technique is rather versatile and the use of a liquid solvent (water most of the time) as a pore forming agent is a strong asset. Freeze-casting has also been developed as a near net shape forming route yielding dense ceramics. In this work we report on porous composite materials synthesized via the ice templating method. Poly(vinyl alcohol) (PVA) is used as matrix and nano-silica (SiO2), nanoclay (NC) and microfibrillated cellulose (MFC) are used as fillers to improve the mechanical stability of the PVA scaffold. We show our results on the porosity and mechanical stability and consider these porous nanocomposites as potential insulation materials with low thermal conductivity and superior mechanical properties.

The AE4 transporter mediates kidney acid-base sensing.

The kidney plays a key role in the correction of systemic acid-base imbalances. Central for this regulation are the intercalated cells in the distal nephron, which secrete acid or base into the urine. How these cells sense acid-base disturbances is a long-standing question. Intercalated cells exclusively express the Na+-dependent Cl-/HCO3- exchanger AE4 (Slc4a9). Here we show that AE4-deficient mice exhibit a major dysregulation of acid-base balance. By combining molecular, imaging, biochemical and integrative approaches, we demonstrate that AE4-deficient mice are unable to sense and appropriately correct metabolic alkalosis and acidosis. Mechanistically, a lack of adaptive base secretion via the Cl-/HCO3- exchanger pendrin (Slc26a4) is the key cellular cause of this derailment. Our findings identify AE4 as an essential part of the renal sensing mechanism for changes in acid-base status.

First HPSE2 missense mutation in urofacial syndrome.

Urofacial syndrome (UFS) describes the combination of urological problems and an inverted facial expression upon attempts to smile. Seventeen independent familial cases from different ethnicities have been described so far. Some of these have been linked to chromosome 10q. Very recently, homozygous loss-of-function mutations affecting the gene HPSE2 were identified in nine cases. Here, we describe a consanguineous UFS family from Pakistan with three of six siblings affected. We establish linkage to the chromosome 10q critical region and identify two non-synonymous HPSE2 variants. In silico analysis and screening of controls defines c.631T>C (p.Y211H) as a novel benign SNP and c.1628A>T (p.N543I) as the disease-causing mutation. Our study exemplifies the challenges in proper clinical diagnosis of UFS and, thereby, supports the hypothesis of the disease being under diagnosed. By identifying the first HPSE2 missense mutation it also provides a starting point for studies aimed at functionally understanding the unusual combination of symptoms as characterizing UFS.

Cost-analysis of PCR-guided pre-emptive antibiotic treatment of Staphylococcus aureus infections: an analytic decision model.

The aim of this study is to examine whether rapid polymerase chain reaction (PCR)-based screening is a cost-efficient tool to optimize pre-emptive antibiotic therapy of methicillin-resistant and methicillin-sensitive Staphylococcus aureus (MRSA and MSSA, respectively) infections. A decision analytic cost model was developed, based on data from the peer-reviewed literature. Sensitivity analyses were undertaken to investigate the impact of variation in the MRSA rate, cost ratio of the cost of inappropriate antibiotic therapy to the cost of appropriate antibiotic therapy, PCR test cost, and total hospital costs per case. At a current MRSA rate of 24.5 % in Germany, PCR-guided treatment regimens are cost-efficient compared to empirical strategies. The costs of alternative treatment strategies differ, on average, up to 1,780 per case. An empirical MRSA treatment strategy is least costly when the cost ratio is less than 1.06. When the total hospital cost per MRSA case is increased, pre-emptive MSSA treatment with PCR tests achieves the lowest average cost. Early verification and adaptation of an initial pre-emptive antibiotic treatment of S. aureus infections using PCR-based tests are advantageous in Germany and other European countries. PCR tests, accordingly, should be considered as elements in antimicrobial stewardship programs.

Transmission rates, screening methods and costs of MRSA--a systematic literature review related to the prevalence in Germany.

Methicillin-resistant Staphylococcus aureus (MRSA) infections represent a serious challenge for health care institutions, which is inherent in the combination of prevalence, transmission rates and costs. Furthermore, performing an MRSA screening requires information on the complex system of effectiveness, accuracy and costs of different screening methods. The purpose of this study was to give an overview of parameters with decisive significance for the burden of MRSA and the selection of a specific MRSA screening strategy. A systematic literature search for peer-reviewed health economic studies associated with MRSA was performed (from 1995 to the present). Eighty-seven different studies met all inclusion and exclusion criteria. Primary outcomes included the prevalence of MRSA, MRSA transmission rates, performance characteristics of MRSA screening methods, costs for pre-emptive isolation precautions and costs per MRSA case. The prevalence rates reported for all inpatients (1.2-5.3 %) as well as for inpatients with risk factors or patients in risk areas (3.85-20.6 %) vary greatly. The range of cross-transmission rates per day reported for patients with MRSA in isolation is 0.00081-0.009 and for carriers not in isolation is 0.00137-0.140, respectively. For polymerase chain reaction (PCR) methods, the mean sensitivity and specificity were 91.09 and 95.79 %, respectively. Culture methods show an average sensitivity of 89.01 % and an average specificity of 93.21 %. The turn-around time for PCR methods averages 15 h, while for the culture method, it can only be estimated as 48-72 h. This review filtered important parameters and cost drivers, and covered them with literature-based averages. These findings serve as an ideal evidence base for further health economic considerations of the cost-effectiveness of different MRSA screening methods.

Protracted course of juvenile ceroid lipofuscinosis associated with a novel CLN3 mutation (p.Y199X).

The juvenile neuronal ceroid lipofuscinosis (JNCL, Batten disease, MIM 204200), is an autosomal recessive lysosomal storage disease, which is characterized by ubiquitous accumulation of the lipopigment material ceroid-lipofuscin. It manifests with loss of vision in childhood due to retinal degeneration, followed by seizures and parkinsonism leading to premature death at around 30 years. Eighty-five percent of JNCL patients carry a disease-causing 1.02 kb deletion in the CLN3 gene on chromosome 16. Here we report on a large consanguineous Lebanese family with five affected siblings. Electron microscopy of lymphocytes revealed the presence of fingerprint profiles suggesting JNCL. However, disease progression, especially of mental and motor function was slower as expected for 'classic' JNCL. We thus confirmed the diagnosis by genetic testing and found a new c.597C>A transversion in exon 8, homozygous in all affected family members and not present in 200 alleles of normal controls. The mutation generates a premature termination codon (p.Y199X) truncating the CLN3 protein by 55%. In heterozygous state mutant mRNA transcripts are expressed at the same levels as the wild-type ones, suggesting the absence of nonsense mediated messenger decay. We discuss a potential residual catalytic function of the truncated protein as a cause for the mild phenotype.

Extracellular matrix contains insulin-like growth factor binding protein-5: potentiation of the effects of IGF-I.

Insulin-like growth factor binding proteins (IGFBPs) have been shown to serve as carrier proteins for the insulin-like growth factors (IGFs) and to modulate their biologic effects. Since extracellular matrix (ECM) has been shown to be a reservoir for IGF-I and IGF-II, we examined the ECM of cultured human fetal fibroblasts and found that IGFBP-5 was incorporated intact into ECM, while mostly inert proteolytic fragments were found in the medium. In contrast, two other forms of IGFBP that are secreted by these cells were either present in ECM in minimal amounts (IGFBP-3) or not detected (IGFBP-4). Likewise, when purified IGFBPs were incubated with ECM, IGFBP-5 bound preferentially. IGFBP-5 was found to bind to types III and IV collagen, laminin, and fibronectin. Increasing salt concentrations inhibited the binding of IGFBP-5 to ECM and accelerated the release of IGFBP-5 from ECM, suggesting an ionic basis for this interaction. ECM-associated IGFBP-5 had a sevenfold decrease in affinity for IGF-I compared to IGFBP-5 in solution. Furthermore, when IGFBP-5 was present in cell culture substrata, it potentiated the growth stimulatory effects of IGF-I on fibroblasts. When IGFBP-5 was present only in the medium, it was degraded to a 22-kD fragment and had no effect on IGF-I-stimulated growth. We conclude that IGFBP-5 is present in fibroblast ECM, where it is protected from degradation and can potentiate the biologic actions of IGF-I. These findings provide a molecular explanation for the association of the IGF's with the extracellular matrix, and suggest that the binding of the IGF's to matrix, via IGFBP-5, may be important in mediating the cellular growth response to these growth factors.

[Economic aspects of the management and control of MRSA]. / Okonomische Aspekte des Hygienemangements von MRSA.

Methicillin resistance of Staphylococcus aureus strains (MRSA) has become a steadily growing cost factor for the German and international health system. Besides direct costs for diagnostics and therapy, indirect and intangible costs are a considerable part in the total expenses. Because of the massive increase of MRSA in medical centres as well as in the population and further worsening of forced rationalizing in the health care systems, the interest in the economic effects of this phenomenon has increased greatly during the last years. For managing MRSA and for effective cost containment, especially in hospitals, infection control measures have not only to prevent the further spread of MRSA but actively reduce the number of colonised patients. In the last years the efficiency of several infection control measures and their combination to prevent the spread of MRSA has been shown. However, these measures are linked to considerable direct costs in the first place. Nevertheless, an overall economic assessment of hygiene measures is only possible after a complete cost-benefit analysis, where costs are compared to effects achieved. Moreover, analyses that focus on effects for single institutions only are as inappropriate as purely monetary analyses or analyses that include the direct additional only. Despite of at first considerable expenditures, not only infection control measures but proactive strategies to reduce the total number of MRSA cases (accordingly of the "Search and Destroy" philosophy) are shown to be cost efficient, even in low prevalence situations, based on examples in literature. Hence, effective programmes to reduce MRSA are not only necessary from a medical and ethical focus but are also cost efficient in medium term for health care providers and the community.

Disruption of KCC2 reveals an essential role of K-Cl cotransport already in early synaptic inhibition.

Synaptic inhibition by GABA(A) and glycine receptors, which are ligand-gated anion channels, depends on the electrochemical potential for chloride. Several potassium-chloride cotransporters can lower the intracellular chloride concentration [Cl(-)](i), including the neuronal isoform KCC2. We show that KCC2 knockout mice died immediately after birth due to severe motor deficits that also abolished respiration. Sciatic nerve recordings revealed abnormal spontaneous electrical activity and altered spinal cord responses to peripheral electrical stimuli. In the spinal cord of wild-type animals, the KCC2 protein was found at inhibitory synapses. Patch-clamp measurements of embryonic day 18.5 spinal cord motoneurons demonstrated an excitatory GABA and glycine action in the absence, but not in the presence, of KCC2, revealing a crucial role of KCC2 for synaptic inhibition.

Mutation and phenotypic spectrum in patients with cardio-facio-cutaneous and Costello syndrome.

Cardio-facio-cutaneous (CFC) and Costello syndrome (CS) are congenital disorders with a significant clinical overlap. The recent discovery of heterozygous mutations in genes encoding components of the RAS-RAF-MAPK pathway in both CFC and CS suggested a similar underlying pathogenesis of these two disorders. While CFC is heterogeneous with mutations in BRAF, MAP2K1, MAP2K2 and KRAS, HRAS alterations are almost exclusively associated with CS. We carried out a comprehensive mutation analysis in 51 CFC-affected patients and 31 individuals with CS. Twelve different BRAF alterations were found in twenty-four patients with CFC (47.0%), two MAP2K1 mutations in five (9.8%) and two MAP2K2 sequence variations in three CFC-affected individuals (5.9%), whereas three patients had a KRAS alteration (5.9%). We identified four different missense mutations of HRAS in twenty-eight cases with CS (90.3%), while KRAS mutations were detected in two infants with a phenotype meeting criteria for CS (6.5%). In 14 informative families, we traced the parental origin of HRAS alterations and demonstrated inheritance of the mutated allele exclusively from the father, further confirming a paternal bias in the parental origin of HRAS mutations in CS. Careful clinical evaluation of patients with BRAF and MAP2K1/2 alterations revealed the presence of slight phenotypic differences regarding craniofacial features in MAP2K1- and MAP2K2-mutation positive individuals, suggesting possible genotype-phenotype correlations.

[Distal spinal-muscular atrophy 1 (DSMA1 or SMARD1)]. / L'amyotrophie spinale distale de type 1 (DSMA1 ou SMARD1).

In this article, we review the clinical, neuropathological and genetic aspects of distal spinal-muscular atrophy 1 (DSMA1; MIM#604320), formerly designated as autosomal recessive spinal muscular atrophy with respiratory distress type 1 (SMARD1) and also known as distal hereditary-motor neuropathy type 6 (dHMN6 or HMN6).

An intronic growth hormone receptor mutation causing activation of a pseudoexon is associated with a broad spectrum of growth hormone insensitivity phenotypes.

CONTEXT: Inherited GH insensitivity (GHI) is usually caused by mutations in the GH receptor (GHR). Patients present with short stature associated with high GH and low IGF-I levels and may have midfacial hypoplasia (typical Laron syndrome facial features). We previously described four mildly affected GHI patients with an intronic mutation in the GHR gene (A(-1)-->G(-1) substitution in intron 6), resulting in the activation of a pseudoexon (6Psi) and inclusion of 36 amino acids. OBJECTIVE: The study aimed to analyze the clinical and genetic characteristics of additional GHI patients with the pseudoexon (6Psi) mutation. DESIGN/PATIENTS: Auxological, biochemical, genetic, and haplotype data from seven patients with severe short stature and biochemical evidence of GHI were assessed. MAIN OUTCOME MEASURES: We assessed genotype-phenotype relationship. RESULTS: One patient belongs to the same extended family, previously reported. She has normal facial features, and her IGF-I levels are in the low-normal range for age. The six unrelated patients, four of whom have typical Laron syndrome facial features, have heights ranging from -3.3 to -6.0 sd and IGF-I levels that vary from normal to undetectable. We hypothesize that the marked difference in biochemical and clinical phenotypes might be caused by variations in the splicing efficiency of the pseudoexon. CONCLUSIONS: Activation of the pseudoexon in the GHR gene can lead to a variety of GHI phenotypes. Therefore, screening for the presence of this mutation should be performed in all GHI patients without mutations in the coding exons.