An evaluation of methods for the isolation of nontuberculous mycobacteria from patients with cystic fibrosis, bronchiectasis and patients assessed for lung transplantation.

BACKGROUND: RGM medium is an agar-based, selective culture medium designed for the isolation of nontuberculous mycobacteria (NTM) from the sputum of patients with cystic fibrosis (CF). We evaluated RGM medium for the detection of NTM in patients with CF (405 samples), bronchiectasis (323 samples) and other lung diseases necessitating lung transplantation (274 samples). METHODS: In total, 1002 respiratory samples from 676 patients were included in the study. Direct culture on RGM medium, with incubation at two temperatures (30 °C and 37 °C), was compared with conventional culture of decontaminated samples for acid-fast bacilli (AFB) using both a solid medium (Löwenstein-Jensen medium) and a liquid medium (the Mycobacterial Growth Indicator Tube; MGIT). RESULTS: For all three patient groups, significantly more isolates of NTM were recovered using RGM medium incubated at 30 °C than by any other method (sensitivity: 94.6% vs. 22.4% for conventional AFB culture; P < 0.0001). Significantly more isolates of Mycobacterium abscessus complex were isolated on RGM at 30 °C than by AFB culture (sensitivity: 96.1% vs. 58.8%; P < 0.0001). The recovery of Mycobacterium avium complex was also greater using RGM medium at 30 °C compared to AFB culture (sensitivity: 83% vs. 70.2%), although this difference was not statistically significant and a combination of methods was necessary for optimal recovery (P = 0.21). CONCLUSIONS: In the largest study of RGM medium to date, we reaffirm its utility for isolation of NTM from patients with CF. Furthermore; we show that it also provides an effective tool for culture of respiratory samples from patients with bronchiectasis and other lung diseases.

A novel COL4A1 frameshift mutation in familial kidney disease: the importance of the C-terminal NC1 domain of type IV collagen.

BACKGROUND: Hereditary microscopic haematuria often segregates with mutations of COL4A3, COL4A4 or COL4A5 but in half of families a gene is not identified. We investigated a Cypriot family with autosomal dominant microscopic haematuria with renal failure and kidney cysts. METHODS: We used genome-wide linkage analysis, whole exome sequencing and cosegregation analyses. RESULTS: We identified a novel frameshift mutation, c.4611_4612insG:p.T1537fs, in exon 49 of COL4A1. This mutation predicts truncation of the protein with disruption of the C-terminal part of the NC1 domain. We confirmed its presence in 20 family members, 17 with confirmed haematuria, 5 of whom also had stage 4 or 5 chronic kidney disease. Eleven family members exhibited kidney cysts (55% of those with the mutation), but muscle cramps or cerebral aneurysms were not observed and serum creatine kinase was normal in all individuals tested. CONCLUSIONS: Missense mutations of COL4A1 that encode the CB3 [IV] segment of the triple helical domain (exons 24 and 25) are associated with HANAC syndrome (hereditary angiopathy, nephropathy, aneurysms and cramps). Missense mutations of COL4A1 that disrupt the NC1 domain are associated with antenatal cerebral haemorrhage and porencephaly, but not kidney disease. Our findings extend the spectrum of COL4A1 mutations linked with renal disease and demonstrate that the highly conserved C-terminal part of the NC1 domain of the α1 chain of type IV collagen is important in the integrity of glomerular basement membrane in humans.

Combinatorial Conflicting Homozygosity (CCH) analysis enables the rapid identification of shared genomic regions in the presence of multiple phenocopies.

BACKGROUND: The ability to identify regions of the genome inherited with a dominant trait in one or more families has become increasingly valuable with the wide availability of high throughput sequencing technology. While a number of methods exist for mapping of homozygous variants segregating with recessive traits in consanguineous families, dominant conditions are conventionally analysed by linkage analysis, which requires computationally demanding haplotype reconstruction from marker genotypes and, even using advanced parallel approximation implementations, can take substantial time, particularly for large pedigrees. In addition, linkage analysis lacks sensitivity in the presence of phenocopies (individuals sharing the trait but not the genetic variant responsible). Combinatorial Conflicting Homozygosity (CCH) analysis uses high density biallelic single nucleotide polymorphism (SNP) marker genotypes to identify genetic loci within which consecutive markers are not homozygous for different alleles. This allows inference of identical by descent (IBD) inheritance of a haplotype among a set or subsets of related or unrelated individuals. RESULTS: A single genome-wide conflicting homozygosity analysis takes <3 seconds and parallelisation permits multiple combinations of subsets of individuals to be analysed quickly. Analysis of unrelated individuals demonstrated that in the absence of IBD inheritance, runs of no CH exceeding 4 cM are not observed. At this threshold, CCH is >97% sensitive and specific for IBD regions within a pedigree exceeding this length and was able to identify the locus responsible for a dominantly inherited kidney disease in a Turkish Cypriot family in which six out 17 affected individuals were phenocopies. It also revealed shared ancestry at the disease-linked locus among affected individuals from two different Cypriot populations. CONCLUSIONS: CCH does not require computationally demanding haplotype reconstruction and can detect regions of shared inheritance of a haplotype among subsets of related or unrelated individuals directly from SNP genotype data. In contrast to parametric linkage allowing for phenocopies, CCH directly provides the exact number and identity of individuals sharing each locus. CCH can also identify regions of shared ancestry among ostensibly unrelated individuals who share a trait. CCH is implemented in Python and is freely available (as source code) from http://sourceforge.net/projects/cchsnp/ .

Spatial variation in the risk of hospitalization with childhood pneumonia and empyema in the North of England.

The aim of this study was to investigate spatial variation in risk of hospitalization in childhood pneumonia and empyema in the North of England and associated risk factors. Data on childhood (0-14 years) hospital admissions with a diagnosis pneumonia or empyema were linked to postcode districts. Bayesian conditional autoregressive models were used to evaluate spatial variation and the relevance of specific spatial covariates in an area-based study using postcode as the areal unit. There was a sixfold variation in the risk of hospitalization due to pneumonia across the study region. Variation in risk was associated with material deprivation, Child Well-being Index (CWI) health domain score, number of children requiring local authority support, and distance to hospital. No significant spatial variation in risk for empyema was found.

Impact of the 7-valent pneumococcal conjugate vaccine on the incidence of childhood pneumonia.

In September 2006, the 7-valent pneumococcal conjugate vaccine (PCV7) was added to the UK immunization programme. We aimed to evaluate the impact of PCV7 on the incidence of all-cause community-acquired pneumonia (CAP) in children. A prospective survey was undertaken in 2008-2009 at 11 hospitals in North East England of children aged 0-16 years with radiologically confirmed pneumonia. Data were compared to those from a similar survey undertaken in the same hospitals in 2001-2002. A total of 542 children were enrolled, of which 74% were aged <5 years. PCV7 uptake was 90∙7%. The incidence of pneumonia was 11∙8/10,000 [95% confidence interval (CI) 10∙9-12∙9], and the hospitalization rate was 9∙9/10,000 (95% CI 9∙0-10∙9). Compared to 2001, there was a 19% (95% CI 8-29) reduction in the rate of CAP in those aged <5 years, and in those <2 years a 33∙1% (95% CI 20-45) reduction in the incidence of CAP and 38∙1% (95% CI 24-50) reduction in hospitalization rates. However, for those unvaccinated aged ≥5 years, there was no difference in the incidence of CAP and hospitalization rate between both surveys. Since 2001, the overall reduction in incidence was 17∙7% (95% CI 8-26) and for hospitalization 18∙5% (95% CI 8-28). For the <5 years age group there was a lower incidence of CAP in PCV7-vaccinated children (25∙2/10,000, 95% CI 22∙6-28∙2) than in those that were not vaccinated (37∙4/10,000, 95% CI 29∙2-47∙1). In conclusion, PCV7 has reduced both incidence and rate of hospitalization of pneumonia in children, particularly in the <2 years age group.

Magnetic order in FeCr(2)S(4)-type chalcogenide spinels.

The thiospinels of composition FeCr(2)S(4) and Fe(1+x)Cr(2-2x)Sn(x)S(4) with 0

Insulin-like growth factor-I protects myoblasts from apoptosis but requires other factors to stimulate proliferation.

Insulin-like growth factor-I (IGF-I) has been shown to stimulate myoblast proliferation for a limited time after which serum is required to reactivate IGF-I-stimulated myoblast proliferation. The aim of these studies was to determine whether IGF-I can stimulate myoblast proliferation and/or inhibit apoptosis alone or whether co-factors are necessary. This was achieved by investigating the proliferative response of L6 myoblasts to IGF-I and horse serum (HS) and by examining the status of cells in terms of cell number, substrate adherence, cell viability and DNA laddering following incubation with IGF-I and HS. L6 myoblasts proliferate in response to IGF-I after 36 h is not due to accumulation of waste products or lack of IGF-I. The addition of a low level (1% v/v) of HS restores the ability of myoblasts to proliferate in response to IGF-I and this supports the existence of a mitogenic competence factor. Furthermore, myoblasts failing to proliferate in response to IGF-I after 36 h regain the capacity to respond to IGF-I for a further period of 36 h when exposed to fetal bovine serum. Following the initial (36 h) phase of IGF-I-stimulated proliferation, removal of both IGF-I and HS led to a dramatic (60%) reduction in the number of cells fully attached to the culture vessel, with 60% of the completely detached cells dead. Agarose gel electrophoresis of extracts from these detached cells revealed higher levels of DNA laddering than extracts prepared from attached cells with IGF-I present. This suggests that IGF-I acts as a survival factor by protecting cells from apoptosis. In conclusion these experiments support the presence of a mitogenic competence factor in horse serum, which restores the ability of cells to proliferate in response to IGF-I. Unlike proliferation, protection against apoptosis is achieved by IGF-I or HS independently of each other.

[Membrane fatty acids and blood antioxidants in 77 patients with HIV infection]. / Acides gras membranaires et anti-oxydants sériques chez 77 patients infectés par le VIH.

We have measured the fatty acid (FA) composition of erythrocyte membranes and plasma anti-oxidants in HIV+ patients. Saturated FA are higher and poly-unsaturated FA lower than in controls (P = 0.02). Selenium (Se) is lower in patients less than 400 CD4 cells/mm3 (P = 0.002). Vitamin A is lower in the HIV+ regardless of the CD4 cell count. Se and vitamin A are correlated to nutritional markers (body mass index and albumin).

Occlusal caries diagnosis in molar teeth from bitewing and panoramic radiographs.

INTRODUCTION: Previous studies have implied that the panoramic radiograph was inferior to the bitewing radiograph for caries diagnosis. However, these clinical studies lacked a method of validation. The aim of this study was to use an electronic caries meter (ECM II, LODE, Groningen, The Netherlands) to validate occlusal caries diagnosis made from bitewing and panoramic radiographs. MATERIALS AND METHOD: Forty-nine Army recruits were examined with the ECM, and had bitewing and panoramic radiographs taken. In total 299 molar occlusal surfaces were available for examination. Seven examiners viewed the bitewing and panoramic radiographs on two separate occasions and assessed each occlusal surface for dentine caries as 1: almost definitely no caries, 2: probably no caries, 3: unsure, 4: caries probably present, and 5: caries almost definitely present. This was repeated on 20% of the radiographs at two further separate sittings. ECM conductance readings greater than 9 were taken to indicate dentine caries. Examiner decisions that caries was probably and definitely considered to be present were taken as positive diagnoses. RESULTS: Bitewing and panoramic radiographs provided sensitivity values of 0.25 and 0.19 and specificity values of 0.93 and 0.97 respectively. ROC analysis indicated no statistically significant difference in diagnostic quality between the bitewing and panoramic radiographs. Intra-examiner reproducibility was found to be poor to moderate (Kappa values for bitewing = 0.31-0.44, panoramic = 0.07-0.54). CONCLUSION: No difference in overall diagnostic performance was found between bitewing and panoramic radiographs for the diagnosis of occlusal dentine caries.

Revealing organic carbon-nitrate linear relationship from UV spectra of freshwaters in agricultural environment.

A strong non linear relationship between nitrate and organic matter (assessed by dissolved organic carbon, DOC) has been recently demonstrated by Taylor and Townsend (2010), namely for freshwaters. In this context, our study explores this relation from the behavior of sets of normalized UV spectra (same area under each spectrum) of different water samples showing a hidden isosbestic point (HIP) around 225 nm. This HIP is linked to the existence of a simple relation between nitrate and DOC, the proportions of which vary according to the sampling location and environmental factors. In a second step, a simple linear model is proposed for nitrate-DOC relationship (α⋅NO3+ß⋅DOC=1) and a validation is proposed for more than 150 samples of different Brittany rivers and lakes. For samples of the largest watershed, a complementary exploitation from data acquired during the different campaigns confirmed the seasonal evolution between spring (high nitrate/low DOC) and autumn (high DOC/low nitrate). Further investigation on other freshwater samples is needed in order to improve the limits of this linear model.

Genetic testing can resolve diagnostic confusion in Alport syndrome.

Alport syndrome (AS) is a familial glomerular disorder resulting from mutations in the genes encoding several members of the type IV collagen protein family. Despite advances in molecular genetics, renal biopsy remains an important initial diagnostic tool. Histological diagnosis is challenging as features may be non-specific, particularly early in the disease course and in females with X-linked disease. We present three families for whom there was difficulty in correctly diagnosing AS or thin basement membrane nephropathy as a result of misinterpretation of non-specific and incomplete histology. We highlight the importance of electron microscopy and immunofluorescence in improving diagnostic yield and also the hazard of interpreting a descriptive histological term as a diagnostic label. Molecular genetic testing allows a definitive diagnosis to be made in index patients and at-risk family members.

Incidence of end-stage renal disease in the Turkish-Cypriot population of Northern Cyprus: a population based study.

BACKGROUND: This is the first report of the incidence and causes of end-stage renal disease (ESRD) of the Turkish-Cypriot population in Northern Cyprus. METHODS: Data were collected over eight consecutive years (2004-2011) from all those starting renal replacement therapy (RRT) in this population. Crude and age-standardised incidence at 90 days was calculated and comparisons made with other national registries. We collected DNA from the entire prevalent population. As an initial experiment we looked for two genetic causes of ESRD that have been reported in Greek Cypriots. RESULTS: Crude and age-standardised incidence at 90 days was 234 and 327 per million population (pmp) per year, respectively. The mean age was 63, and 62% were male. The age-adjusted prevalence of RRT in Turkish-Cypriots was 1543 pmp on 01/01/2011. The incidence of RRT is higher than other countries reporting to the European Renal Association - European Dialysis and Transplant Association, with the exception of Turkey. Diabetes is a major cause of ESRD in those under 65, accounting for 36% of incident cases followed by 30% with uncertain aetiology. 18% of the incident population had a family history of ESRD. We identified two families with thin basement membrane nephropathy caused by a mutation in COL4A3, but no new cases of CFHR5 nephropathy. CONCLUSIONS: This study provides the first estimate of RRT incidence in the Turkish-Cypriot population, describes the contribution of different underlying diagnoses to ESRD, and provides a basis for healthcare policy planning.

Magnetic interaction in ferrous antimonite, FeSb2O4, and some derivatives.

Ferrous antimonite, FeSb(2)O(4), which is isostructural with Pb(3)O(4), and some lead- and cobalt-doped variants of composition FeSb(1.5)Pb(0.5)O(4) and Co(0.5)Fe(0.5)Sb(1.5)Pb(0.5)O(4) have been examined by (57)Fe and (121)Sb Mössbauer spectroscopy. Antimony is present as Sb(3+). The presence of Pb(2+) on the antimony site induces partial oxidation of Fe(2+) to Fe(3+). There is no Verwey-type transition in which electrons are shared between iron in different oxidation states. The quasi-one-dimensional magnetic structure gives rise to situations in which weakly coupled Fe(2+) ions can coexist in a non-magnetic state alongside Fe(3+) ions in a magnetically ordered state.

Hematologic responses to carbon monoxide and altitude: a comparative study.

Groups of 40- and 90-day-old rats were exposed to 500 ppm CO (HbCO=31-43%) and simulated altitude (15,000 ft). Resulting hematologic changes were monitored after 1 day, 1 wk, 3-4 wk, and 9-11 wk of exposure. The two treatments resulted in similar changes in hemoglobin (Hb) and hematocrit ratio (Hct) in the young rats, while in the older group there were several small, but significant differences in these parameters. Mean corpuscular hemoglobin concentration (MCHC) fell 12.4-15.1% during the 1st wk of exposure in all groups except the young altitude rats, which remained at the control value. The former groups returned to control levels by the 3rd to 4th wk of exposure. Red cell 2,3-diphosphoglycerate (2,3-DPG) increased 21.6-26.1% above control during 3-4 wk of altitude exposure, but later (9-11 wk) returned toward control levels. CO exposure resulted in a sharp decrease in 2,3-DPG after 1 day of exposure only in the young rats. 2,3-DPG level progressively decreased with age in control rats. The effects of CO and altitude although similar in several respects (i.e., Hb, Hct, MCHC) differ with regard to red cell 2,3-DPG.

The role of Sb(5+) in the structure of Sb(2)O(3)-B(2)O(3) binary glasses--an NMR and Mössbauer spectroscopy study.

Glasses of general formula xSb(2)O(3) (1-x)B(2)O(3) (0

G-protein beta gamma subunits mediate specific phosphorylation of the protein-tyrosine phosphatase SH-PTP1 induced by lysophosphatidic acid.

SH-PTP1 is a protein-tyrosine phosphatase preferentially expressed in hematopoietic cells and bearing two SH2 (src homology-2) domains. In the human megakaryocytic cell line Dami, lysophosphatidic acid (LPA) promoted a rapid increase in SH-PTP1 phosphorylation on both serine and tyrosine residues. Only tyrosine phosphorylation was significantly inhibited by pertussis toxin and by the protein kinase C inhibitor GF109203X. Moreover, SH-PTP1 was phosphorylated upon challenge with other agonists acting via G-protein-coupled receptors such as alpha-thrombin, epinephrine, and ADP, whereas the closely related protein-tyrosine phosphatase SH-PTP2 failed to share such a regulation in Dami cells. We developed an in vitro assay that reproduced LPA-dependent phosphorylation of SH-PTP1 in a cell-free system. The fusion protein glutathione S-transferase-beta-adrenergic receptor kinase 1-(495-689) or the transducin subunit Galphat-GDP, which act as specific antagonists of Gbetagamma, inhibited SH-PTP1 phosphorylation. Moreover, purified transducin Gbetagamma subunits mimicked the effect of LPA. Finally, stable expression of beta-adrenergic receptor kinase 1-(495-689) in Dami cells resulted in the inhibition of SH-PTP1 as a specific target of protein kinases linked to G-protein-coupled receptors via Gbetagamma subunits.