Critical values for Lawshe's content validity ratio: revisiting the original methods of calculation

YesThe content validity ratio originally proposed by Lawshe is widely used to quantify content validity and yet methods used to calculate the original critical values were never reported. Methods for original calculation of critical values are suggested along with tables of exact binomial probabilities

Neutrophil activation in systemic capillary leak syndrome (Clarkson disease)

Systemic capillary leak syndrome (SCLS; Clarkson disease) is a rare orphan disorder characterized by transient yet recurrent episodes of hypotension and peripheral oedema due to diffuse vascular leakage of fluids and proteins into soft tissues. Humoral mediators, cellular responses and genetic features accounting for the clinical phenotype of SCLS are virtually unknown. Here, we searched for factors altered in acute SCLS plasma relative to matched convalescent samples using multiplexed aptamer‐based proteomic screening. Relative amounts of 612 proteins were changed greater than twofold and 81 proteins were changed at least threefold. Among the most enriched proteins in acute SCLS plasma were neutrophil granule components including bactericidal permeability inducing protein, myeloperoxidase and matrix metalloproteinase 8. Neutrophils isolated from blood of subjects with SCLS or healthy controls responded similarly to routine pro‐inflammatory mediators. However, acute SCLS sera activated neutrophils relative to remission sera. Activated neutrophil supernatants increased permeability of endothelial cells from both controls and SCLS subjects equivalently. Our results suggest systemic neutrophil degranulation during SCLS acute flares, which may contribute to the clinical manifestations of acute vascular leak

Role of laeA in the regulation of alb1, gliP , Conidial Morphology and Virulence in Aspergillus fumigatus

The alb1 (pksP) gene has been reported as a virulence factor controlling the pigmentation and morphology of conidia in Aspergillus fumigatus. A recent report suggested that laeA regulates alb1 expression and conidial morphology but not pigmentation in the A. fumigatus strain AF293. laeA has also been reported to regulate the synthesis of secondary metabolites, such as gliotoxin. We compared the role of laeA in the regulation of conidial morphology and the expression of alb1 and gliP in strains B-5233 and AF293, which differ in colony morphology and nutritional requirements. Deletion of laeA did not affect conidial morphology or pigmentation in these strains, suggesting that laeA is not involved in alb1 regulation during conidial morphogenesis. Deletion of laeA, however, caused down-regulation of alb1 during mycelial growth in a liquid medium. Transcription of gliP, involved in the synthesis of gliotoxin, was drastically reduced in B-5233laeAΔ, and the gliotoxin level found in the culture filtrates was 20% of wild-type concentrations. While up-regulation of gliP in AF293 was comparable to that in B-5233, the relative mRNA level in AF293laeAΔ was about fourfold lower than that in B-5233laeAΔ. Strain B-5233lae4Δ caused slower onset of fatal infection in mice relative to that with B-5233. Histopathology of sections from lungs of infected mice corroborated the survival data. Culture filtrates from B-5233laeAΔ caused reduced death in thymoma cells and were less inhibitory to a respiratory burst of neutrophils than culture filtrates from B-5233. Our results suggest that while laeA is not involved in the regulation of alb1 function in conidial morphology, it regulates the synthesis of gliotoxin and the virulence of A. fumigatus

Hepatic abnormalities in patients with chronic granulomatous disease

Chronic granulomatous disease (CGD) is a rare congenital disorder characterized by repeated bacterial and fungal infections. Aside from a high incidence of liver abscess, little is known about hepatic involvement in CGD. The aim of this study was to describe the spectrum of liver abnormalities seen in CGD. The charts of 194 patients with CGD followed at the NIH were reviewed, with a focus on liver abnormalities. Liver enzyme elevations occurred on at least one occasion in 73% of patients during a mean of 8.9 years of follow-up. ALT elevations were generally transient. Although transient alkaline phosphatase (ALP) elevations were also common, persistent ALP elevations lasting up to 17.6 years were seen in 25% of patients. Liver abscess occurred in 35% of patients. Drug-induced hepatotoxicity was documented in 15% of patients but likely occurred more frequently. Hepatomegaly was found in 34% and splenomegaly in 56% of patients. Liver histology showed granulomata in 75% and lobular hepatitis in 90% of specimens. Venopathy of the portal vein was common (80%) and associated with splenomegaly. Venopathy of the central vein was also common (63%) and was associated with the number of abscess episodes. Nodular regenerative hyperplasia (NRH) was seen in 9 patients, including 6 of 12 autopsy specimens. CONCLUSION: Liver enzyme abnormalities occur frequently in patients with CGD. In addition to liver abscesses and granulomata, drug hepatotoxicity is likely underappreciated. Vascular lesions such as venopathy and--to a lesser extent--NRH are common. The cause and clinical consequences of venopathy await prospective evaluation

Simultaneous Host-Pathogen Transcriptome Analysis during Granulibacter bethesdensis Infection of Neutrophils from Healthy Subjects and Patients with Chronic Granulomatous Disease

Polymorphonuclear leukocytes (PMN) from patients with chronic granulomatous disease (CGD) fail to produce microbicidal concentrations of reactive oxygen species (ROS) due to mutations in NOX2. Patients with CGD suffer from severe, life-threatening infections and inflammatory complications. Granulibacter bethesdensis is an emerging Gram-negative pathogen in CGD that resists killing by PMN of CGD patients (CGD PMN) and inhibits PMN apoptosis through unknown mechanisms. Microarray analysis was used to study mRNA expression in PMN from healthy subjects (normal PMN) and CGD PMN during incubation with G. bethesdensis and, simultaneously, in G. bethesdensis with normal and CGD PMN. We detected upregulation of antiapoptotic genes (e.g., XIAP and GADD45B) and downregulation of proapoptotic genes (e.g., CASP8 and APAF1) in infected PMN. Transcript and protein levels of inflammation- and immunity-related genes were also altered. Upon interaction with PMN, G. bethesdensis altered the expression of ROS resistance genes in the presence of normal but not CGD PMN. Levels of bacterial stress response genes, including the ClpB gene, increased during phagocytosis by both normal and CGD PMN demonstrating responses to oxygen-independent PMN antimicrobial systems. Antisense knockdown demonstrated that ClpB is dispensable for extracellular growth but is essential for bacterial resistance to both normal and CGD PMN. Metabolic adaptation of Granulibacter growth in PMN included the upregulation of pyruvate dehydrogenase. Pharmacological inhibition of pyruvate dehydrogenase by triphenylbismuthdichloride was lethal to Granulibacter. This study expands knowledge of microbial pathogenesis of Granulibacter in cells from permissive (CGD) and nonpermissive (normal) hosts and identifies potentially druggable microbial factors, such as pyruvate dehydrogenase and ClpB, to help combat this antibiotic-resistant pathogen.This is the publisher’s final pdf. The published article is copyrighted by American Society for Microbiology and can be found at: http://iai.asm.org

Genes Differentially Expressed in Conidia and Hyphae of Aspergillus fumigatus upon Exposure to Human Neutrophils

Aspergillus fumigatus is the most common etiologic agent of invasive aspergillosis in immunocompromised patients. Several studies have addressed the mechanism involved in host defense but only few have investigated the pathogen's response to attack by the host cells. To our knowledge, this is the first study that investigates the genes differentially expressed in conidia vs hyphae of A. fumigatus in response to neutrophils from healthy donors as well as from those with chronic granulomatous disease (CGD) which are defective in the production of reactive oxygen species.Transcriptional profiles of conidia and hyphae exposed to neutrophils, either from normal donors or from CGD patients, were obtained by using the genome-wide microarray. Upon exposure to either normal or CGD neutrophils, 244 genes were up-regulated in conidia but not in hyphae. Several of these genes are involved in the degradation of fatty acids, peroxisome function and the glyoxylate cycle which suggests that conidia exposed to neutrophils reprogram their metabolism to adjust to the host environment. In addition, the mRNA levels of four genes encoding proteins putatively involved in iron/copper assimilation were found to be higher in conidia and hyphae exposed to normal neutrophils compared to those exposed to CGD neutrophils. Deletants in several of the differentially expressed genes showed phenotypes related to the proposed functions, i.e. deletants of genes involved in fatty acid catabolism showed defective growth on fatty acids and the deletants of iron/copper assimilation showed higher sensitivity to the oxidative agent menadione. None of these deletants, however, showed reduced resistance to neutrophil attack.This work reveals the complex response of the fungus to leukocytes, one of the major host factors involved in antifungal defense, and identifies fungal genes that may be involved in establishing or prolonging infections in humans