Gene-encoded surface antigens of Legionella pneumophila and their role in pathogenicity

The pneumonial agent, Legionella pneumophila, is the predominant bacterium responsible for Legionnaires\u27 disease. Experimentally, these organisms have demonstrated the ability to adhere to host cells without the presence of a mucopolysaccharide layer. A recombinant plasmid, pLP 116, resulting from the ligation of a Hae III digest from the L. pneumophila, Nottingham N\sb7 genome and Sma I-digested pUC 19 vector was shown to encode for a 25 kilodalton (kDa) major outer membrane protein (MOMP) of L. pneumophila by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS). This protein was detected on the surface of an E. coli clone (LP 116) by immunoassays. Virulence testing using the fertile chicken egg lethality assay determined that the clone experienced increased virulence over that of the parent strain. The E. coli parent strain was found to be non-adherent to U937 cells in culture while the clone LP 116 experienced a 40-55% increase in adherence. The L. pneumophila N\sb7 strain demonstrated 100% binding; however, L. pneumophila and clone LP 116 incubated with MOMP-specific monoclonal antibody experienced a complete loss of adherence to U937 cells. Organisms coated with the monoclonal antibody did not infect fertile chicken eggs at any dilution. The outer membrane protein (OMP) profiles of the attenuated derivative of the L. pneumophila isolate used in this study showed a decrease in the 25 kDa protein and the presence of a 31 kDa protein not found in the OMP profile of the virulent strain. This laboratory strain experienced an increase in lethal dose (LD\sb{50}) values in the chicken embryonated egg lethality assay. When the recombinant plasmid pLP 116 was electroporated (electrically transformed) into the attenuated L. pneumophila derivative the 25 kDa protein was produced in greater amounts and the 31 kDa band was no longer present. The LD\sb{50} values of the transformed attenuated L. pneumophila N\sb7 strain decreased to that of the original isolate. This study has shown the first reported difference between what appears to be genotypically and phenotypically similar organisms. It also has demonstrated that the 25 kDa MOMP of L. pneumophila plays an important role in adherence of the organism and that the 25 kDa MOMP can be recognized as a virulence factor related to the ability of the organism to cause infection

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs

Cognitive functioning throughout adulthood and illness stages in individuals with psychotic disorders and their unaffected siblings.

Important questions remain about the profile of cognitive impairment in psychotic disorders across adulthood and illness stages. The age-associated profile of familial impairments also remains unclear, as well as the effect of factors, such as symptoms, functioning, and medication. Using cross-sectional data from the EU-GEI and GROUP studies, comprising 8455 participants aged 18 to 65, we examined cognitive functioning across adulthood in patients with psychotic disorders (n = 2883), and their unaffected siblings (n = 2271), compared to controls (n = 3301). An abbreviated WAIS-III measured verbal knowledge, working memory, visuospatial processing, processing speed, and IQ. Patients showed medium to large deficits across all functions (ES range = -0.45 to -0.73, p < 0.001), while siblings showed small deficits on IQ, verbal knowledge, and working memory (ES = -0.14 to -0.33, p < 0.001). Magnitude of impairment was not associated with participant age, such that the size of impairment in older and younger patients did not significantly differ. However, first-episode patients performed worse than prodromal patients (ES range = -0.88 to -0.60, p < 0.001). Adjusting for cannabis use, symptom severity, and global functioning attenuated impairments in siblings, while deficits in patients remained statistically significant, albeit reduced by half (ES range = -0.13 to -0.38, p < 0.01). Antipsychotic medication also accounted for around half of the impairment in patients (ES range = -0.21 to -0.43, p < 0.01). Deficits in verbal knowledge, and working memory may specifically index familial, i.e., shared genetic and/or shared environmental, liability for psychotic disorders. Nevertheless, potentially modifiable illness-related factors account for a significant portion of the cognitive impairment in psychotic disorders.The European Community’s Seventh Framework Programme under grant agreement No. HEALTH-F2-2010-241909 (EU-GEI)

DNA methylation patterns identify subgroups of pancreatic neuroendocrine tumors with clinical association

Here we report the DNA methylation profile of 84 sporadic pancreatic neuroendocrine tumors (PanNETs) with associated clinical and genomic information. We identified three subgroups of PanNETs, termed T1, T2 and T3, with distinct patterns of methylation. The T1 subgroup was enriched for functional tumors and ATRX, DAXX and MEN1 wild-type genotypes. The T2 subgroup contained tumors with mutations in ATRX, DAXX and MEN1 and recurrent patterns of chromosomal losses in half of the genome with no association between regions with recurrent loss and methylation levels. T2 tumors were larger and had lower methylation in the MGMT gene body, which showed positive correlation with gene expression. The T3 subgroup harboured mutations in MEN1 with recurrent loss of chromosome 11, was enriched for grade G1 tumors and showed histological parameters associated with better prognosis. Our results suggest a role for methylation in both driving tumorigenesis and potentially stratifying prognosis in PanNETs

Immune Responses to AAV in Canine Muscle Monitored by Cellular Assays and Noninvasive Imaging

We previously demonstrated that direct intramuscular injection of rAAV2 or rAAV6 in wild-type dogs resulted in robust T-cell responses to viral capsid proteins, and others have shown that cellular immunity to adeno-associated virus (AAV) capsid proteins coincided with liver toxicity and elimination of transgene expression in a human trial of hemophilia B. Here, we show that the heparin-binding ability of a given AAV serotype does not determine the induction of T-cell responses following intramuscular injection in dogs, and identify multiple epitopes in the AAV capsid protein that are recognized by T cells elicited by AAV injection. We also demonstrate that noninvasive magnetic resonance imaging (MRI) can accurately detect local inflammatory responses following intramuscular rAAV injection in dogs. These studies suggest that pseudotyping rAAV vectors to remove heparin-binding activity will not be sufficient to abrogate immunogenicity, and validate the utility of enzyme-linked immunosorbent spot (ELISpot) assay and MRI for monitoring immune and inflammatory responses following intramuscular injection of rAAV vectors in preclinical studies in dogs. These assays should be incorporated into future human clinical trials of AAV gene therapy to monitor immune responses