Immunodiagnosis of tuberculosis: new questions, new tools conference 2008

Human infection with Mycobacterium tuberculosis exists as a spectrum of conditions ranging from asymptomatic infection to active disease. Novel, accurate tuberculosis immunodiagnostics have been introduced over the last decade, but it remains challenging to timely diagnose active disease and to accurately distinguish asymptomatic M. tuberculosis infection from immune memory resulting from a prior infection eradicated by the host response. The conference titled Immunodiagnosis of Tuberculosis: New Questions, New Tools, which was held on September 21-23, 2008 in Virginia Beach, Virginia, United States, brought together basic scientists and clinical experts to discuss recent progress in tuberculosis research and diagnosis. Global analyses of M. tuberculosis biology and the host immune response, with emphasis on systems approaches to the study of host-pathogen interactions, were presented. Moreover, conference participants discussed new tests in the pipeline and reviewed new technologies leading to novel assay formats. The discussion included technologies ranging from simple, inexpensive point-of-care tests to automated molecular platforms for detection of multiple infections based on the “lab on a chip” concept. It was also recognized that the utility of any new diagnostic relies on laboratory capacity, accessibility, costs, and test deployment. The conference included lessons from the field. For example, the application of existing technologies to neglected areas, such as diagnosis in children and HIV+ populations, was discussed

Prevalence of human papillomavirus cervical infection in an Italian asymptomatic population

BACKGROUND: In the last decade many studies have definitely shown that human papillomaviruses (HPVs) are the major cause of cervical carcinogenesis and, in the last few years, HPV testing has been proposed as a new and more powerful tool for cervical cancer screening. This issue is now receiving considerable attention in scientific and non scientific press and HPV testing could be considered the most important change in this field since the introduction of cervical cytology. This paper reports our prevalence data of HPV infection collected in the '90s, while a follow up of these patients is ongoing. METHODS: For this study we used polymerase chain reaction (PCR) to search HPV DNA sequences in cervical cell scrapings obtained from 503 asymptomatic women attending regular cervical cancer screening program in the city of Genova, Italy. All patients were also submitted to a self-administered, standardized, questionnaire regarding their life style and sexual activity. On the basis of the presence of HPV DNA sequences women were separated into two groups: "infected" and "non infected" and a statistical analysis of the factors potentially associated with the infection group membership was carried out. RESULTS: The infection rate was 15.9% and the most frequent viral type was HPV 16. CONCLUSION: Our HPV positivity rate (15.9%) was consistent to that reported by other studies on European populations

The T1405N Carbamoyl Phosphate Synthetase Polymorphism Does Not Affect Plasma Arginine Concentrations in Preterm Infants

A C-to-A nucleotide transversion (T1405N) in the gene that encodes carbamoyl-phosphate synthetase 1 (CPS1) has been associated with changes in plasma concentrations of L-arginine in term and near term infants but not in adults. In preterm infants homozygosity for the CPS1 Thr1405 variant (CC genotype) was associated with an increased risk of having necrotizing enterocolitis (NEC). Plasma L-arginine concentrations are decreased in preterm infants with NEC.To examine the putative association between the CPS1 T1405N polymorphism and plasma arginine concentrations in preterm infants.Prospective multicenter cohort study. Plasma and DNA samples were collected from 128 preterm infants (<30 weeks) between 6 and 12 hours after birth. Plasma amino acid and CPS1 T1405N polymorphism analysis were performed.Distribution of genotypes did not differ between the preterm (CC:CA:AA = 55.5%:33.6%:10.9%, n = 128) and term infants (CC:CA:AA = 54.2%:35.4%:10.4%, n = 96). There was no association between the CPS1 genotype and plasma L-arginine or L-citrulline concentration, or the ornithine to citrulline ratio, which varies inversely with CPS1 activity. Also the levels of asymmetric dimethylarginine, and symmetric dimethylarginine were not significantly different among the three genotypes.The present study in preterm infants did not confirm the earlier reported association between CPS1 genotype and L-arginine levels in term infants

OP0063 QUANTITATIVE COMPUTED TOMOGRAPHY PREDICTS 10-YEAR MORTALITY IN INTERSTITIAL LUNG DISEASE RELATED TO SYSTEMIC SCLEROSI

Background: Interstitial lung disease (ILD) is the main cause of death in Systemic Sclerosis (SSc). Chest CT is the gold standard in detecting ILD although it is not easy to understand its prognostic value. ILD qualitative assessment is almost worthless. Goh et al. semi quantitative score of ILD extent is related to mortality risk but it is burdened by relevant inter/intra-readers variability. An operator independent algorithm based on voxel-wise analysis proved to identify SSc patients with an increased risk of mortality according to prediction models. Objectives: To verify if quantitative analysis of chest CT (QCT) predict 10 years-mortality in SSc patients. Methods: SSc patients with availability of a chest CT were enrolled in 13 different centers. The CT voxel-wise analysis with a free software (www.horosproject.com) provided QCT indexes: kurtosis, skewness, mean lung attenuation and standard deviation. Patients

The young stellar content of the giant H II regions M 8, G333.6-0.2, and NGC 6357 with VLT/KMOS

Context. The identification and characterisation of populations of young massive stars in (giant) H II regions provides important constraints on (i) the formation process of massive stars and their early feedback on the environment, and (ii) the initial conditions for population synthesis models predicting the evolution of ensembles of stars. Aims. We identify and characterise the stellar populations of the following young giant H II regions: M 8, G333.6-0.2, and NGC 6357. Methods. We have acquired H-and K-band spectra of around 200 stars using the K-band Multi Object Spectrograph on the ESO Very Large Telescope. The targets for M 8 and NGC 6357 were selected from the Massive Young Star-Forming Complex Study in Infrared and X-ray (MYStIX), which combines X-ray observations with near-infrared (NIR) and mid-infrared data. For G333.6-0.2, the sample selection is based on the NIR colours combined with X-ray data. We introduce an automatic spectral classification method in order to obtain temperatures and luminosities for the observed stars. We analysed the stellar populations using their photometric, astrometric, and spectroscopic properties and compared the position of the stars in the Hertzprung-Russell diagram with stellar evolution models to constrain their ages and mass ranges. Results. We confirm the presence of candidate ionising sources in the three regions and report new ones, including the first spectroscopically identified O stars in G333.6-0.2. In M 8 and NGC 6357, two populations are identified: (i) OB main-sequence stars (M > 5 M· ) and (ii) pre-main sequence stars (M ≈ 0.5ℓ-ℓ 5 M· ). The ages of the clusters are ∼1-3 Myr, 90% probability of being members of the H II region, whereas a selection based on NIR colours leads to a membership probability of only ∼70%

A role of BRCA1 and BRCA2 germline mutations in breast cancer susceptibility within Sardinian population

<p>Abstract</p> <p>Background</p> <p>In recent years, numerous studies have assessed the prevalence of germline mutations in <it>BRCA1 </it>and <it>BRCA2 </it>genes in various cohorts. We here extensively investigated the prevalence and geographical distribution of <it>BRCA1-2 </it>mutations in the entire genetically-homogeneous Sardinian population. The occurrence of phenotypic characteristics which may be predictive for the presence of <it>BRCA1-2 </it>germline mutations was also evaluated.</p> <p>Methods</p> <p>Three hundred and forty-eight breast cancer patients presenting a familial recurrence of invasive breast or ovarian carcinoma with at least two affected family members were screened for <it>BRCA1-2 </it>mutations by DHPLC analysis and DNA sequencing. Association of <it>BRCA1 </it>and <it>BRCA2 </it>mutational status with clinical and pathological parameters was evaluated by Pearson's Chi-Squared test.</p> <p>Results and Conclusion</p> <p>Overall, 8 <it>BRCA1 </it>and 5 <it>BRCA2 </it>deleterious mutations were detected in 35/348 (10%) families; majority (23/35;66%) of mutations was found in <it>BRCA2 </it>gene. The geographical distribution of <it>BRCA1-2 </it>mutations was related to three specific large areas of Sardinia, reflecting its ancient history: <it>a</it>) the Northern area, linguistically different from the rest of the island (where a <it>BRCA2 c.8764_8765delAG </it>mutation with founder effect was predominant); <it>b</it>) the Middle area, land of the ancient Sardinian population (where <it>BRCA2 </it>mutations are still more common than <it>BRCA1 </it>mutations); and <it>c</it>) the South-Western area, with many Phoenician and Carthaginian locations (where <it>BRCA1 </it>mutations are prevalent). We also found that phenotypic features such as high tumor grading and lack of expression of estrogen/progesterone receptors together with age at diagnosis and presence of ovarian cancer in the family may be predictive for the presence of <it>BRCA1-2 </it>germline mutations.</p

Kita Driven Expression of Oncogenic HRAS Leads to Early Onset and Highly Penetrant Melanoma in Zebrafish

Melanoma is the most aggressive and lethal form of skin cancer. Because of the increasing incidence and high lethality of melanoma, animal models for continuously observing melanoma formation and progression as well as for testing pharmacological agents are needed.Using the combinatorial Gal4-UAS system, we have developed a zebrafish transgenic line that expresses oncogenic HRAS under the kita promoter. Already at 3 days transgenic kita-GFP-RAS larvae show a hyper-pigmentation phenotype as earliest evidence of abnormal melanocyte growth. By 2-4 weeks, masses of transformed melanocytes form in the tail stalk of the majority of kita-GFP-RAS transgenic fish. The adult tumors evident between 1-3 months of age faithfully reproduce the immunological, histological and molecular phenotypes of human melanoma, but on a condensed time-line. Furthermore, they show transplantability, dependence on mitfa expression and do not require additional mutations in tumor suppressors. In contrast to kita expressing melanocyte progenitors that efficiently develop melanoma, mitfa expressing progenitors in a second Gal4-driver line were 4 times less efficient in developing melanoma during the three months observation period.This indicates that zebrafish kita promoter is a powerful tool for driving oncogene expression in the right cells and at the right level to induce early onset melanoma in the presence of tumor suppressors. Thus our zebrafish model provides a link between kita expressing melanocyte progenitors and melanoma and offers the advantage of a larval phenotype suitable for large scale drug and genetic modifier screens

Altered maturation of peripheral blood dendritic cells in patients with breast cancer

Tumours have at least two mechanisms that can alter dendritic cell (DC) maturation and function. The first affects the ability of haematopoietic progenitors to differentiate into functional DCs; the second affects their differentiation from CD14+ monocytes, promoting an early but dysfunctional maturation. The aim of this study was to evaluate the in vivo relevance of these pathways in breast cancer patients. For this purpose, 53 patients with invasive breast cancer were compared to 68 healthy controls. To avoid isolation or culture procedures for enrichment of DCs, analyses were directly performed by flow cytometry on whole-blood samples. The expression of surface antigens and intracellular accumulation of regulatory cytokines upon LPS stimulation were evaluated. The number of DCs, and in particular of the myeloid subpopulation, was markedly reduced in cancer patients (P < 0.001). Patient DCs were characterized by a more mature phenotype compared with controls (P = 0.016), and had impaired production of IL-12 (P < 0.001), These alterations were reverted by surgical resection of the tumour. To investigate the possible role of some tumour-related immunoactive soluble factors, we measured the plasmatic levels of vascular endothelial growth factor, IL-10 and spermine. A significant inverse correlation between spermine concentration and the percentage of DCs expressing IL-12 was found. Evidence was also obtained that in vitro exposure of monocyte-derived DCs to spermine promoted their activation and maturation, and impaired their function. Taken together, our results suggest that both the above-described mechanisms could concomitantly act in breast cancer to affect DC differentiation, and that spermine could be a mediator of dysfunctional maturation of DCs

Induction of cortical plasticity and improved motor performance following unilateral and bilateral transcranial direct current stimulation of the primary motor cortex

BACKGROUND: Transcranial direct current stimulation (tDCS) is a non-invasive technique that modulates the excitability of neurons within the primary motor cortex (M1). Research shows that anodal-tDCS applied over the non-dominant M1 (i.e. unilateral stimulation) improves motor function of the non-dominant hand. Similarly, previous studies also show that applying cathodal tDCS over the dominant M1 improves motor function of the non-dominant hand, presumably by reducing interhemispheric inhibition. In the present study, one condition involved anodal-tDCS over the non-dominant M1 (unilateral stimulation) whilst a second condition involved applying cathodal-tDCS over the dominant M1 and anodal-tDCS over non-dominant M1 (bilateral stimulation) to determine if unilateral or bilateral stimulation differentially modulates motor function of the non-dominant hand. Using a randomized, cross-over design, 11 right-handed participants underwent three stimulation conditions: 1) unilateral stimulation, that involved anodal-tDCS applied over the non-dominant M1, 2) bilateral stimulation, whereby anodal-tDCS was applied over the non-dominant M1, and cathodal-tDCS over the dominant M1, and 3) sham stimulation. Transcranial magnetic stimulation (TMS) was performed before, immediately after, 30 and 60 minutes after stimulation to elucidate the neural mechanisms underlying any potential after-effects on motor performance. Motor function was evaluated by the Purdue pegboard test. RESULTS: There were significant improvements in motor function following unilateral and bilateral stimulation when compared to sham stimulation at all-time points (all P 0.05). Furthermore, changes in corticomotor plasticity were not related to changes in motor performance. CONCLUSION: These results indicate that tDCS induced behavioural changes in the non-dominant hand as a consequence of mechanisms associated with use-dependant cortical plasticity that is independent of the electrode arrangement

A Prognostic Model for Estimating the Time to Virologic Failure in HIV-1 Infected Patients Undergoing a New Combination Antiretroviral Therapy Regimen

<p>Abstract</p> <p>Background</p> <p>HIV-1 genotypic susceptibility scores (GSSs) were proven to be significant prognostic factors of fixed time-point virologic outcomes after combination antiretroviral therapy (cART) switch/initiation. However, their relative-hazard for the time to virologic failure has not been thoroughly investigated, and an expert system that is able to predict how long a new cART regimen will remain effective has never been designed.</p> <p>Methods</p> <p>We analyzed patients of the Italian ARCA cohort starting a new cART from 1999 onwards either after virologic failure or as treatment-naïve. The time to virologic failure was the endpoint, from the 90^thday after treatment start, defined as the first HIV-1 RNA > 400 copies/ml, censoring at last available HIV-1 RNA before treatment discontinuation. We assessed the relative hazard/importance of GSSs according to distinct interpretation systems (Rega, ANRS and HIVdb) and other covariates by means of Cox regression and random survival forests (RSF). Prediction models were validated via the bootstrap and c-index measure.</p> <p>Results</p> <p>The dataset included 2337 regimens from 2182 patients, of which 733 were previously treatment-naïve. We observed 1067 virologic failures over 2820 persons-years. Multivariable analysis revealed that low GSSs of cART were independently associated with the hazard of a virologic failure, along with several other covariates. Evaluation of predictive performance yielded a modest ability of the Cox regression to predict the virologic endpoint (c-index≈0.70), while RSF showed a better performance (c-index≈0.73, p < 0.0001 vs. Cox regression). Variable importance according to RSF was concordant with the Cox hazards.</p> <p>Conclusions</p> <p>GSSs of cART and several other covariates were investigated using linear and non-linear survival analysis. RSF models are a promising approach for the development of a reliable system that predicts time to virologic failure better than Cox regression. Such models might represent a significant improvement over the current methods for monitoring and optimization of cART.</p