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PURPOSE. Caspase-associated recruitment domain-9 (CARD9) deficiency is an inborn error of immunity that typically predisposes otherwise healthy patients to single fungal infections and the occurrence of multiple invasive fungal infections is rare. It has been described as the first known condition that predisposes to extrapulmonary Aspergillus infection with preserved lungs. We present a patient that expands the clinical variability of CARD9 deficiency. MATERIALS AND METHODS. Genetic analysis was performed by Sanger sequencing. Neutrophils and mononuclear phagocyte response to fungal stimulation were evaluated through luminol-enhanced chemiluminescence and whole blood production of the proinflammatory mediator IL-6 respectively. RESULTS. We report a 56-year-old Argentinean woman, whose invasive Exophiala spinifera infection at the age of 32 years was unexplained and reported in year 2004. At the age of 49 years, she presented with chronic pulmonary disease due to Aspergillus nomius. After partial improvement following treatment with caspofungin and posaconazole, right pulmonary bilobectomy was performed. Despite administration of multiple courses of antifungals, sustained clinical remission could not be achieved. We recently found that the patient’s blood showed an impaired production of interleukin (IL)-6 when stimulated with zymosan. We also found that she is homozygous for a previously reported CARD9 loss-of-function mutation (Q289*). CONCLUSIONS. This is the first report of a patient with inherited CARD9 deficiency and chronic invasive pulmonary aspergillosis (IPA) due to A. nomius. Inherited CARD9 deficiency should be considered in otherwise healthy children and adults with one or more invasive fungal diseases.

Several exon skipping antisense oligonucleotides (eteplirsen, golodirsen, viltolarsen, and casimersen) have been approved for the treatment of Duchenne muscular dystrophy, but many more are in development targeting an array of different DMD exons. Preclinical screening of the new oligonucleotide sequences is routinely performed using patient-derived cell cultures, and evaluation of their efficacy may be performed at RNA and/or protein level. While several methods to assess exon skipping and dystrophin expression in cell culture have been developed, the choice of methodology often depends on the availability of specific research equipment.In this chapter, we describe and indicate the relevant bibliography of all the methods that may be used in this evaluation and describe in detail the protocols routinely followed at our institution, one to evaluate the efficacy of skipping at RNA level (nested PCR) and the other the restoration of protein expression (myoblot), which provide good results using equipment largely available to most research laboratories.

Contains fulltext : 248204.pdf (Publisher’s version ) (Open Access) Nucleic acid therapeutics is a growing field aiming to treat human conditions that has gained special attention due to the successful development of mRNA vaccines against SARS-CoV-2. Another type of nucleic acid therapeutics is antisense oligonucleotides, versatile tools that can be used in multiple ways to target pre-mRNA and mRNA. While some years ago these molecules were just considered a useful research tool and a curiosity in the clinical market, this has rapidly changed. These molecules are promising strategies for personalized treatments for rare genetic diseases and they are in development for very common disorders too. In this chapter, we provide a brief description of the different mechanisms of action of these RNA therapeutic molecules, with clear examples at preclinical and clinical stages.

Electroencephalographic (EEG) alpha oscillations have been related to heart rate variability (HRV) and both change in Alzheimer's disease (AD). We explored if task switching reveals altered alpha power and HRV in cognitively healthy individuals with AD pathology in cerebrospinal fluid (CSF) and whether HRV improves the AD pathology classification by alpha power alone. We compared low and high alpha event-related desynchronization (ERD) and HRV parameters during task switch testing between two groups of cognitively healthy participants classified by CSF amyloid/tau ratio: normal (CH-NAT, n=19) or pathological (CH-PAT, n=27). For the task switching paradigm, participants were required to name the color or word for each colored word stimulus, with two sequential stimuli per trial. Trials include color (cC) or word (wW) repeats with low load repeating, and word (cW) or color switch (wC) for high load switching. HRV was assessed for RR interval, standard deviation of RR-intervals (SDNN) and root mean squared successive differences (RMSSD) in time domain, and low frequency (LF), high frequency (HF), and LF/HF ratio in frequency domain. Results showed that CH-PATs compared to CH-NATs presented: 1) increased (less negative) low alpha ERD during low load repeat trials and lower word switch cost (low alpha: p=0.008, Cohen's d=-0.83, 95% confidence interval -1.44 to -0.22, and high alpha: p=0.019, Cohen's d=-0.73, 95% confidence interval -1.34 to -0.13); 2) decreasing HRV from rest to task, suggesting hyper-activated sympatho-vagal responses. 3) CH-PATs classification by alpha ERD was improved by supplementing HRV signatures, supporting a potentially compromised brain-heart interoceptive regulation in CH-PATs. Further experiments are needed to validate these findings for clinical significance.

Utrophin is an autosomal paralogue of dystrophin, a protein whose deficit causes Duchenne and Becker muscular dystrophies (DMD/BMD). Utrophin is naturally overexpressed at the sarcolemma of mature dystrophin-deficient fibres in DMD and BMD patients as well as in the mdx Duchenne mouse model. Dystrophin and utrophin can co-localise in human foetal muscle, in the dystrophin-competent fibres from DMD/BMD carriers, and revertant fibre clusters in biopsies from DMD patients. These findings suggest that utrophin overexpression could act as a surrogate, compensating for the lack of dystrophin, and, as such, it could be used in combination with dystrophin restoration therapies. Different strategies to overexpress utrophin are currently under investigation. In recent years, many compounds have been reported to modulate utrophin expression efficiently in preclinical studies and ameliorate the dystrophic phenotype in animal models of the disease. In this manuscript, we discuss the current knowledge on utrophin protein and the different mechanisms that modulate its expression in skeletal muscle. We also include a comprehensive review of compounds proposed as utrophin regulators and, as such, potential therapeutic candidates for these muscular dystrophies. This work was supported by funding from Health Institute Carlos III (ISCIII, Spain) and the European Regional Development Fund, (ERDF/FEDER), `A way of making Europe': Grant PI15/00333; Basque Government (grants 2016111029, 2018222035 and 2020333012) and Duchenne Parent Project Spain (grant 05/2016). P. S--M holds a Rio Hortega Fellowship from ISCIII (CM19/00104). V.A--G holds a Miguel Servet Fellowship from the ISCIII (CPII17/00004), part-funded by ERDF/FEDER. A. L--M acknowledges funding by Biocruces Bizkaia Health Research Institute (BC/I/DIV/19/001). V. A--G also acknowledges funding from Ikerbasque (Basque Foundation for Science). None of this funding represents a conflict of interest with the content of this review.

Gene editing methods are an attractive therapeutic option for Duchenne muscular dystrophy, and they have an immediate application in the generation of research models. To generate myoblast cultures that could be useful in in vitro drug screening, we have optimised a CRISPR/Cas9 gene edition protocol. We have successfully used it in wild type immortalised myoblasts to delete exon 52 of the dystrophin gene, modelling a common Duchenne muscular dystrophy mutation; and in patient’s immortalised cultures we have deleted an inhibitory microRNA target region of the utrophin UTR, leading to utrophin upregulation. We have characterised these cultures by demonstrating, respectively, inhibition of dystrophin expression and overexpression of utrophin, and evaluating the expression of myogenic factors (Myf5 and MyH3) and components of the dystrophin associated glycoprotein complex (α-sarcoglycan and β-dystroglycan). To demonstrate their use in the assessment of DMD treatments, we have performed exon skipping on the DMDΔ52-Model and have used the unedited DMD cultures/ DMD-UTRN-Model combo to assess utrophin overexpression after drug treatment. While the practical use of DMDΔ52-Model is limited to the validation to our gene editing protocol, DMD-UTRN-Model presents a possible therapeutic gene edition target as well as a useful positive control in the screening of utrophin overexpression drugs.

This study aimed to estimate the number of incident cases and hospital admissions attributable to secondhand smoke (SHS) exposure at home for asthma, otitis media (OM), and lower respiratory infections (LRI) in children in Spain. The burden of respiratory disease caused by SHS exposure was estimated in terms of incident cases and hospitalized cases for asthma, OM, and LRI. Estimates were calculated using the population attributable fraction. The age-specific (0-1 year, 0-4 years, 5-11 years, and 0-11 years) prevalence of SHS exposure in children was estimated through a telephone survey performed in a representative sample of Spanish households with children in 2016. The risk estimates for all diseases were selected from international meta-analyses. The number of hospitalized cases was obtained for each disease from the Hospital Minimum Data Set provided by the Ministry of Health of Spain. Incident cases were obtained from the Global Health Data Exchange. In 2015, SHS exposure caused an estimated total of 136,403 incident cases of the following respiratory diseases: 9058 (8.5%) cases of asthma, 120,248 (8.5%) of OM, and 7097 (13.5%) of LRI in children aged 0–14 years old in Spain. Likewise, SHS exposure caused a total of 3028 hospitalized cases, with 379 (8.5%) for asthma and 167 (8.5%) for OM in children 0–11 years old, and 2482 (11.6%) for LRI in children

Background This study aims to identify sociodemographic characteristics associated with secondhand smoke (SHS) exposure and the adoption of smoking bans in homes with children in Spain Methods We performed, in 2016, a cross-sectional study to a representative sample of Spanish households with children under 12 years old. We administered a telephone survey to the parents asking about smoking patterns at home, children’s SHS exposure and sociodemographic characteristics. Poisson regression models with robust variance were built to assess sociodemographic characteristics associated with household SHS exposure and the adoption of smoking rules. Results In this study participated 2411 families, 25.8% of which reported exposure at home and 84.4% implemented smoking bans. SHS exposure was associated with having one (aPR = 2.09; 95% CI: 1.43–3.04) or two Spanish parents (aPR = 1.71; 95% CI: 1.24–2.36), lower educational attainment (primary: aPR = 1.74; 95% CI: 1.45–2.10; secondary: aPR = 1.37; 95% CI: 1.17–1.60 compared with university studies), a family structure different from two-parent family (aPR = 1.38; 95% CI: 1.14–1.67) and parents between 31 and 40 years (aPR = 0.75; 95% CI: 0.57–0.99) and 41–50 years (aPR = 0.62; 95% CI: 0.47–0.81) compared with 18- to 30-year-old parents. The adoption of smoking bans was associated with two-parent family (aPR = 1.09; 95% CI: 1.01–1.17), living with non-smokers (aPR = 1.46; 95% CI: 1.31–1.62), parents of foreign origin (aPR = 1.09; 95% CI: 1.04–1.14) and younger children (0–3 years: aPR = 1.05; 95% CI: 1.01–1.09) compared with the oldest children (8–11 years). Conclusion The parent’s origin and the family structure were associated with SHS exposure and the adoption of smoking bans at home. Moreover, the number of smokers living at home was relevant for the adoption of smoking bans, and the educational attainment for SHS exposure. These factors should be taken into consideration when designing or implementing smoke-free home programmes.

Paracoccidioidomycosis (Pm) is a systemic disease, endemic in the American continent. There are two different clinical forms, the infant-juvenile or subacute form (PmS) and the chronic adult form (PmC). The human immunodeficiency virus (HIV) associated paracoccidioidomycosis (PmHIV) shares characteristics with both of the previously mentioned forms. The objective of this work was to describe the epidemiological, clinical and laboratory features of the PmHIV and to compare them with the ones of PmS and the PmC. A retrospective analysis of 119 patients with paracoccidioidomycosis was performed. Ninety four suffered the chronic form, 11 the subacute one and 14 were coinfected with HIV. Patients with PmHIV presented a CD4+ T lymphocytes median of 70.5 cells/μl, 71.4% had fever, 64.3% had a miliary pattern on the chest radiography, 64.3% had hepatosplenomegaly, 64.3% had mucosal lesions and 50% had skin lesions. One patient died during his hospitalization. The clinical presentation of Pm in patients with HIV resembled the subacute form with fever, hepatomegaly and skin lesions. However, they also tended to present mucosal lesions, positive serology for Pm and pulmonary parenchyma lesions as usually seen in PmC (9/14 PmHIV patients had overlapping features, while 4/14 PmHIV patients clinically resembled PmS and 1/14 PmC). The incidence of Pm has not changed with the burden of AIDS as it has happened with other fungal infections but it appears clinically different from the classic clinical forms of the disease.

Since the gene responsible for Duchenne muscular dystrophy was first described in 1987 [1], knowledge of neuromuscular diseases (NMD) has evolved rapidly. [...]

Despite the international guidelines on the containment of the coronavirus disease 2019 (COVID-19) pandemic, the European scientific community was not sufficiently prepared to coordinate scientific efforts. To improve preparedness for future pandemics, we have initiated a network of nine European-funded Cooperation in Science and Technology (COST) Actions that can help facilitate inter-, multi-, and trans-disciplinary communication and collaboration., M.H.V. has grants from FEDER (Fundo Europeu de Desenvolvimento Regional) through COMPETE 2020 and from FCT (Foundation for Science and Technology; POCI-01-0145-FEDER-030457 ; POCI-01-0145-FEDER-016390 : CANCEL STEM). V.A-G. holds a Miguel Servet Fellowship from the ISCIII (CPII17/00004), part-funded by ERDF/FEDER , and acknowledges funding from Ikerbasque (Basque Foundation for Science). T.B. has grants from Carraresi Foundation , EU Horizon 2020 research and innovation programme No 828835 FetOpen and MIND Food’s HUB-Regione Lombardia. J.D.L.R. acknowledges funding from ISCIII (project PI18/00591 ) cofounded by the EU FEDER funds. D.P-S. has grants RTI2018-097624-B-I00 ( AEI, MCI, Spain, and ERDF ), RD16/0006/0021 ( ISCIII, Spain, and ERDF ), and CSIC PTI Global Health ( PIE 202020E223/CSIC-COV19-100 ). H.H.H.W.S. has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 777111 (REPO-TRIAL). I.S. acknowledges funding from FCT (Fundação para a Ciência e Tecnologia), Portugal. C.R. is supported by AIRC (Italian Association for Cancer Research; IG21408 ). C.R. has funding from PharmaMar . Y.D. acknowledges funding from the National Research Fund (grants C14/BM/8225223 , C17/BM/11613033 , and COVID-19/2020-1/14719577/miRCOVID ), the Ministry of Higher Education and Research, and the Heart Foundation – Daniel Wagner of Luxembourg. P.T. acknowledges the support of H2020 project iPC “individualized Paediatric Cure” ( 826121 ).

Background Recurrent vulvovaginitis is a growing problem that affects millions of women worldwide. In many cases it is treated as vulvovaginal candidiasis, but there is not always microbiological confirmation. Aims To determine the etiology of vulvovaginitis in a group of patients. Methods This is a cross-sectional study in which the data from the medical records of 316 adult patients who consulted for vulvovaginitis were analyzed. Eighty nine percent of the cases had already suffered previous episodes. Results The median age was 34 (265 patients were between 16 and 45 years old). Yeasts were isolated in culture from 211 (66.8%) patients, although pseudo-hyphae and yeasts were observed in only 166 samples (52.5%) in the direct microscopic examination. Multiple predisposing factors were found, among which the use of contraceptives or previous antibiotics stand out. Most of the patients (almost 90%) had been treated with antifungals, with or without microbiological confirmation. Candida albicans was isolated in 187 (88.6%) patients, followed by Candida glabrata in 6 (2.8%) patients. Association with bacterial vaginosis was found in 35.1% and with intermediate bacterial microbiota in 33.2% of the cases. A remarkably high proportion of C. albicans isolates resistant to fluconazole (80.1%) and itraconazole (58.8%) was found. Conclusions A microbiological analysis is essential to confirm the diagnosis of vulvovaginal candidiasis, whether simple, complicated, or recurrent. Identifying the isolated yeast species and determining its susceptibility to antifungal agents are particularly important.

Paracoccidioidomycosis (PCM) is a life-threatening systemic mycosis widely reported in the Gran Chaco ecosystem. The disease is caused by different species from the genus Paracoccidioides, which are all endemic to South and Central America. Here, we sequenced and analyzed 31 isolates of Paracoccidioides across South America, with particular focus on isolates from Argentina and Paraguay. The de novo sequenced isolates were compared with publicly available genomes. Phylogenetics and population genomics revealed that PCM in Argentina and Paraguay is caused by three distinct Paracoccidioides genotypes, P. brasiliensis (S1a and S1b) and P. restrepiensis (PS3). P. brasiliensis S1a isolates from Argentina are frequently associated with chronic forms of the disease. Our results suggest the existence of extensive molecular polymorphism among Paracoccidioides species, and provide a framework to begin to dissect the connection between genotypic differences in the pathogen and the clinical outcomes of the disease. Fil: Teixeira, Marcus de Melo. Universidade do Brasília; Brasil Fil: Cattana, Maria Emilia. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste; Argentina Fil: Matute, Daniel R.. University of North Carolina; Estados Unidos Fil: Muñoz, José F.. Broad Institute Of Mit And Harvard; Estados Unidos Fil: Arechavala, Alicia. Hospital Francisco J Muñiz; Argentina Fil: Isbell, Kristin. University of North Carolina; Estados Unidos Fil: Schipper, Rafael. Universidade do Brasília; Brasil Fil: Santiso, Gabriela Maria. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; Argentina Fil: Tracogna, Fernanda. Gobierno de la Provincia de Chaco. Hospital Julio Cecilio Perrando.; Argentina Fil: Sosa, María de los Ángeles. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina Fil: Cech, Norma. Hospital 4 de Junio; Argentina Fil: Alvarado, Primavera. Instituto de Biomedicina Dr. Jacinto Convit; Venezuela Fil: Barreto, Laura. Instituto Superior de Formación Docente Salome Ureña; República Dominicana Fil: Chacón, Yone. Provincia de Salta. Ministerio de Salud Pública. Hospital del Milagro; Argentina Fil: Ortellado, Juana. Universidad Nacional de Asunción; Paraguay Fil: Lima, Cleoni Mendes de. Universidade Federal de Rondonia; Brasil Fil: Chang, Marilene Rodrigues. Universidade Federal do Mato Grosso do Sul; Brasil Fil: Niño Vega, Gustavo. Universidad de Guanajuato; México Fil: Yasuda, Maria Aparecida Shikanai. Universidade de Sao Paulo; Brasil Fil: Felipe, Maria Sueli Soares. Universidade Catolica de Brasilia; Brasil Fil: Negroni, Ricardo. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; Argentina Fil: Cuomo, Christina A.. Broad Institute of MIT And Harvard; Estados Unidos Fil: Barker, Bridget. Tgen Northern Arizona University; Estados Unidos Fil: Giusiano, Gustavo Emilio. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina. Universidad Nacional del Nordeste. Facultad de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste; Argentina

A descriptive observational and cross-sectional study was carried out. The clinical characteristics, etiologic agents, treatments and outcome of 33 cases of tinea capitis in the Mycology Unit at Francisco J. Muñiz Hospital of Buenos Aires City between January 2015 and December 2019 were analyzed. The median age of the patients was 7 years, 21 of whom were male, 3 were HIV-positive and 22 had pets. The isolated etiologic agents were the following: Microsporum canis in 22 cases, Trichophyton tonsurans in 8, Nannizzia gypsea in 2 and Trichophyton mentagrophytes in one patient. Suppurative tinea capitis (krion Celsi) was detected in 10 cases and the same number of patients presented other skin locations of their dermatophytosis in addition to those in the scalp. Twenty-one cases were orally treated with griseofulvin and 12 with terbinafine. Those patients with suppurative tinea capitis received drops of betamethasone by mouth besides the antifungal drugs. All patients had good clinical and mycological response to the treatments, all lesions disappeared, and mycological studies turned negative by the end of the treatments. We conclude that both drugs were effective for the treatment of tinea capitis; however, lesions in those cases receiving terbinafine involuted more slowly.

Myotonic dystrophy type I (DM1) is the most common form of adult muscular dystrophy, caused by expansion of a CTG triplet repeat in the 3′ untranslated region (3′UTR) of the myotonic dystrophy protein kinase (DMPK) gene. The pathological CTG repeats result in protein trapping by expanded transcripts, a decreased DMPK translation and the disruption of the chromatin structure, affecting neighboring genes expression. The muscleblind-like (MBNL) and CUG-BP and ETR-3-like factors (CELF) are two families of tissue-specific regulators of developmentally programmed alternative splicing that act as antagonist regulators of several pre-mRNA targets, including troponin 2 (TNNT2), insulin receptor (INSR), chloride channel 1 (CLCN1) and MBNL2. Sequestration of MBNL proteins and up-regulation of CELF1 are key to DM1 pathology, inducing a spliceopathy that leads to a developmental remodelling of the transcriptome due to an adult-to-foetal splicing switch, which results in the loss of cell function and viability. Moreover, recent studies indicate that additional pathogenic mechanisms may also contribute to disease pathology, including a misregulation of cellular mRNA translation, localization and stability. This review focuses on the cause and effects of MBNL and CELF1 deregulation in DM1, describing the molecular mechanisms underlying alternative splicing misregulation for a deeper understanding of DM1 complexity. To contribute to this analysis, we have prepared a comprehensive list of transcript alterations involved in DM1 pathogenesis, as well as other deregulated mRNA processing pathways implications.

IntroductionExposure to secondhand aerosol from e-cigarette (SHA) may pose harmful effects to bystanders. This study aims to investigate the prevalence, duration and determinants of SHA exposure in various indoor settings in 12 European countries.MethodsIn 2017–2018, we conducted a cross-sectional study, the TackSHS survey, on a representative sample of the population aged ≥15 years in 12 European countries (Bulgaria, England, France, Germany, Greece, Ireland, Italy, Latvia, Poland, Portugal, Romania and Spain). We described the prevalence and duration of exposure to SHA in several indoor settings among 11 604 e-cigarette non-users. Individual-level and country-level characteristics associated with SHA exposure were also explored using multilevel logistic regression analyses.ResultsOverall, 16.0% of e-cigarette non-users were exposed to SHA in any indoor setting at least weekly, ranging from 4.3% in Spain to 29.6% in England. The median duration of SHA exposure among those who were exposed was 43 min/day. ‘Other indoor settings’ (eg, bar and restaurant) was reported as the place where most of e-cigarette non-users were exposed (8.3%), followed by workplace/educational venues (6.4%), home (5.8%), public transportation (3.5%) and private transportation (2.7%). SHA exposure was more likely to occur in certain groups of non-users: men, younger age groups, those with higher level of education, e-cigarette past users, current smokers, those perceiving SHA harmless and living in countries with a higher e-cigarette use prevalence.ConclusionsWe found inequalities of SHA exposure across and within European countries. Governments should consider extending their tobacco smoke-free legislation to e-cigarettes to protect bystanders, particularly vulnerable populations such as young people.Trial registration numberNCT02928536.

Our aim was to provide a systematic review of studies on the burden of disease due to second-hand smoke (SHS) exposure, reviewing methods, exposure assessment, diseases causally linked to SHS, health outcomes, and estimates available to date. A literature review of studies on the burden of disease from SHS exposure, available in PubMed and SCOPUS, published 2007–2018 in English language, was carried out following the PRISMA recommendations. Overall, 588 studies were first identified, and 94 were eligible. Seventy-two studies were included in the systematic review. Most of them were based on the comparative risk assessment approach, assessing SHS exposure using mainly surveys on exposure at home/workplaces. Diseases more frequently studied were: lung cancer, ischemic heart disease, stroke, chronic obstructive pulmonary disease, asthma and breast cancer in adults; lower respiratory tract infection, otitis media, asthma, sudden infant death syndrome and low birth weight in children. The SHS exposure assessment and the reported population attributable fractions (PAF) were largely heterogeneous. As an example, the PAF from lung cancer varied between 0.6% and 20.5%. Moreover, PAF were estimated applying relative risks and SHS exposures with no consistent definitions or with different age classes. The research gap on the SHS exposure burden is shrinking. However, estimates are not yet available for a number of countries, particularly the Middle Eastern and African countries, and not all diseases with the strongest evidence of causation, such as sudden infant death syndrome, have been explored. Moreover, in some cases the applied methodology revealed relatively low quality of data. info:eu-repo/semantics/acceptedVersion

Smoke-free legislation reduced second-hand smoke (SHS) exposure in public places, and indirectly promoted private smoke-free settings. Nevertheless, a large proportion of adults is still exposed to SHS at home. The aim of this paper is to quantify the burden of disease due to home SHS exposure among adults in the 28-European Union (EU) countries for year 2017. The burdens by gender from lung cancer, chronic obstructive pulmonary disease (COPD), breast cancer, ischemic heart disease (IHD), stroke, asthma, and diabetes were estimated in an original research analysis using the comparative risk assessment method. Relative risks of death/diseases by gender for adults exposed to SHS at home compared to not exposed ones were estimated updating existing meta-analyses. Prevalence of home SHS exposure by gender was estimated using a multiple imputation procedure based on Eurobarometer surveys. Data on mortality and disability adjusted life years (DALYs) were obtained from the Global Burden of Disease, Injuries and Risk Factors Study. In 2017, 526,000 DALYs (0.36% of total DALYs) and 24,000 deaths (0.46% of total deaths) were attributable to home SHS exposure in the 28-EU countries, mainly from COPD and IHD. South-Eastern EU countries showed the highest burden, with proportion of DALYs/deaths attributable to SHS exposure on total higher than 0.50%/0.70%, whereas northern EU-countries showed the lowest burden, with proportions of DALYs/deaths lower than 0.25%/0.34%. The burden from SHS exposure is still significant in EU countries. More could be done to raise awareness of the health risks associated with SHS exposure at home.

The current study examined symptoms of irritation reported by non-smokers passively exposed to e-cigarette aerosols and their timing and association with the concentrations of volatile organic compounds (VOCs) produced.40 healthy non-smoking adults were exposed to e-cigarette aerosols for 30 min in a 35 m20 males and 20 females, with a mean age of 24.6 years (SD = 4.3) and exhaled CO 7 ppm participated. PM concentrations in both experimental sessions were higher than the Control (p 0.001). The most commonly reported symptoms were burning, dryness, sore throat, cough, breathlessness and headache. During both experimental sessions, ocular, nasal, throat-respiratory symptoms and general complaints increased significantly (p 0.05). Ocular and nasal symptoms returned to baseline by tA 30-min exposure to SHA provoked symptoms of sensory irritation and general complaints that lasted up to 30 min after the exposure and were positively associated with the concentrations of the VOC mixture emitted.

A 21 year-old man, HIV infected, and with poor adherence to antiretroviral treatment, consulted us due to a papular rash on trunk and extremities, showing simultaneously hypochromic stains on his back. He was afebrile and hemodynamically stable. In the mycological study of the clinical samples taken from different lesions, yeasts compatible with Malassezia were observed. Oral itraconazole was prescribed, with an excellent clinical response. Finding the same etiolologic agent in 2different clinical pictures on a single patient is extremely rare.

Background: Duchenne muscular dystrophy is a lethal disease caused by lack of dystrophin. Skipping of exons adjacent to out-of-frame deletions has proven to restore dystrophin expression in Duchenne patients. Exon 51 has been the most studied target in both preclinical and clinical settings and the availability of standardized procedures to quantify exon skipping would be advantageous for the evaluation of preclinical and clinical data. Objective: To compare methods currently used to quantify antisense oligonucleotide–induced exon 51 skipping in the DMD transcript and to provide guidance about the method to use. Methods: Six laboratories shared blinded RNA samples from Duchenne patient-derived muscle cells treated with different amounts of exon 51 targeting antisense oligonucleotide. Exon 51 skipping levels were quantified using five different techniques: digital droplet PCR, single PCR assessed with Agilent bioanalyzer, nested PCR with agarose gel image analysis by either ImageJ or GeneTools software and quantitative real-time PCR. Results: Differences in mean exon skipping levels and dispersion around the mean were observed across the different techniques. Results obtained by digital droplet PCR were reproducible and showed the smallest dispersion. Exon skipping quantification with the other methods showed overestimation of exon skipping or high data variation. Conclusions: Our results suggest that digital droplet PCR was the most precise and quantitative method. The quantification of exon 51 skipping by Agilent bioanalyzer after a single round of PCR was the second-best choice with a 2.3-fold overestimation of exon 51 skipping levels compared to digital droplet PCR.

This study aimed to estimate the association between second-hand smoke (SHS) exposure in children and asthma, wheezing and perceived health.A cross-sectional study based on a telephone survey was performed on a representative sample of 2411 children under 12 years old in Spain. Exposure to SHS in private and public settings, and the prevalence of asthma, wheezing and perceived poor health were described. The association between health indicators and SHS exposure was analyzed using multivariate Poisson regression models with robust variance according to age and educational level.The prevalence of SHS exposure in children was 29.2% in private settings and 42.5% in public settings. There was no association between SHS exposure and asthma, wheezing and perceived poor health in children ≤5 years. In children aged 6-11 years with parents with primary/secondary education, presenting asthma (adjusted prevalence ratio [aPR]: 2.1; 95% confidence interval [95%CI]: 1.2-3.8) and worse perceived health (aPR: 1.6; 95%CI: 1.1-2.1) were positively associated with SHS exposure in private settings. In children with parents with university studies, a negative association between SHS exposure and asthma (aPR: .3; 95%CI: 0.1-0.7) and wheezing (aPR: 0.3; 95%CI: 0.1-0.8) was observed.There are differences in the association between SHS exposure and asthma, wheezing and poor perceived health according to educational level. Interventions with an equity perspective aimed at reducing SHS exposure in childhood should be implemented.

Cryptococcosis is a life-threatening fungal infection caused by the encapsulated yeasts Cryptococcus neoformans and C. gattii, acquired from the environment. In Latin America, as occurring worldwide, C. neoformans causes more than 90% of the cases of cryptococcosis, affecting predominantly patients with HIV, while C. gattii generally affects otherwise healthy individuals. In this region, cryptococcal meningitis is the most common presentation, with amphotericin B and fluconazole being the antifungal drugs of choice. Avian droppings are the predominant environmental reservoir of C. neoformans, while C. gattii is associated with several arboreal species. Importantly, C. gattii has a high prevalence in Latin America and has been proposed to be the likely origin of some C. gattii populations in North America. Thus, in the recent years, significant progress has been made with the study of the basic biology and laboratory identification of cryptococcal strains, in understanding their ecology, population genetics, host-pathogen interactions, and the clinical epidemiology of this important mycosis in Latin America. Fil: Firacative, Carolina. University of Sydney; Australia Fil: Lizarazo, Jairo. Universidad de Pamplona; España Fil: Illnait Zaragozí, María Teresa. Tropical Medicine Institute Pedro Kourí; Cuba Fil: Castañeda, Maria Elizabeth. Instituto Nacional de Salud; Colombia. Latin American Cryptococcal Study Group; Brasil Fil: Arechavala, Alicia. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; Argentina. Latin American Cryptococcal Study Group; Brasil Fil: Córdoba, Susana Beatríz. Latin American Cryptococcal Study Group; Brasil. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C. G. Malbrán”; Argentina Fil: Mazza, Mariana. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C. G. Malbrán”; Argentina. Latin American Cryptococcal Study Group; Brasil Fil: Taverna, Constanza Giselle. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C. G. Malbrán”; Argentina. Latin American Cryptococcal Study Group; Brasil Fil: Isla, Guillermina. Latin American Cryptococcal Study Group; Brasil. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C. G. Malbrán”; Argentina Fil: Chiapello, Laura Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina. Latin American Cryptococcal Study Group; Brasil Fil: Vergara, Mario León Silva. Universidade Federal do Triangulo Mineiro; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Melhem, Marcia S. C.. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Szeszs, Maria Walderez. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Martins, Marilena dos Anjos. Latin American Cryptococcal Study Group; Brasil. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil Fil: Bonfietti, Lucas Xavier. Latin American Cryptococcal Study Group; Brasil. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil Fil: Oliveira, Rogério Antonio de. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Oliveira, Lidiane de. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Santos, Dayane Christine Silva. Latin American Cryptococcal Study Group; Brasil. Governo do Estado de São Paulo. Secretaria da Saúde. Instituto Adolfo Lutz; Brasil Fil: Lazera, Marcia S.. Latin American Cryptococcal Study Group; Brasil. Fundación Oswaldo Cruz; Brasil Fil: Wanke, Bodo. Fundación Oswaldo Cruz; Brasil. Latin American Cryptococcal Study Group; Brasil Fil: Díaz, María Cristina. Latin American Cryptococcal Study Group; Brasil. Universidad de Chile; Chile Fil: Escandón, Patricia. Instituto Nacional de Salud; Colombia. Latin American Cryptococcal Study Group; Brasil Fil: Noguera, María Clara. Latin American Cryptococcal Study Group; Brasil. Universidad Metropolitana; Colombia Fil: Andreu, Carlos Manuel Fernández. Latin American Cryptococcal Study Group; Brasil Fil: Castril­Lón, Laura. Universidad Nacional Autónoma de México; México. Latin American Cryptococcal Study Group; Brasil Fil: Bustamante, Beatriz. Hospital Cayetano Heredia. Instituto de Medicina Tropical Alexander von Humboldt; Perú. Hospital Cayetano Heredia; Perú. Latin American Cryptococcal Study Group; Brasil Fil: Dolande, Maribel. Universidad Central de Venezuela; Venezuela. Latin American Cryptococcal Study Group; Brasil Fil: Ferrara, Giussepe. Universidad Central de Venezuela; Venezuela. Latin American Cryptococcal Study Group; Brasil

Background The introduction of € smoke-free laws' has reduced the population's exposure to second-hand smoke (SHS), although SHS is still an issue in homes and other public places. Children are vulnerable to its health effects, and their greatest exposure occurs at home. Objectives To assess airborne nicotine concentration of the living room and children's bedroom of homes with children under 13 years of age, and to analyse factors associated with these levels. Methods We conducted a cross-sectional study in Barcelona in 2015-2016, selecting a convenience sample from families with at least one child under 13 years of age. The sample comprised 50 families with smokers and 50 without. We measured airborne nicotine concentrations in the living room and children's bedroom, and, using a questionnaire administered to the parents, collected information about smoking habits at home. Results Homes without smokers showed nicotine concentrations below the limit of detection (

Cryptococcosis is a life-threatening fungal infection caused by the encapsulated yeasts Cryptococcus neoformans and C. gattii, acquired from the environment. In Latin America, as occurring worldwide, C. neoformans causes more than 90% of the cases of cryptococcosis, affecting predominantly patients with HIV, while C. gattii generally affects otherwise healthy individuals. In this region, cryptococcal meningitis is the most common presentation, with amphotericin B and fluconazole being the antifungal drugs of choice. Avian droppings are the predominant environmental reservoir of C. neoformans, while C. gattii is associated with several arboreal species. Importantly, C. gattii has a high prevalence in Latin America and has been proposed to be the likely origin of some C. gattii populations in North America. Thus, in the recent years, significant progress has been made with the study of the basic biology and laboratory identification of cryptococcal strains, in understanding their ecology, population genetics, host-pathogen interactions, and the clinical epidemiology of this important mycosis in Latin America.

Histoplasmosis is a mycosis with a high prevalence in HIV/AIDS patients. Clinical presentation includes a wide spectrum of manifestations and diagnosis usually takes up to several weeks in patients who do not present cutaneous lesions.To determine the clinical and microbiological characteristics as well as some biochemical parameters in patients with AIDS-associated histoplasmosis without tegumentary lesions, in order to develop a guideline which enables an early empiric treatment in cases of difficult diagnosis.Medical records of 86 patients with histoplasmosis were reviewed; 31 patients with diagnosis of AIDS-associated histoplasmosis without cutaneous lesions were analyzed.Fever was the most frequent symptom (96.7%), lung involvement was observed in 22 patients (70.9%), the most commonly radiological pattern was miliary pattern [(12/22), 54.5%]. Nineteen patients presented with splenomegaly. Blood culture sensitivity was 93.3% (28/30) and serology was positive only in 23.5% of the cases. Eight patients died (25.8%). Patients in which CD4+ T cell lymphocytes count was50 cells/μl, albumin levels2.5 g/dl and who presented with pancytopenia had an unfavorable outcome.In HIV seropositive patients with fever associated to splenomegaly and bilateral miliar pattern in chest radiography, the empiric treatment with amphotericin B must be considered if signs and symptoms of unfavorable outcome are present and due to the time that it takes to arrive at an accurate diagnosis. In order to confirm the diagnosis, all microbiological samples should be collected prior to initiating therapy.

Aims New therapies for neuromuscular disorders are often mutation specific and require to be studied in patient's cell cultures. In Duchenne muscular dystrophy (DMD) dystrophin restoration drugs are being developed but as muscle cell cultures from DMD patients are scarce and do not grow or differentiate well, only a limited number of candidate drugs are tested. Moreover, dystrophin quantification by western blotting requires a large number of cultured cells; so fewer compounds are as thoroughly screened as is desirable. We aimed to develop a quantitative assessment tool using fewer cells to contribute in the study of dystrophin and to identify better drug candidates. Methods An 'in-cell western' assay is a quantitative immunofluorescence assay performed in cell culture microplates that allows protein quantification directly in culture, allowing a higher number of experimental repeats and throughput. We have optimized the assay ('myoblot') to be applied to the study of differentiated myoblast cultures. Results After an exhaustive optimization of the technique to adapt it to the growth and differentiation rates of our cultures and the low intrinsic expression of our proteins of interests, our myoblot protocol allows the quantification of dystrophin and other muscle-associated proteins in muscle cell cultures. We are able to distinguish accurately between the different sets of patients based on their dystrophin expression and detect dystrophin restoration after treatment. Conclusions We expect that this new tool to quantify muscle proteins in DMD and other muscle disorders will aid in their diagnosis and in the development of new therapies.

BackgroundThe introduction of ‘smoke-free laws’ has reduced the population’s exposure to second-hand smoke (SHS), although SHS is still an issue in homes and other public places. Children are vulnerable to its health effects, and their greatest exposure occurs at home.ObjectivesTo assess airborne nicotine concentration of the living room and children’s bedroom of homes with children under 13 years of age, and to analyse factors associated with these levels.MethodsWe conducted a cross-sectional study in Barcelona in 2015–2016, selecting a convenience sample from families with at least one child under 13 years of age. The sample comprised 50 families with smokers and 50 without. We measured airborne nicotine concentrations in the living room and children’s bedroom, and, using a questionnaire administered to the parents, collected information about smoking habits at home.ResultsHomes without smokers showed nicotine concentrations below the limit of detection (3), while those with at least one smoker showed 0.16 µg/m3 in the living room and 0.12 µg/m3 in the bedroom. When smoking was allowed inside home, these values increased to 1.04 and 0.48 µg/m3, respectively. Moreover, nicotine concentrations in both rooms were strongly correlated (r=0.89), and higher nicotine levels were associated with the number of cigarettes smoked in the living room, smoking rules, the number of smokers living at home and tobacco smell.ConclusionsHomes with smokers present SHS in the living room and in the children’s bedroom. Therefore, programmes focused on reducing children’s SHS exposure are urgently needed.

A 48 year-old immunocompetent woman, who had a nodular lesion in the neck and a dense infiltrate at the lower lobe of the left lung, presented at the Mycology Unit of Muñiz Hospital of Buenos Aires City. The pulmonary infiltrate disappeared spontaneously 3 months later. The histopathological study of the nodular lesion showed capsulated yeasts (mucicarmin and alcian blue positive stains) compatible with Cryptococcus. The mycological study of a new sample, obtained by a nodular puncture, allowed the isolation of yeasts, identified as Cryptococcus gattii (VGII). Latex test for Cryptococcus capsular antigen in serum was positive (1/100). CSF cultures rendered negative results. Fluconazole at a daily dose of 800mg was given during 45 days with partial improvement; as cultures from a new clinical sample were positive for Cryptococcus, the antimycotic was changed to itraconazole 400mg/day for 5 months, with an excellent clinical response.

The case of a 60 year old woman with hemoptysis and a thin-walled cavitary lesion at the upper lobe of the right lung is presented. The woman presented at the Mycology Unit of the Muñiz Hospital in Buenos Aires City 3 months after the beginning of her clinical manifestations. A hyaline micelial fungus with chlamido-arthroconidias was isolated from the bronchoalveolar lavage. Immunodiffusion and counter-immnunoelectrophoresis with coccidioidin and histoplasmin rendered positive results against both antigents, and skin tests with coccidioidin and histoplasmin were also positive with strong reactions. The isolated fungus was identified as Coccidioides posadasii at the National Microbiology Institute Carlos Malbrán, by means of a molecular technique. The patient was treated with itraconazole by oral route at a daily dose of 200mg with good clinical response, but due to the persistence of the lung cavity, a surgical removal of the upper lobe of the right lung had to be scheduled.

The use of splice-switching antisense therapy is highly promising, with a wealth of pre-clinical data and numerous clinical trials ongoing. Nevertheless, its potential to treat a variety of disorders has yet to be realized. The main obstacle impeding the clinical translation of this approach is the relatively poor delivery of antisense oligonucleotides to target tissues after systemic delivery. We are a group of researchers closely involved in the development of these therapies and would like to communicate our discussions concerning the validity of standard methodologies currently used in their pre-clinical development, the gaps in current knowledge and the pertinent challenges facing the field. We therefore make recommendations in order to focus future research efforts and facilitate a wider application of therapeutic antisense oligonucleotides., Fundación Ramón Areces and from Spanish Ministry of Economy and Competitiveness and European Regional Development Fund [grant reference SAF2013-43005-R]. B.S. was supported by a grant from the Tromsø Research Foundation. F.P.R. was supported by the Association Française contre les Myopathies (AFM-téléthon). M.A.D. was supported by Telethon Italia [grant reference GGP08244] and Italian Ministry of Health [grant reference GR-2008-1136933 and RF-2011-02347694]. S.L. was supported by The Dutch Duchenne Parent Project NL (DPP NL) and Association Française contre les Myopathies (AFM-téléthon). G.N.G. holds a Miguel Servet Fellowship from the ISCIII (grant reference CP14/00032] and a Fondo de Investigaciones Sanitarias Grant [grant reference PI15/01756], both are partfunded by the European Regional Development Fund (ERDF/FEDER). V.S. was supported by SKIP-NMD project, a European Community’s Seventh

A retrospective study was carried out on 171 cases of disseminated histoplasmosis diagnosed in HIV/AIDS patients during the period 2009-2014. Although HIV diagnosis rates remained stable over the study period, a sensible increase in the number of histoplasmosis cases was observed in the last three years. Disseminated histoplasmosis was prevalent in males with an average age of 37.8 years. At diagnosis, only 54/171 (31.6%) were receiving HAART, and CD4+ T-lymphocyte counts ranged from 4 to 264 cells/upsilon. Cutaneous lesions, including ulcerated papules or molluscoid plaques, were present in 110/171 (64.3%), with Histoplasma capsulatum being observed in all skin scraping specimens upon Giemsa staining. Respiratory manifestations were second in frequency with bronchoalveolar lavage showing a high diagnostic performance. Radiological findings included milliary patterns, interstitial infiltrates, and focalized condensations. Out of 141 blood cultures performed, H. capsulatum was isolated in 82 (58.2%). No significant difference in diagnostic performance was found between blood cultures and skin scraping (p = 0.6164). Other opportunistic infections were observed in 70/171 (40.9%) prior to or concomitantly with histoplasmosis. Association with Mycobacterium tuberculosis was recorded in 16/171 (9.4%) and one had a multi-drug resistant isolate. The severity of histoplasmosis determined the monotherapy with amphotericin B deoxycholate in 115 (67.3%), itraconazole in 42 (24.5%), and combined therapies in 14 (8.2%). Mortality was 19.9% (34/171). Finally, we emphasize that the higher prevalence in the last three years of the study should prompt the medical community to consider the diagnosis of histoplasmosis to reduce mortality of AIDS patients.

Background Cryptococcosis is still a life-threatening mycosis that continues to be of serious concern in Latin American countries, especially among HIV+positive population. However, there is not any reliable information about the prevalence of this disease in this region. Aims The aim of this study is to report data of 2041 patients with cryptococcosis that were attended at the Infectious Diseases Hospital F. J. Muniz over a 30 year-period. Methods Information about demographic and clinical data, survival time and the applied treatment, was taken from the Mycology Unit database. Mycological exams from different clinical samples were performed. Cryptococcal capsular antigen in serum and cerebrospinal fluid was detected through the latex agglutination technique. Cryptococcus isolates were phenotypically identified and the genotype was determined in some of them. Susceptibility tests were carried out following M27-A3 document. Results Seventy five percent of HIV+positive patients and 50% of the HIV-negative population were males. Mean ages were 34.1 in HIV+positive patients and 44.8 in the HIV-negative. Cryptococcosis was associated with AIDS in 98% of the cases. Meningeal compromise was seen in 90% of the patients. Although cerebrospinal fluid rendered more positive results, blood culture was the first diagnostic finding in some cases. Cryptococcal antigen showed positive results in 96.2% of the sera samples and in the 93.1% of the cerebrospinal fluid samples. Most of the isolates were Cryptococcus neoformans and belonged to genotype VNI. Minimal inhibitory concentration values were mostly below the epidemiological cutoff values. Conclusions We observed that thanks to a high level of clinical suspicion, early diagnosis, combined therapy and intracranial pressure control by daily lumbar punctures, the global mortality rate has markedly decreased through the years in the analyzed period.

Summary Duchenne muscular dystrophy is a rare, progressive, muscle-wasting disease leading to severe disability and premature death. Treatment is currently symptomatic, but several experimental therapies are in development. Implemented care standards, validated outcome measures correlating with clinical benefit, and comprehensive information about the natural history of the disease are essential for regulatory approval of any treatment. However, for Duchenne muscular dystrophy and other rare diseases, these requirements are not always in place when potential therapies enter the clinical trial phase. A cooperative effort of stakeholders in Duchenne muscular dystrophy—including representatives from patients' groups, academia, industry, and regulatory agencies—is aimed at addressing this shortfall by identifying strategies to overcome challenges, developing the tools needed, and collecting relevant data. An open and constructive dialogue among European stakeholders has positively affected development of treatments for Duchenne muscular dystrophy; this approach could serve as a paradigm for development of treatments for rare diseases in general.

Classic histoplasmosis is a systemic endemic mycosis due to Histoplasma capsulatum var. capsulatum. A significant reduction in the morbidity and mortality of AIDS-related histoplasmosis has been observed since the introduction of highly active antiretroviral therapy (HAART) and secondary antifungal prophylaxis.The aim of this study was to determine the current state of prognosis and treatment response of HIV-positive patients with histoplasmosis in the Francisco J. Muñiz Infectious Diseases Hospital in Buenos Aires City.A retrospective study was conducted using the demographic, clinical, immunological and treatment data of 80 patients suffering from AIDS-related histoplasmosis.Of the 80 cases studied 65 were male, the median age was 36 years, with 73.7% of the patients being drug addicts, 82.5% of the patients was not receiving HAART at diagnosis, and 58.7% of the cases had less than 50 CD4+ cells/μl at the beginning of the treatment. The initial phase of treatment consisted of intravenous amphotericin B and/or oral itraconazole for 3 months, with 78.7% of the cases showing a good clinical response. Only 26/63 patients who were discharged from hospital continued with the follow-up of the HAART, secondary prophylaxis with itraconazole or amphotericin B. Secondary prophylaxis was stopped after more than one year of HAART if the patients were asymptomatic, had two CDThe treatment of histoplasmosis in HIV-positive patients was effective in 78.8% of the cases. The combination of HAART and secondary antifungal prophylaxis is safe, well tolerated, and effective. The low adherence of patients to HAART and the lack of laboratory kits for rapid histoplasmosis diagnosis should be addressed in the future. The usefulness of primary antifungal prophylaxis for cryptococcosis and histoplasmosis HIV-positive patients should be studied.

Objectives: To describe food consumption frequency in adolescents in the context of the financial crisis in 2012, and to analyse potential fluctuations in excess body weight between 2008 and 2012. Method: A cross-sectional study of eating habits and excess body weight was conducted in adolescents aged 13 to 19 years old from public, subsidised and private secondary schools in Barcelona, Spain. The FRESC lifestyle risk factors survey was used, and food frequency consumption, food recommendations and body mass index were analysed according to gender, year of education and socioeconomic status. Results: Girls ate vegetables and fruits more frequently than boys, while the prevalence of junk food consumption was higher in boys. The prevalence of compliance with food recommendations was lower than 50% for all foods, and gender and socioeconomic differences were found for eggs, red meat and soft drinks. Regarding excess body weight, boys had a higher prevalence than girls in the 2 years analysed. Furthermore, a reduction in excess body weight was observed among girls in secondary education in the highest socioeconomic groups (28.7% [95% CI: 24.8-32.6%] in 2008 to 20.5% [95% CI: 17.1-23.8%] in 2012). Conclusions: The prevalence of adolescents following food recommendations is low, and gender differences were found in terms of food consumption frequency, even in the context of financial crisis. There is a need to promote programmes and policies to reduce inequalities related to eating habits and excess body weight in adolescents. Resumen: Objetivos: Describir la frecuencia de consumo de alimentos en adolescentes en un contexto de crisis económica en el año 2012, y analizar los potenciales cambios en el exceso de peso entre los años 2008 y 2012. Diseño: Estudio transversal de los hábitos alimentarios y el exceso de peso en adolescentes de entre 13 y 19 años de edad pertenecientes a escuelas públicas, concertadas y privadas de Barcelona. Se utilizó la encuesta FRESC sobre factores de riesgo asociados a estilos de vida y se analizaron, según sexo, curso académico y nivel socioeconómico, las frecuencias de consumo, recomendaciones alimentarias e índice de masa corporal. Resultados: Las chicas realizaban un mayor consumo de frutas y verduras, mientras que los chicos consumían comida no saludable con mayor frecuencia. La prevalencia de cumplimiento de las recomendaciones alimentarias es inferior al 50% para todos los alimentos, y se encontraron diferencias según el sexo y el nivel socioeconómico en el consumo de huevos, carne roja y refrescos. En relación al exceso de peso, la prevalencia fue mayor en los chicos que en las chicas en los 2 años analizados. Además, se observó una reducción del exceso de peso en las chicas cursando Educación Secundaria Obligatoria y de nivel socioeconómico alto (de 28,7% [IC95%: 24,8-32,6%] en 2008 a 20,5% [IC95%: 17,1-23,8%] en 2012). Conclusiones: La prevalencia de adolescentes que siguen las recomendaciones de consumo es baja. Hay diferencias de sexo en las frecuencias de consumo de los alimentos, incluso en un contexto de crisis económica. Existe una necesidad de promover programas y políticas para reducir las desigualdades relacionadas con los hábitos alimentarios y el exceso de peso en los adolescentes. Keywords: Adolescent, Food consumption, Overweight, Economic recession, Diet surveys, Palabras clave: Adolescente, Ingestión de alimentos, Sobrepeso, Recesión económica, Encuestas sobre dietas

Duchenne muscular dystrophy (DMD) is the most common childhood neuromuscular disorder. It is caused by mutations in the DMD gene that disrupt the open reading frame (ORF) preventing the production of functional dystrophin protein. The loss of dystrophin ultimately leads to the degeneration of muscle fibres, progressive weakness and premature death. Antisense oligonucleotides (AOs) targeted to splicing elements within DMD pre-mRNA can induce the skipping of targeted exons, restoring the ORF and the consequent production of a shorter but functional dystrophin protein. This approach may lead to an effective disease modifying treatment for DMD and progress towards clinical application has been rapid. Less than a decade has passed between the first studies published in 1998 describing the use of AOs to modify the DMD gene in mice and the results of the first intramuscular proof of concept clinical trials. Whilst phase II and III trials are now underway, the heterogeneity of DMD mutations, efficient systemic delivery and targeting of AOs to cardiac muscle remain significant challenges. Here we review the current status of AO-mediated therapy for DMD, discussing the preclinical, clinical and regulatory hurdles and their possible solutions to expedite the translation of AO-mediated exon skipping therapy to clinic.

The aim of this study was to develop and characterize antigens for the diagnosis of aspergillosis. Nine strains of Aspergillus species Aspergillus fumigatus , Aspergillus flavus , and Aspergillus niger were grown in Sabouraud and Smith broth to produce exoantigens. The antigens were tested by immunodiffusion against sera from patients with aspergillosis and other systemic mycoses. The protein fraction of the antigens was detected by SDS–PAGE; Western blot and representative bands were assessed by mass spectrometry coupled to a nano Acquity UltraPerformance LC and analyzed by the Mascot search engine. Concurrently, all sera were tested with Platelia Aspergillus EIA. The most reactive antigens to sera from patients infected by A. fumigatus were produced by A. fumigatus MG2 Sabouraud and pooled A. fumigatus Sabouraud samples, both with a sensitivity of 93% and specificity of 100% and 97%, respectively. Aspergillus niger and A. flavus antigens were reactive against A. niger and A. flavus sera, each one with a sensitivity and specificity of 100%. Two proteins, probably responsible for antigenic activity, β-glucosidase in A. fumigatus and α-amylase in A. niger were attained. The commercial kit had a specificity of 22%, sensitivity of 100%, positive predictive value of 48%, and negative predictive value of 100%. The antigens produced showed high sensitivity and specificity and can be exploited for diagnostics of aspergilloma.

We previously conducted a proof of principle; dose escalation study in Duchenne muscular dystrophy (DMD) patients using the morpholino splice-switching oligonucleotide AVI-4658 (eteplirsen) that induces skipping of dystrophin exon 51 in patients with relevant deletions, restores the open reading frame and induces dystrophin protein expression after intramuscular (i.m.) injection. We now show that this dystrophin expression was accompanied by an elevated expression of α-sarcoglycan, β-dystroglycan (BDG) and--in relevant cases--neuronal nitric oxide synthase (nNOS) at the sarcolemma, each of which is a component of a different subcomplex of the dystrophin-associated glycoprotein complex (DAPC). As expected, nNOS expression was relocalized to the sarcolemma in Duchenne patients in whom the dystrophin deletion left the nNOS-binding domain (exons 42-45) intact, whereas this did not occur in patients with deletions that involved this domain. Our results indicate that the novel internally deleted and shorter dystrophin induced by skipping exon 51 in patients with amenable deletions, can also restore the dystrophin-associated complex, further suggesting preserved functionality of the newly translated dystrophin.

Summary Background We report clinical safety and biochemical efficacy from a dose-ranging study of intravenously administered AVI-4658 phosphorodiamidate morpholino oligomer (PMO) in patients with Duchenne muscular dystrophy. Method We undertook an open-label, phase 2, dose-escalation study (0·5, 1·0, 2·0, 4·0, 10·0, and 20·0 mg/kg bodyweight) in ambulant patients with Duchenne muscular dystrophy aged 5–15 years with amenable deletions in DMD. Participants had a muscle biopsy before starting treatment and after 12 weekly intravenous infusions of AVI-4658. The primary study objective was to assess safety and tolerability of AVI-4658. The secondary objectives were pharmacokinetic properties and the ability of AVI-4658 to induce exon 51 skipping and dystrophin restoration by RT-PCR, immunohistochemistry, and immunoblotting. The study is registered, number NCT00844597. Findings 19 patients took part in the study. AVI-4658 was well tolerated with no drug-related serious adverse events. AVI-4658 induced exon 51 skipping in all cohorts and new dystrophin protein expression in a significant dose-dependent (p=0·0203), but variable, manner in boys from cohort 3 (dose 2 mg/kg) onwards. Seven patients responded to treatment, in whom mean dystrophin fluorescence intensity increased from 8·9% (95% CI 7·1–10·6) to 16·4% (10·8–22·0) of normal control after treatment (p=0·0287). The three patients with the greatest responses to treatment had 21%, 15%, and 55% dystrophin-positive fibres after treatment and these findings were confirmed with western blot, which showed an increase after treatment of protein levels from 2% to 18%, from 0·9% to 17%, and from 0% to 7·7% of normal muscle, respectively. The dystrophin-associated proteins α-sarcoglycan and neuronal nitric oxide synthase were also restored at the sarcolemma. Analysis of the inflammatory infiltrate indicated a reduction of cytotoxic T cells in the post-treatment muscle biopsies in the two high-dose cohorts. Interpretation The safety and biochemical efficacy that we present show the potential of AVI-4658 to become a disease-modifying drug for Duchenne muscular dystrophy. Funding UK Medical Research Council; AVI BioPharma.

Cryptococcus neoformans causes disseminated infection in 7-8% of HIV positive patients admitted to Hospital F. J. Muñiz in Buenos Aires. Meningoencephalitis is the most frequent clinical manifestation and is one of the main causes of death in those patients with AIDS. The standard treatment for this mycosis consists of amphotericin B followed by fluconazole until two successive cultures of CFS are negative. Although resistance to these drugs is infrequent, minimal inhibitory concentrations (MIC) of some antifungals can be high. Since it is important to know the susceptibility levels of this fungus to the antifungal drugs usually employed in our institution, we analyzed the susceptibility test results of C. neoformans with two diffusion methods (Etest and NeoSensitabs tablets) employing Mueller-Hinton agar with 2% glucose and 0.5 microg/ml methylene blue. These results were compared with MICs obtained through the use of the broth microdilution reference method (CLSI). Results showed good agreement with the reference method, with no very major errors and only two major errors for fluconazole using NeoSensitabs tablets. For all the above mentioned, we confirm the usefulness of Mueller-Hinton agar to evaluate C. neoformans susceptibility to amphotericin B and fluconazole with these two agar diffusion methods.

Background: Aspergillus fumigatus can cause a wide variety of clinical syndromes, especially in the three largest immunocompromised groups, such as HIV-infected patients. Primary renal aspergillosis is an extremely rare entity. Aims: We report an unusual case of renal abscess due to Aspergillus fumigatus in a patient with AIDS. Methods: We review clinical and laboratory records, and provide follow up of the patient. Results: A 38-year-old man, HIV seropositive, was admitted to our hospital with fever, lumbar pain and respiratory symptoms. Abdominal ultrasound and computerised tomography showed a single and large lesion consistent with an abscess located in the left kidney. Aspergillus fumigatus was isolated from clinical sample obtained by ultrasound-guided needle aspiration. Despite a correct treatment based on amphotericin B and drainage of the abscess, surgery was necessary and nephrectomy was carried out. Histopathological examination of the surgical specimen confirmed the diagnosis of renal aspergillosis. Systemic antifungal therapy based on intravenous and oral voriconazole and highly active antiretroviral therapy was started after surgery. The patient had a good response to the established treatment and he remains in a good clinical condition at one year of follow up. Conclusions: Combined medical and surgical treatment is the elective therapy for renal abscesses due to Aspergillus when percutaneous drainage and the administration of systemic antifungal drugs, such as amphotericin B and/or oral voriconazole or itraconazole, fail. This case emphasizes renal fungal infections should be included in the differential diagnosis of kidney abscesses in AIDS patients.

Duchenne muscular dystrophy (DMD) is characterised by the absence of dystrophin in muscle biopsies, although residual dystrophin can be present, either as dystrophin-positive (revertant) fibres or traces. As restoration of dystrophin expression is the end point of clinical trials, such residual dystrophin is a key factor in recruitment of patients and may also confound the analysis of dystrophin restoration in treated patients, if, as previously observed in the mdx mouse, revertant fibres increase with age. In 62% of the diagnostic biopsies reports of 65 DMD patients studied, traces or revertants were recorded with no correlation between traces or revertants, the patients' performance, or corticosteroids response. In nine of these patients, there was no increase in traces or revertants in biopsies taken a mean of 8.23 years (5.8-10.4 years) after the original diagnostic biopsy. This information should help in the design and execution of clinical trials focused on dystrophin restoration strategies. (C) 2010 Elsevier B.V. All rights reserved.

Stem cell therapy holds promise for treating muscle diseases. Although satellite cells regenerate skeletal muscle, they only have a local effect after intra-muscular transplantation. Alternative cell types, more easily obtainable and systemically-deliverable, were therefore sought. Human synovial stem cells (hSSCs) have been reported to regenerate muscle fibres and reconstitute the satellite cell pool. We therefore determined if these cells are able to regenerate skeletal muscle after intra-muscular injection into cryodamaged muscles of Rag2-/γ chain-/C5-mice. We found that hSSCs possess only limited capacity to undergo myogenic differentiation in vitro or to contribute to muscle regeneration in vivo. However, this is enhanced by over-expression of human MyoD1. Interestingly, hSSCs express extracellular matrix components laminin α2 and collagen VI within grafted muscles. Therefore, despite their limited capacity to regenerate skeletal muscle, hSSCs could play a role in treating muscular dystrophies secondary to defects in extracellular matrix proteins.