Advances in primary immunodeficiency diseases in Latin America: epidemiology, research, and perspectives

Primary immunodeficiencies (PIDs) are genetic disorders of the immune system comprising many different phenotypes. Although previously considered rare, recent advances in their clinical, epidemiological, and molecular definitions are revealing how much we still need to learn about them. For example, geographical and ethnic variations as well as the impact of certain practices influence their frequency and presentation, making it necessary to consider their study in terms of regions. The Latin American Society for Immunodeficiencies was established as an organization dedicated to provide scientific support for basic and clinical research and to develop tools and educational resources to promote awareness in the medical community. Initiatives such as these are positively influencing the way PIDs are tackled in these countries, as shown by recent reports and publications. This paper provides a historical compilation and a current view of the many issues faced by scientists studying these diseases in these countries, highlighting the diverse scientific contributions and offering a promising perspective for the further developments in this field in Latin America

Epidemiology Insights

This book represents an overview on the diverse threads of epidemiological research, brings together the expertise and enthusiasm of an international panel of leading researchers to provide a state-of-the art overview of the field. Topics include the epidemiology of dermatomycoses and Candida spp. infections, the epidemiology molecular of methicillin-resistant Staphylococcus aureus (MRSA) isolated from humans and animals, the epidemiology of varied manifestations neuro-psychiatric, virology and epidemiology, epidemiology of wildlife tuberculosis, epidemiologic approaches to the study of microbial quality of milk and milk products, Cox proportional hazards model, epidemiology of lymphoid malignancy, epidemiology of primary immunodeficiency diseases and genetic epidemiology family-based. Written by experts from around the globe, this book is reading for clinicians, researchers and students, who intend to address these issues

Bacterial Meningitis

This timely collection of expert papers draws attention to the global burden of meningitis and the challenges faced by the WHO’s roadmap to defeat meningitis by 2030. The three main goals of the meningitis roadmap are to eliminate epidemics of bacterial meningitis, reduce cases and deaths from vaccine-preventable bacterial meningitis, and reduce disability and improve quality of life after meningitis of any cause. This book includes a wide range of original research and reviews on epidemiology and vaccination of bacterial meningitis that have direct relevance to advancing the goals of the roadmap

Helicobacter pylori in childhood : aspects of prevalence, diagnosis and treatment

In this dissertation we present the results of our research on Helicobacter pylori infections in childhood, focusing on the prevalence, diagnosis and treatment of the infection. Our studies were conducted in the Netherlands, Europe and Indonesia. We discuss diagnostic tests, therapeutic regimens, resistance and preventive measurements. We highlight clinical and pathophysiological aspects of the infection and describe which particular strains are prevalent and how transmission occurs. Presently, there are no established correlations between a Helicobacter pylori infection and recurrent abdominal pain, gastroesophageal reflux disease or growth retardation. We present data on the prevalence of Helicobacter pylori in young infants in the Netherlands and observe that children with at least one non-Dutch parent form a risk group. We assess risk factors in a Europe-wide study on gastroduodenal erosions and ulcers in childhood. In our study, Helicobacter pylori infection and gastrotoxic medications were relatively little implicated as etiology of that pathology. The prevalence of Helicobacter pylori infection in Indonesian young children is relatively high and points at an early acquisition of the infection. Finally, the resistance of Helicobacter pylori to clarithromycin and metronidazole was assessed for adults and children in the Netherlands. Low resistance rates were found, but the resistance in adults is increasing. We conclude that a test-and-treat regimen is justified for the Netherlands.UBL - phd migration 201

Identificação de múltiplos marcadores polimórficos em pneumocystis jirovecii: relação com a evolução clínica

Pneumocystis jirovecii é um importante agente infeccioso, causador de pneumonia (PPc) em doentes com infecção vírus da imunodeficiência humana/síndroma de imunodeficiência adquirida (VIH/sida), tendo sido, também, já descritos casos de infecção em imunocompetentes ou em doentes com graus moderados de imunodeficiência. Sequências polimórficas de ácido desoxirribonucleico (DNA) são detectadas frequentemente em isolados de P. jirovecii, sugerindo que a variação genética é comum neste microrganismo e que diversos subtipos genéticos podem existir. Permanecem por esclarecer as razões pelas quais os doentes sofrem de diferentes graus de severidade de PPc. Alguns estudos apontam para que determinados indicadores do prognóstico clínico possam estar relacionados com a apresentação e a evolução da infecção por P. jirovecii. Por outro lado, também é sugerido que polimorfismos específicos possam determinar características epidemiológicas do microrganismo, como a distribuição geográfica de determinados subtipos, os modos de transmissão, ou mesmo, fenómenos de resistência a fármacos, ou diferentes graus de virulência. Coloca-se a hipótese de que a virulência de um determinado subtipo de P. jirovecii possa estar dependente de múltiplos genes e, portanto, da associação de polimorfismos múltiplos que ocorrem em diversas regiões do genoma. Com o objectivo de identificar genes potencialmente associados com factores da infecção por P. jirovecii e com o propósito de desenvolver técnicas moleculares de alto rendimento, efectuou-se o estudo da variabilidade genética de isolados deste microrganismo, em Portugal. O trabalho envolveu a recolha de espécimes pulmonares de doentes infectados por P. jirovecii com diferentes características clínicas e integrou as metodologias de imunofluorescência indirecta com anticorpos monoclonais (IFI/AcM), reacção de polimerização em cadeia (PCR), sequenciação directa, análise de fragmentos de restrição (RFLP), PCR multiplex, genotipagem por reacção de polimerização de base única (SBE), PCR em tempo real (RT-PCR) e DNA pooling. Os genes candidatos considerados no presente estudo, foram seleccionados com base numa extensa pesquisa bibliográfica: o gene da subunidade grande do rRNA mitocondrial (mtLSU rRNA), o gene do citocromo b (CYB), o gene da superóxido dismutase (SOD), o gene da -tubulina ( TUB), o gene da tiorredoxina reductase (TRR1), o gene da timidilato sintetase (TS), o gene da dihidrofolato reductase (DHFR), o gene da dihidropteroato sintetase (DHPS), a região conservada (UCS) dos genes da família das glicoproteínas major de superfície (MSG) e o gene da protease kexin-like (KEX1). Este procedimento inicial, permitiu avaliar o interesse relativo destes 10 loci, verificar a informação sobre as respectivas sequências, sua variabilidade, importância a nível metabólico e a potencialidade para estas regiões poderem estar relacionadas com características da infecção, como resistência a fármacos, ou factores de virulência. A diversidade genética, as frequências de distribuição de genótipos e a relação entre os genótipos observados e diversos parâmetros da infecção, foram investigadas por PCR seguido de sequenciação directa, ou RFLP. A análise dos resultados permitiu confirmar o declínio das mutações associadas a fenómenos de resistência aos fármacos da família das sulfas, em Portugal, provavelmente como consequência do decréscimo no uso de profilaxia anti-P. jirovecii com sulfas, após a introdução da terapêutica anti-retrovírica potente (HAART). Nos 10 loci de P. jirovecii analisados, foram identificados e caracterizados, um total de 23 polimorfismos, dos quais, quatro (mt85, SOD110, SOD215, DHFR312) demonstraram estar relacionados com parâmetros clínicos da infecção, e, por isso, foram considerados como relevantes no seguimento da investigação. Verificou-se que um genótipo específico (SOD110C/SOD215T) demonstrou estar associado a casos de PPc com maior severidade, enquanto que outros dois genótipos (mt85C/SOD215C e SOD110T/SOD215C) foram associados a casos de PPc com menor severidade. A análise cruzada das sequências permitiu detectar correlações estatisticamente significativas entre diversos polimorfismos, sugerindo a existência de potenciais haplótipos. Observou-se um elevado grau de recombinação entre os genótipos multilocus (MLGs) de P. jirovecii, sendo que, um dos MLGs (MLG E) se encontrava sobre-representado na população, sugerindo uma estrutura epidémica, na qual o fenómeno de recombinação genética é frequente, mas onde ocasionalmente, um clone bem sucedido, tende, rapidamente, a aumentar a sua frequência originando um clone epidémico. A técnica de PCR multiplex/SBE foi aplicada com sucesso na identificação de polimorfismos genéticos de P. jirovecii em pools de DNA, demonstrando que é possível uma análise num formato múltiplo, abrangendo um elevado número de amostras. A metodologia proposta demonstrou ser uma ferramenta útil na caracterização do perfil genético de P. jirovecii, permitindo a amplificação e caracterização de diversos fragmentos, de uma forma rápida, implicando redução de custos e de tempo de operação, e possibilitando, a aplicação em larga escala. A associação das técnicas de PCR multiplex/SBE e de DNA pooling por RT-PCR permitiu determinar a distribuição das frequências relativas dos polimorfismos de base única (SNPs, do inglês single nucleotide polymorphims) DHFR312, mt85, SOD215 e SOD110 em subgrupos de doentes, com diferentes características clínicas. Esta abordagem, possibilitou a identificação de SNPs presentes nos diferentes subgrupos, permitindo a classificação de polimorfismos, aparentemente, não relevantes (ex. DHFR312), e, conseguindo seleccionar outros polimorfismos, potencialmente relacionados com diferentes parâmetros da infecção (ex. mt85, SOD215 e SOD110). Assim, os SNPs seleccionados podem, de alguma forma, estar associados a haplótipos de P. jirovecii com características distintas, ou seja, potencialmente implicados em diferentes graus de severidade da doença. Os resultados obtidos com o presente estudo, permitem colocar a hipótese de que haplótipos de P. jirovecii com características distintas podem existir, e que, regiões genéticas específicas podem funcionar como marcadores desses mesmos organismos. A aplicação das metodologias de alto rendimento, propostas nesta investigação, pode acelerar a identificação de polimorfismos de P. jirovecii relevantes a nível clínico, permitindo uma aproximação baseada em haplótipos para caracterização clínica deste microrganismo patogénico, auxiliando e adequando a escolha do tratamento mais indicado, possibilitando assim, um melhor controlo da doença. A robustez, sensibilidade e especificidade, demonstradas pelas técnicas de PCR multiplex/SBE e de DNA pooling, fazem com que sejam indicadas, não só para o estudo de P. jirovecii, mas também para o estudo de inúmeros microrganismos, em especial de microrganismos não cultiváveis, nos quais, a limitação da amostra, faz com que sejam requeridas técnicas de alta capacidade, para que seja possível a caracterização de isolados, tal como ficou demonstrado, no presente estudo, para o caso de P. jirovecii.Pneumocystis jirovecii pneumonia (PcP) is a common and serious disease in persons infected with the human immunodeficiency virus (HIV). P. jirovecii has also been isolated from immunocompetent persons and patients with moderate degrees of immunodeficiency. Polymorphic desoxiribonucleic acid (DNA) sequences are frequently detected in P. jirovecii isolates suggesting that genetic variation is common in this organism and different genetic subtypes may exist. Several aspects of the disease remain unknown; particularly the reasons why some patients suffer from moderate PcP with favorable response to the anti-P. jirovecii drugs, while others suffer from severe PcP, which could be fatal, despite therapy. While several prognostic indicators have been associated with the outcome of PcP, advances in understanding the genetic diversity of P. jirovecii have shown that specific polymorphisms could determine epidemiological profiles of the pathogen, including geographical distribution, modes of transmission, drug resistance, and virulence of particular subtypes. Data suggests that the virulence of a specific P. jirovecii genetic subtype may be dependent on multiple polymorphisms that occur at several genomic regions. The aims of the present study were to: (i) investigate possible associations between relevant polymorphic sequences and P. jirovecii infection parameters; (ii) develop a novel molecular approach based on new high-throughput methodologies. A description of genetic diversity in P. jirovecii isolates, collected from Portuguese infected patients presenting different clinical parameters, based on the identification of multiple single nucleotide polymorphisms (SNPs), was achieved using indirect immunofluorescence with monoclonal antibodies (IFI/AcM), polymerase chain reaction (PCR), DNA sequencing, restriction fragment length polymorphism analysis (RFLP), multiplex-PCR, single base extension (SBE), real time PCR (RT-PCR), and DNA pooling. The candidate P. jirovecii DNA regions were selected based on literature research. A total of 10 distinct loci were considered the most promising sequences: the mitochondrial large-subunit rRNA (mtLSU rRNA), the cytochrome b (CYB), the superoxide dismutase (SOD), the -tubulin ( TUB), the thioredoxin reductase (TRR1), the thymidylate synthase (TS), the dihydrofolate reductase (DHFR),the dihydropteroate synthase (DHPS), the upstream conserved region (UCS) of the major surface glycoprotein gene (MSG), and the kexin-like serine protease (KEX1). The importance of the candidate loci were evaluated based on the bibliographic research. This initial step provided substantial information about sequence variation, metabolic pathways, and the relationship between these sequences and parameters of infection, such as drug resistance or virulence. The genetic diversity, the frequency distribution of genotypes and the relationship between observed genotypes and different parameters of infection were investigated using PCR followed by DNA sequencing or RFLP. The results confirmed a decline of P. jirovecii mutations associated with resistance to sulfa drugs in Portugal, probably as a result of the decreased use of sulfa prophylaxis after the introduction of the highly active antiretroviral therapy (HAART). Twenty three SNPs were detected and characterized among the 10 loci studied, of which four (mt85, SOD110, SOD215, DHFR312) were demonstrated to be related to clinical parameters of infection, and therefore were considered as relevant sequence variations to be analyzed in the subsequent investigation. It was found that a particular genotype (SOD110C/SOD215T) showed to be associated with more virulent PcP episodes, whereas two other genotypes (mt85C/SOD215C and SOD110T/SOD215C) were associated with less virulent PcP episodes. The study of associated variations and cross-genetic analysis demonstrated significant statistical associations between several SNPs, suggesting the existence of P. jirovecii haplotypes. The results also suggested large recombination between most P. jirovecii multilocus genotypes (MLGs). However, one specific MLG (MLG E) occurred at a higher frequency than would be expected according to panmitic expectations, suggesting linkage disequilibrium and clonal propagation. The persistence of this specific MLG may be a consequence of clonal reproduction of this successful genotypic array in a P. jirovecii population with epidemic structure. Multiplex amplification of genomic DNA associated with single base extension genotyping showed to be a suitable high-throughput methodology for large-scale P. jirovecii SNPs screening. The association of multiplex-PCR/SBE and DNA pooling by RT-PCR showed to be a reliable methodology to estimate P. jirovecii SNPs allele frequencies in P. jirovecii DNA pools. The proposed methodology proved to be a useful tool to study the P. jirovecii genetic profile, providing the characterization of multiplefragments in a cost-saving fast format, in which equal amounts of DNA from a large number of individual samples are pooled and then genotyped. In the present study, the relative frequencies of DHFR312, mt85, SOD215, and SOD110 were calculated in several subgroups of patients with different clinical features. This strategy provided a final selection of effectively important polymorphisms, allowing the exclusion of P. jirovecii polymorphisms apparently without clinical relevance (e.g. DHFR312), and the classification of other polymorphisms, correlated with specific parameters of PcP (e.g. mt85, SOD215 and SOD110). These SNPs may be associated with specific P. jirovecii haplotypes potentially involved with different degrees of virulence in PcP episodes. These results suggest that P. jirovecii haplotypes with distinct characteristics may exist and that specific genetic regions are expected to be suitable molecular markers for epidemiological and clinical applications. The high-throughput methodologies proposed in the present research were designed to accelerate the identification of clinical relevant P. jirovecii polymorphisms. Further application of these techniques will supply a haplotype based approach for clinical characterization of this pathogenic microorganism, leading to tailored treatment choices, and better management of the disease. The robustness and sensitivity shown by multiplex-PCR/SBE and DNA pooling demonstrated that these techniques are indicated for the study of P. jirovecii SNPs with clinical relevance, and for the study of SNPs of other microorganisms, especially those without an in vitro culture system, in which high-sensitivity techniques are required due to sample limitation, as demonstrated in the present study for P. jirovecii

Frameshift mutations at the C-terminus of HIST1H1E result in a specific DNA hypomethylation signature

BACKGROUND: We previously associated HIST1H1E mutations causing Rahman syndrome with a specific genome-wide methylation pattern. RESULTS: Methylome analysis from peripheral blood samples of six affected subjects led us to identify a specific hypomethylated profile. This "episignature" was enriched for genes involved in neuronal system development and function. A computational classifier yielded full sensitivity and specificity in detecting subjects with Rahman syndrome. Applying this model to a cohort of undiagnosed probands allowed us to reach diagnosis in one subject. CONCLUSIONS: We demonstrate an epigenetic signature in subjects with Rahman syndrome that can be used to reach molecular diagnosis

Genetics of Pompe Disease: The secrets within the coding region of the GAA gene and beyond

The main subject addressed in this thesis is the genotype-phenotype relationship in Pompe disease. Pompe disease is a rare, autosomal recessive disorder caused by disease-associated variants (mutations) in the gene coding for the enzyme acid-α-glucosidase (GAA). The function of GAA is to degrade glycogen to glucose in the lysosome. Deficiency of GAA will result in glycogen accumulation in the lysosome, causing damage to the cells and tissues, leading to loss of function. The research described in this thesis could define the best methods and samples of diagnosing Pompe disease enzymatically. Moreover, molecular analysis is an essential step to conclude the diagnostic procedure of a Pompe patient. The results showed the need for additional analyses to detect new disease-associated GAA variants when conventional methods are insufficient. Furthermore, most of the work showed and discussed in this thesis explores the genotype-phenotype correlation in Pompe disease

European position paper on rhinosinusitis and nasal polyps 2020

The European Position Paper on Rhinosinusitis and Nasal Polyps 2020 is the update of similar evidence based position papers published in 2005 and 2007 and 2012. The core objective of the EPOS2020 guideline is to provide revised, up-to-date and clear evidence-based recommendations and integrated care pathways in ARS and CRS. EPOS2020 provides an update on the literature published and studies undertaken in the eight years since the EPOS2012 position paper was published and addresses areas not extensively covered in EPOS2012 such as paediatric CRS and sinus surgery. EPOS2020 also involves new stakeholders, including pharmacists and patients, and addresses new target users who have become more involved in the management and treatment of rhinosinusitis since the publication of the last EPOS document, including pharmacists, nurses, specialised care givers and indeed patients themselves, who employ increasing self-management of their condition using over the counter treatments. The document provides suggestions for future research in this area and offers updated guidance for definitions and outcome measurements in research in different settings. EPOS2020 contains chapters on definitions and classification where we have defined a large number of terms and indicated preferred terms. A new classification of CRS into primary and secondary CRS and further division into localized and diffuse disease, based on anatomic distribution is proposed. There are extensive chapters on epidemiology and predisposing factors, inflammatory mechanisms, (differential) diagnosis of facial pain, allergic rhinitis, genetics, cystic fibrosis, aspirin exacerbated respiratory disease, immunodeficiencies, allergic fungal rhinosinusitis and the relationship between upper and lower airways. The chapters on paediatric acute and chronic rhinosinusitis are totally rewritten. All available evidence for the management of acute rhinosinusitis and chronic rhinosinusitis with or without nasal polyps in adults and children is systematically reviewed and integrated care pathways based on the evidence are proposed. Despite considerable increases in the amount of quality publications in recent years, a large number of practical clinical questions remain. It was agreed that the best way to address these was to conduct a Delphi exercise . The results have been integrated into the respective sections. Last but not least, advice for patients and pharmacists and a new list of research needs are included