Bayesian Methods in Brain Connectivity Change Point Detection with EEG Data and Genetic Algorithm

Human brain is processing a great amount of information everyday, and our brain regions are organized optimally for this information processing. There have been increasing number of studies focusing on functional or effective connectivity in human brain regions in the last decade. In this dissertation, Bayesian methods in Brain connectivity change point detection are discussed. First, a review of state-of-the-art Bayesian-inference-based methods applied to functional magnetic resonance imaging (fMRI) data is carried out, three methods are reviewed and compared. Second, the Bayesian connectivity change point model is extended to change point analysis in electroencephalogram (EEG) data, and the ability of EEG measures of frontal and temporo-parietal activity during mindfulness therapy to track response to dysfunctional anxiety patients\u27 treatment is tested successfully. Then an optimized method for Bayesian connectivity change point model with genetic algorithm (GA) is proposed and proved to be more efficient in change point detection. And due to the good parallel performance of GA, the change point detection method can be parallelized in GPU or multi-processor computers as a future work. Furthermore, a more advanced Bayesian bi-cluster connectivity change point model is developed to simultaneously detect change point of each subject within a group, and cluster subjects into different groups according to their change point distribution and connectivity dynamics. The method is also validated on experimental datasets. After discussing brain change point detection, a review of Bayesian analysis of complex mutations in HBV HCV and HIV studies is also included as part of my Ph.D. work. Finally, conclusions are drawn and future work is discussed

Bayesian analysis of complex mutations in HBV, HCV, and HIV studies

This article provides a review of the Bayesian-inference-based methods applied to Hepatitis B Virus (HBV), Hepatitis C Virus (HCV), and Human Immunodeficiency Virus (HIV) studies with a focus on the detection of the viral mutations and various problems which are correlated to these mutations. The authors also provide a summary of the Bayesian methods' applications toward these viruses' studies, where several important and useful results have been discovered

Evolutionary history and phylogeography of the hepatitis C and hepatitis B visuses in Portugal

Em 2019, a Organização Mundial de Saúde estimou 354 milhões de pessoas com hepatite B e C crónica a nível mundial. Em Portugal, o mais recente inquérito transversal a nível nacional parece revelar uma diminuição das prevalências de VHB e VHC anteriormente estimadas. A informação sobre a evolução da diversidade viral continua limitada, desconhecendo-se se a epidemiologia molecular das infeções por VHB e VHC tem vindo a ser alterada em Portugal. O objetivo deste estudo foi investigar a história evolutiva de ambos os vírus e a sua filogeografia em Portugal. Os ácidos nucleicos virais foram extraídos do plasma de doentes portugueses com VHB e VHC, os genes pol do VHB e NS5A e NS5B do VHC foram amplificados e sequenciados. A reconstrução de árvores filogenéticas de máxima verossimilhança no software IQtree v1.6.11 foi usada para genotipar as estirpes virais. A história epidémica dos subtipos do VHC em Portugal foi reconstruída através de métodos Bayesianos, conforme implementados no software BEAST v1.10.4, tal como a investigação da origem e das rotas de dispersão de ambos os vírus. As resistências aos antivirais nos dois clades do subtipo 1a do VHC foram analisadas online no Geno2Pheno [HCV] 0.92. Os nossos dados revelaram que o subgenótipo D4 do VHB foi o primeiro a ser introduzido em Portugal cerca de 1857, seguido pelo D3 e A2 algumas décadas mais tarde. Os genótipos E e A1 do VHB foram introduzidos posteriormente, quase em simultâneo. Os nossos resultados revelaram um papel muito importante de Portugal na exportação de D4 e A2 para o Brasil e Cabo Verde, respetivamente, no início do século XX. Em Portugal foram identificados subtipos distintos de VHC que entraram no país ao longo do tempo: subtipo 1b (1930-1960), subtipos 3a (1960s) e 1a (1980s), possivelmente associados a transfusões de sangue contaminado, ao início do uso de drogas intravenosas e ao seu uso generalizado, respetivamente. Os subtipos 4a e 4d, emergiram mais recentemente, possivelmente com o ressurgimento do uso de opiáceos. O subtipo 1a é claramente o mais frequente em Portugal apresentando dois clades diferentes (I e II) a circular na população, que possivelmente possuem vias de transmissão diferentes. Os primeiros países a introduzir os clades I e II em Portugal foram os Estados Unidos da América (1965) e Espanha (1955), respetivamente. Dois subclades, classificados como X e Y, foram identificados entre as estirpes do clade I. As RAS basais no gene NS5A foram encontradas principalmente nas estirpes do clade I/subclade Y, sendo a mutação mais frequente a L31M, que se revelou ausente nas estirpes do clade I/subclade X e do clade II. Este trabalho permitiu conhecer a história epidemiológica do VHB e VHC em Portugal, mostrando que Portugal teve um papel importante na dispersão global do VHB e fornecendo novos conhecimentos sobre a epidemiologia molecular, origem e dinâmica de dispersão do VHC em Portugal. Também indicou algumas estirpes virais de VHC como mais propensas a adquirir RAS e que a resistência aos antivirais deve ser investigada no contexto dos clades/subclades do subtipo 1a.In 2019, the World Health Organization estimated 354 million people with chronic hepatitis B and C worldwide. In Portugal, the most recent national cross-sectional survey indicates a decrease in previously estimated HBV and HCV prevalence. Information on the evolution of viral diversity remains limited, and it is not known whether the molecular epidemiological profile of HBV and HCV infections has been changing in Portugal. The aim of this study was to investigate the molecular epidemiology and evolutionary history of both viruses and their phylogeography in Portugal. Viral nucleic acids were extracted from the plasma of Portuguese patients infected with HBV and HCV, the pol gene of HBV and NS5A and NS5B genes of HCV were amplified and sequenced. The reconstruction of maximum likelihood phylogenetic trees in the IQtree v1.6.11 software was used for genotyping purposes. The epidemic history of HCV subtypes in Portugal was reconstructed using Bayesian methods, as implemented in the BEAST v1.10.4 software, as in the investigation of the origin and routes of spread of both viruses. Antiviral drug resistance in the two HCV subtype 1a clades were analyzed online in Geno2Pheno [HCV] 0.92. Our data indicated that the D4 subgenotype of HBV was the first to be introduced in Portugal around 1857, followed by D3 and A2 a few decades later. HBV genotypes E and A1 were introduced later, almost simultaneously. Our results revealed a very important role of Portugal in the exportation of D4 and A2 to Brazil and Cape Verde, respectively, in the beginning of the 20th century. Distinct HCV subtypes that entered Portugal over time were identified: subtype 1b (1930- 1960), subtypes 3a (1960s) and 1a (1980s), possibly associated with transfusions of contaminated blood, with the beginning of intravenous drug use and its widespread use, respectively. Subtypes 4a and 4d have emerged more recently, possibly with the resurgence of opiate use. Subtype 1a is clearly the most frequent in Portugal with two different clades (I and II) circulating in the population, which possibly have different transmission routes. The first countries to introduce clades I and II in Portugal were the United States of America Spain (1965) and Spain (1955), respectively. Two subclades, classified as X and Y, were identified among the clade I strains. The basal RAS in the NS5A gene were found mainly in the clade I/subclade Y strains, the most frequent mutation being L31M, which was absent in the strains of clade I/subclade X and clade II. This work allowed us to understand the epidemiological history of HBV and HCV in Portugal, showing that Portugal played an important role in the global spread of HBV and providing new knowledge about the molecular epidemiology, origin and dispersion dynamics of HCV in Portugal. It also highlighted that some viral strains of HCV may be more likely to acquire RAS and that antiviral resistance should be further investigated in the context of subtype 1a clades/subclades

Novel HBsAg markers tightly correlate with occult HBV infection and strongly affect HBsAg detection.

Occult HBV infection (OBI) is a threat for the safety of blood-supply, and has been associated with the onset of HBV-related hepatocellular carcinoma and lymphomagenesis. Nevertheless, genetic markers in HBsAg (particularly in D-genotype, the most common in Europe) significantly associated with OBI in vivo are missing. Thus, the goal of this study is to define: (i) prevalence and clinical profile of OBI among blood-donors; (ii) HBsAg-mutations associated with OBI; (iii) their impact on HBsAg-detection. OBI was searched among 422,278 blood-donors screened by Nucleic-Acid-Testing. Following Taormina-OBI-definition, 26 (0.006%) OBI-patients were identified. Despite viremia <50IU/ml, HBsAg-sequences were obtained for 25/26 patients (24/25 genotype-D). OBI-associated mutations were identified by comparing OBI-HBsAg with that of 82 chronically-infected (genotype-D) patients as control. Twenty HBsAg-mutations significantly correlated for the first time with OBI. By structural analysis, they localized in the major HBV B-cell-epitope, and in HBsAg-capsid interaction region. 14/24 OBI-patients (58.8%) carried in median 3 such mutations (IQR:2.0-6.0) against 0 in chronically-infected patients. By co-variation analysis, correlations were observed for R122P+S167L (phi=0.68, P=0.01), T116N+S143L (phi=0.53, P=0.03), and Y100S+S143L (phi=0.67, p<0.001). Mutants (obtained by site-directed mutagenesis) carrying T116N, T116N+S143L, R122P, R122P+Q101R, or R122P+S167L strongly decreased HBsAg-reactivity (54.9±22.6S/CO, 31.2±12.0S/CO, 6.1±2.4S/CO, 3.0±1.0S/CO and 3.9±1.3S/CO, respectively) compared to wild-type (306.8±64.1S/CO). Even more, Y100S and Y100S+S143L supernatants show no detectable-HBsAg (experiments in quadruplicate). In conclusions, unique HBsAg-mutations in genotype-D, different than those described in genotypes B/C (rarely found in western countries), tightly correlate with OBI, and strongly affect HBsAg-detection. By altering HBV-antigenicity and/or viral-particle maturation, they may affect full-reliability of universal diagnostic-assays for HBsAg-detection

Statistische Analyse von Sequenzpopulationen in der Virologie und Immunologie

In this thesis I have examined various topics regarding the relationship between viruses and the human immune system. I expanded and refined a tool (which can now be found as R-package SeqFeatR on C-RAN) for the analysis of sequence data and features of this sequences like HLA type or tropism (see chapter 4) and checked with this tool if there are differences between some multiple correction approaches for sequence data, and how Bayesian inference could be used in this context (see chapter 5). It could be shown that Bayesian inference is superior to the frequentistic methods for this kind of problem, because multiple correction approaches ignore the fact that different positions in a sequence alignment may be connected in the protein product of this sequence and are therefor not independent. Furthermore, I have examined sequences from HCV with a form of bootstrap algorithm to find sequence areas which can be used in unknown transmission cases in court. Two areas were found, one in the hypervariable region and the other at the end of the non-structural protein NS5B (see chapter 9). Proteasomal cleavage of alien amino acid sequences inside human cells leads to a presentation of fragments of these sequences on the surface of the cell as epitopes. To present such a fragment, not only must it bind to the MHC, but also needs to be in the correct length to be presented. Therefore viral evolution should favor those viruses, which cannot be cut into presentable epitopes. With epitope data from IEDB and predicted viral sequences which bind the MHC, I searched for amino acids inside the flanking regions around the epitope that may indicate a possible escape mutation against the proteasomal cleavage processes. Fourteen such amino acids and positions were found (see chapter 7). I created a model of HBV reverse transcriptase to check if mutations in certain positions could influence binding with the nucleotide analogue reverse transcriptase inhibitor Tenofovir. Mutations which were inside the binding pocket for Tenofovir showed, in an experimental design by the group of Mengji Lu, a decreased affinity towards the drug (see chapter 10). Together with Ralf Küppers group I examined NGS from different types of B cells to search for almost identical sequences between those. We found similar to identical sequences from two, three and even four kinds of cells in the blood samples of both donors (see chapter 6).In dieser Dissertation bearbeitete ich verschiedene Themen aus dem Bereich der humanpatho-genen Viren und des menschlichen Immunsystems. Zu diesem Zweck entwarf ich ein Programm (welches auf dem R-Archiv C-RAN unter dem Namen SeqFeatR zu finden ist) mit dem sich der Zusammenhang zwischen Sequenzdaten und spezifischen Eigenschaften, wie etwa HLA Typ oder Tropismus, analysieren läßt (s.h Kapitel 4). Mit diesem Programm untersuchte ich ob ein Unterschied zwischen den Verfahren zur Korrektur von Alphafehler-Kumulierung bei Sequenzdaten besteht und in welchem Maße die Verfahren der Bayesschen Statistik besser für diese geeignet sind (s.h. Kapitel 5). Dabei stellte sich heraus, dass letztere für diese Klasse von Problemen eher verwendet werden sollten, da Alphafehler-Kumulierungskorrekturen möglichen Abhängigkeite zwischen verschiedenen Sequen-zpositionen, welche sich unter Umständen erst im fertigen Protein offenbaren, ignorieren. Weiterhin untersuchte ich HCV Sequenzen mittels einer Variante des Bootstrap-Algorithmus um jene Sequenz-Bereiche zu finden, die im Falle von ungeklärten Übertragungswegen zur Identifizierung dieser genutzt werden können. Dabei stellten sich zwei Bereiche als besonders geeignet heraus: Die hypervariable Region sowie ein Bereich am Ende des Nicht-Struktur Protein NS5B (s.h. Kapitel 9). Die Spaltung von fremden Aminosäuresequenzen innerhalb von menschlichen Zellen durch das Proteasom kann zu einer Präsentation dieser Fragmente auf der Zelloberfläche als Epitope führen. Um solche Fragmente präsentieren zu können, müssen diese nicht nur an das spezifische MHC Molekül binden, sondern auch eine optimale Länge besitzen. Daher sollte der evolutionäre Prozess solche Viren fördern, deren Sequenzen sich nicht in entsprechende Stücke zerteilen lassen. Durch eine Kombination von Epitopdaten aus der IEDB und vorhergesagten viralen Sequenzen, welche sicher an MHC Moleküle binden, untersuchte ich, ob innerhalb der flankierenden Regionen um das jeweilige Epitop Sequenzpositionen existieren, welche auf eine Mutation hinweisen, die den Schnittmechanismus der Zelle verhindert. Ich fand vierzehn Aminosäuren und Positionen, die einen solchen Zusammenhang besitzen können (s.h. Kapitel 7). Um heraus zu finden ob es in der reversen Transkriptase von HBV Positionen gibt, welche die Bindung mit dem nukleotidischen Reverse-Transkriptase-Inhibitor Tenofovir beeinflussen, erstellte ich ein Modell dieses Enzyms. Mutationen, die innerhalb der Bindetasche für Tenofovir lagen, führten in einer Versuchsreihe von der Gruppe von Mengji Lu zu einer verringerten Affinität zw ischen Enzym und Medikament (s.h. Kapitel 10). Zusammen mit der Gruppe von Ralf Küppers untersuchte ich Hoch-Durchsatz-Sequenzdaten von verschiedenen Arten von B Zellen um ähnliche Sequenzen zu finden. Wir fanden ähnliche und sogar identische Sequenzen zwischen zwei, drei und sogar allen vier Arten von Zellen jeweils innerhalb der Blutproben jedes der beiden Spender (s.h Kapitel 6)

Evaluation of Intra-Host Variants of the Entire Hepatitis B Virus Genome

Genetic analysis of hepatitis B virus (HBV) frequently involves study of intra-host variants, identification of which is commonly achieved using short regions of the HBV genome. However, the use of short sequences significantly limits evaluation of genetic relatedness among HBV strains. Although analysis of HBV complete genomes using genetic cloning has been developed, its application is highly labor intensive and practiced only infrequently. We describe here a novel approach to whole genome (WG) HBV quasispecies analysis based on end-point, limiting-dilution real-time PCR (EPLD-PCR) for amplification of single HBV genome variants, and their subsequent sequencing. EPLD-PCR was used to analyze WG quasispecies from serum samples of patients (n = 38) infected with HBV genotypes A, B, C, D, E and G. Phylogenetic analysis of the EPLD-isolated HBV-WG quasispecies showed the presence of mixed genotypes, recombinant variants and sub-populations of the virus. A critical observation was that HBV-WG consensus sequences obtained by direct sequencing of PCR fragments without EPLD are genetically close, but not always identical to the major HBV variants in the intra-host population, thus indicating that consensus sequences should be judiciously used in genetic analysis. Sequence-based studies of HBV WG quasispecies should afford a more accurate assessment of HBV evolution in various clinical and epidemiological settings

Genetic Diversity of Hepatitis B and Hepatitis C Viruses in Ethiopia

Hepatitis B and C viruses are a major cause of mortality and morbidity worldwide. In 2015 alone, viral hepatitis caused 1.34 million deaths, a number comparable to annual deaths caused by tuberculosis but higher than those caused by either HIV or malaria. However, in contrast to these three infectious diseases, viral hepatitis has received relatively little attention. Worldwide, only 9% of HBV-infected and 20% of HCV-infected persons have access to affordable hepatitis testing. Even more worrisome is the fact that only 8% of those diagnosed with HBV and 7.4% of those diagnosed with HCV had started treatment. This is even more problematic in low-income countries like Ethiopia where more than 80% of the population are living in rural areas with a very limited access to healthcare and where viral hepatitis has never been considered a major health priority. Most importantly, the clinical and treatment outcomes of HBV and HCV infections are largely influenced by the high genetic diversity (genotype, subgenotype, recombinants and quasispecies variants) of these viruses that also display distinct geographical distribution worldwide. In this thesis, therefore, we studied the seroepidemiology of HBV and HCV infection and determined the molecular epidemiology and genetic diversity of HBV and HCV in different geographic regions of Ethiopia. We identified that genotype A (A1) and genotype D (D2 and novel D10) of HBV and genotype 4 ( 4d and 4r ) of HCV are the most prevalent in Ethiopia. We further analyzed HBV and HCV quasispecies variants and their association with patients’ clinical characteristics using ultra-deep sequencing approach and found some important variants

Audit of Antenatal Testing of Sexually Transmissible Infections and Blood Borne Viruses at Western Australian Hospitals

In August 2007, the Western Australian Department of Health (DOH) released updated recommendations for testing of sexually transmissible infections (STI) and blood-borne viruses (BBV) in antenates. Prior to this, the Royal Australian & New Zealand College of Obstetricians & Gynaecologists (RANZCOG) antenatal testing recommendations had been accepted practice in most antenatal settings. The RANZCOG recommends that testing for HIV, syphilis, hepatitis B and C be offered at the first antenatal visit. The DOH recommends that in addition, chlamydia testing be offered. We conducted a baseline audit of antenatal STI/BBV testing in women who delivered at selected public hospitals before the DOH recommendations. We examined the medical records of 200 women who had delivered before 1st July 2007 from each of the sevenWAhospitals included in the audit. STI and BBV testing information and demographic data were collected. Of the 1,409 women included, 1,205 (86%) were non-Aboriginal and 200 (14%) were Aboriginal. High proportions of women had been tested for HIV (76%), syphilis (86%), hepatitis C (87%) and hepatitis B (88%). Overall, 72% of women had undergone STI/BBV testing in accordance with RANZCOG recommendations. However, chlamydia testing was evident in only 18% of records. STI/BBV prevalence ranged from 3.9% (CI 1.5– 6.3%) for chlamydia, to 1.7% (CI 1–2.4%) for hepatitis C, 0.7% (CI 0.3–1.2) for hepatitis B and 0.6% (CI 0.2–1) for syphilis. Prior to the DOH recommendations, nearly three-quarters of antenates had undergone STI/BBV testing in accordance with RANZCOG recommendations, but less than one fifth had been tested for chlamydia. The DOH recommendations will be further promoted with the assistance of hospitals and other stakeholders. A future audit will be conducted to determine the proportion of women tested according to the DOH recommendations. The hand book from this conference is available for download Published in 2008 by the Australasian Society for HIV Medicine Inc © Australasian Society for HIV Medicine Inc 2008 ISBN: 978-1-920773-59-