Metagenomic analysis of the saliva microbiome with merlin

In recent years, metagenomics has demonstrated to play an essential role on the study of the microorganisms that live in microbial communities, particularly those who inhabit the human body. Several bioinformatics tools and pipelines have been developed for the analysis of these data, but they usually only address one topic: to identify the taxonomic composition or to address the metabolic functional profile. This work aimed to implement a computational framework able to answer the two questions simultaneously. Merlin, a previously released software aiming at the reconstruction of genome-scale metabolic models for single organisms, was extended to deal with metagenomics data. It has an user-friendly and intuitive interface, being suitable for those with limited bioinformatics skills. The performance of the tool was evaluated with samples from the Human Microbiome Project, particularly from saliva. Overall, the results show the same patterns reported before: while the pathways needed for microbial life remain relatively stable, the community composition varies extensively among individuals

Taxonomic and functional analysis of metagenomes

Dissertação de mestrado em BioinformáticaOver the years, metagenomics has demonstrated to play an essential role on the study of the microorganisms that live in microbial communities, particularly those who inhabit the human body. Several bioinformatic tools and pipelines have been developed, but usually they only address one question: "Who is there?" or "What are they doing?". This work aimed to develop a computational framework to answer the two questions simultaneously, that is, perform a taxonomic and functional analysis of microbial communities. Merlin, a previously developed software designed for the construction of genome-scale metabolic models for single organisms, was extended to deal with metagenomics data. It has an userfriendly and intuitive interface, not requiring command-line knowledge and further libraries dependencies or installation, as many other tools. The extended version of Merlin can predict the taxonomic composition of an environmental sample based on the results of homology searches, where the proportions of phyla and genera present are discriminated. Regarding the metabolic analysis, it allows to identify which enzymes are present and calculate their abundance, as well as to nd out which metabolic pathways are e ectively present. The performance of the tool was evaluated with samples from the Human Microbiome Project, particularly from the saliva. The taxonomic membership predicted in Merlin was in agreement with other tools, despite some di erences in the proportions. The functional characterization showed a conserved pool of pathways through di erent samples, although Merlin sometimes presented less pathways than expected because the routine is highly dependent on the enzymes annotation. Overall, the results showed the same pattern as reported before: while the pathways needed for microbial life remain relatively stable, the community composition varies extensively among individuals. In the end, Merlin demonstrated to be a reliable standalone alternative to web services for those scientists that have concerns about sharing data.Ao longo dos anos, a metagenómica demonstrou ter um papel essencial no estudo dos microorganismos que vivem em comunidades bacterianas, particularmente aqueles que habitam o corpo humano. Várias ferramentas e pipelines bioinformáticas foram desenvolvidas, mas normalmente estas apenas abordam uma destas questões: "Quem está lá?" ou "O que é que estão a fazer?" Este trabalho teve como objectivo o desenvolvimento duma ferramenta computacional para responder aos dois problemas em simultâneo, isto é, realizar tanto uma análise taxonómica como funcional de comunidades microbianas. O Merlin, um software anteriormente desenvolvido para construir modelos metabólicos à escala genómica para um organismo, foi estendido para tratar dados de metagenómica. O programa possui uma interface intuitiva e amiga do utilizador, não necessitando de conhecimentos de linha de comandos nem de dependências de bibliotecas ou instalação de aplicações adicionais. Esta versão estendida do Merlin prevê a composição taxonómica global dum metagenoma baseado nos resultados de procuras de sequências homólogas, onde as proporções dos fila e géneros são apresentadas. No que diz respeito à análise metabólica, o Merlin permite identificar quais as enzimas presentes e calcular a sua abundância, bem como identificar quais as vias metabólicas que estão efectivamente presentes. O desempenho da ferramenta foi avaliado com amostras do Projecto do Microbioma Humano, particularmente com amostras da saliva. A composição taxonómica prevista no Merlin esteve de acordo com outras ferramentas, apesar de algumas diferenças observadas nas proporções. A caracterização funcional mostrou um conjunto conservado de vias metabólicas nas diferentes amostras, mesmo que o Merlin tenha identificado menos enzimas que o esperado, pois o método é bastante dependente do processo anotação. Globalmente, os resultados revelaram o mesmo padrão reportado anteriormente: enquanto as vias metabólicas necessárias para a vida microbiana se mantêm estáveis, a composição taxonómica varia bastante entre indivíduos. No final, o Merlin demonstrou ser uma alternativa fidedigna a serviços web para aqueles cientistas que têm restrições em divulgar os seus dados não publicados num website.Fundação para a Ciência e a Tecnologia (FCT) - Projeto COMPETE FCOMP-01-0124-FEDER-015079.ERDF - European Regional Development Fund atrav es do programa COMPETE (programa operacional para a competitividade)

Fecal microRNAs as innovative biomarkers of intestinal diseases and effective players in host-microbiome interactions

Over the past decade, short non-coding microRNAs (miRNAs), including circulating and fecal miRNAs have emerged as important modulators of various cellular processes by regulating the expression of target genes. Recent studies revealed the role of miRNAs as powerful biomarkers in disease diagnosis and for the development of innovative therapeutic applications in several human conditions, including intestinal diseases. In this review, we explored the literature and summarized the role of identified dysregulated fecal miRNAs in intestinal diseases, with particular focus on colorectal cancer (CRC) and celiac disease (CD). The aim of this review is to highlight one fascinating aspect of fecal miRNA function related to gut microbiota shaping and bacterial metabolism influencing. The role of miRNAs as "messenger" molecules for inter kingdom communications will be analyzed to highlight their role in the complex host-bacteria interactions. Moreover, whether fecal miRNAs could open up new perspectives to develop novel suitable biomarkers for disease detection and innovative therapeutic approaches to restore microbiota balance will be discussed

The Artificial Intelligence Workbench: a retrospective review

Last decade, biomedical and bioinformatics researchers have been demanding advanced and user-friendly applications for real use in practice. In this context, the Artificial Intelligence Workbench, an open-source Java desktop application framework for scientific software development, emerged with the goal of provid-ing support to both fundamental and applied research in the domain of transla-tional biomedicine and bioinformatics. AIBench automatically provides function-alities that are common to scientific applications, such as user parameter defini-tion, logging facilities, multi-threading execution, experiment repeatability, work-flow management, and fast user interface development, among others. Moreover, AIBench promotes a reusable component based architecture, which also allows assembling new applications by the reuse of libraries from existing projects or third-party software. Ten years have passed since the first release of AIBench, so it is time to look back and check if it has fulfilled the purposes for which it was conceived to and how it evolved over time

Daily intake of Lactobacillus gasseri CP2305 relieves fatigue and stress-related symptoms in male university Ekiden runners : A double-blind, randomized, and placebo-controlled clinical trial

The heat-inactivated, enteric-colonizing Lactobacillus gasseri CP2305 (CP2305) ameliorates psychological stress-related symptoms. In this study, we examined effects of CP2305 on top athletes experiencing physical and mental stresses. Forty-nine male university Ekiden (long distance relay race) runners daily took the CP2305-containing beverage for 12 weeks during training for and competing in All-Japan university championships. The CP2305 intake significantly facilitated recovery from fatigue and relieved anxiety and depressive mood, compared with placebo intake. The CP2305 intake significantly prevented the training-induced reduction of hemoglobin and facilitated exercise-induced increase in serum growth hormone levels. The CP2305 intake significantly increased the alpha- and beta-diversities of fecal microbiota, and the compositions of Bifidobacterium and Faecalibacterium. Gene expression profiling of peripheral blood leukocytes indicated that CP2305 prevented the stress-induced changes in the expression of genes related to mitochondrial functions. Our results suggest that daily intake of paraprobiotic CP2305 may be beneficial to athletes facing stressful situations

Antibiotic Resistance in Oral Streptococci: The prevalence, diversity, stability, and fitness cost of Tn916 -Tn1545 family in oral streptococcal isolates

Oral streptococci are important members of the oral microbiome that are gathering more attention due to the presence of antimicrobial resistance determinants and their association with mobile genetic elements. Mobile genetic elements such as the ICE, Tn916 are abundant in bacterial populations and have been implicated in the spread of antibiotic resistance. Despite their abundance, little is known regarding the presence, diversity, and evolutionary dynamics of these ICEs in oral streptococci. The work presented in this thesis addresses the prevalence, diversity, stability, and fitness cost of the Tn916 family in oral streptococcal isolates. We report that the wild-type Tn916 is the most prevalent member of this family detected was wild-type Tn916. In terms of diversity, we detected two additional members of this family of ICEs, besides Tn916, namely Tn6815 and Tn6816. As the conjugative transfer of these Tn916-Tn1545 family elements was high, we assessed the effect these elements have on the new host (Streptococcus oralis) in the absence of selective pressure. We, therefore, determined the relative fitness cost associated with the acquisition of these elements and their stability in the absence of selective pressure. The findings of this work showed that upon acquisition Tn916, imposes a reduction in relative fitness ranging from 6% to 25% at time zero (T0). Amelioration of the observed fitness cost was observed within 500 generations, and the ICEs were stable in the absence of selection for up to 1000 generations. Analysis of the number of copies of Tn916 in the transconjugants with both digital droplet PCR (ddPCR) copy number analysis and whole genome sequencing (WGS) indicated no changes in the copy number, elements sequence and insertion sites in the evolution experiment. Taken together, the findings of this work reaffirm the role that Tn916 elements play in the spread of antibiotic resistance among oral streptococci

Metagenomic analysis and culture-based methods to examine the prevalence and distribution of antimicrobial resistance on two New Zealand dairy farms : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Veterinary Science at Massey University, Palmerston North, New Zealand

Antimicrobial resistance (AMR) is a global threat to human and animal health, with the misuse and overuse of antimicrobials being suggested as the main driver of resistance. In a global context, New Zealand (NZ) is a relatively low user of antimicrobials in animal production. However, antimicrobial usage on NZ dairy farms and its potential for driving the spread of AMR within the dairy farm environment is under-researched. This research addresses the hypothesis that antimicrobial use on NZ dairy farms influences the prevalence of AMR in dairy farm environments, taking into consideration seasonality and contrasting farm management practices. The aims of this study, focused on two NZ dairy farm environments over an 15 month period, were to (i) determine the prevalence and distribution of AmpC- and extended-spectrum β-lactamase (ESBL)-producing Escherichia coli, utilising culture-based methods, and (ii) to determine the abundance and diversity of antimicrobial resistance genes (ARGs), utilising a metagenomic approach, and lastly (iii), to assess the impact of systemic antimicrobial treatment on the bovine faecal microbiome. Overall, the research presented in this thesis has shown a low sample level prevalence of ESBL-producing E. coli from two NZ dairy farms (faeces 0%, 1.7%; farm dairy effluent (FDE) 0%, 6.7% from Dairy 1 and Dairy 4, respectively) but AmpC-producing E. coli were more frequently isolated across both farms (faeces 3.3%, 8.3%; FDE 38.4%, 6.7% from Dairy 1 and Dairy 4, respectively). AmpC- and ESBL-producing E. coli were isolated in spring and summer, during months with varying levels of antimicrobial use. Analysis at the individual animal level showed a decrease in bacterial diversity and richness during systemic antimicrobial treatment and in many cases the microbiome diversity recovered post-treatment when the cow re-entered the milking herd. Compared to overseas data in a similar context, NZ dairy farm environments had a low abundance of ARGs, with the highest abundance detected in soil (0.20 - 0.63 copies of ARG per 16S rRNA gene). However, many of the ARGs identified in soil are not frequently found in human pathogens or acquired genes. FDE had a lower ARG abundance but the ARGs were more diverse (0.03 - 0.37 copies of ARG per 16S rRNA gene). There was no association between the normalised ARG abundance and antimicrobial use or collection date, however the low ARG abundance in the farm samples may have made any associations difficult to detect. AMR is a burden for human, animal and environmental health and requires a holistic "One Health" approach to address. The outcomes from this research improve our understanding of the current levels of AMR on two NZ dairy farms and identifies areas for future research. Prevention is better than a cure and urgent action is required to slow the development and dissemination of AMR and to improve antimicrobial stewardship in humans and animals

Research needs for optimising wastewater-based epidemiology monitoring for public health protection

This is the final version. Available on open access from IWA Publishing via the DOI in this recordData availability statement: All relevant data are included in the paper or its Supplementary Information.Wastewater-based epidemiology (WBE) is an unobtrusive method used to observe patterns in illicit drug use, poliovirus, and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The pandemic and need for surveillance measures have led to the rapid acceleration of WBE research and development globally. With the infrastructure available to monitor SARS-CoV-2 from wastewater in 58 countries globally, there is potential to expand targets and applications for public health protection, such as other viral pathogens, antimicrobial resistance (AMR), pharmaceutical consumption, or exposure to chemical pollutants. Some applications have been explored in academic research but are not used to inform public health decision-making. We reflect on the current knowledge of WBE for these applications and identify barriers and opportunities for expanding beyond SARS-CoV-2. This paper critically reviews the applications of WBE for public health and identifies the important research gaps for WBE to be a useful tool in public health. It considers possible uses for pathogenic viruses, AMR, and chemicals. It summarises the current evidence on the following: (1) the presence of markers in stool and urine; (2) environmental factors influencing persistence of markers in wastewater; (3) methods for sample collection and storage; (4) prospective methods for detection and quantification; (5) reducing uncertainties; and (6) further considerations for public health use.Natural Environment Research Council (NERC)Engineering and Physical Sciences Research Council (EPSRC