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Made available in DSpace on 2022-04-29T08:38:06Z (GMT). No. of bitstreams: 0 Previous issue date: 2022-03-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Strongyloidiasis, a parasitosis caused by Strongyloides stercoralis in humans, is a very prevalent infection in tropical or subtropical areas. Gaps on public health strategies corroborates to the high global incidence of strongyloidiasis especially due to challenges involved on its diagnosis. Based on the lack of a gold-standard diagnostic tool, we aimed to present a metabolomic study for the assessment of stool metabolic alterations. Stool samples were collected from 25 patients segregated into positive for strongyloidiasis (n = 10) and negative control (n = 15) and prepared for direct injection high-resolution mass spectrometry analysis. Using metabolomics workflow, 18 metabolites were annotated increased or decreased in strongyloidiasis condition, from which a group of 5 biomarkers comprising caprylic acid, mannitol, glucose, lysophosphatidylinositol and hydroxy-dodecanoic acid demonstrated accuracy over 89% to be explored as potential markers. The observed metabolic alteration in stool samples indicates involvement of microbiota remodeling, parasite constitution, and host response during S. stercoralis infection. Innovare Biomarkers Laboratory School of Pharmaceutical Sciences University of Campinas Department of Internal Medicine Botucatu Medical School São Paulo State University Department of Animal Biology Institute of Biology University of Campinas Department of Clinical Pathology School of Medical Sciences University of Campinas Department of Internal Medicine Botucatu Medical School São Paulo State University FAPESP: 2016/07137-0 FAPESP: 2019/05718-3 to JD

Finding specific molecular targets and the mechanism of action of praziquantel in the treatment of schistosomiasis remains a challenging task. Our efforts were focused on obtaining further information on worm composition before and after exposure to praziquantel in the treatment of schistosomiasis to elucidate the potential sites of action of this drug. Evidence indicates that the lipid bilayer is changed by treatment with praziquantel. Following this rationale, we employed a mass spectrometry imaging-based approach that helped to characterise lipids in specific locations, which are directly involved in the biochemical pathways of the BH strain of Schistosoma mansoni, as well as differentiating the molecular response that each worm sex presents in vivo. Our findings demonstrated significant differences between the chemical markers found in adult worms before and after praziquantel exposure, especially in phospholipids, which were predominantly identified as chemical markers in all samples. Results also indicate that distinct molecular pathways in both male and female worms could be differentially affected by praziquantel treatment. These data shine new light on the mechanism of action of praziquantel, taking a further step towards its full understanding.

Background: Cystic fibrosis is a disabling genetic disease with an increased prevalence in European heritage populations. Currently, the most used technique for collection of cystic fibrosis samples and diagnosis is provided through uncomfortable tests, with uncertain results, mostly based on chloride concentration in sweat. Since cystic fibrosis mutation induces many metabolic changes in patients, exploring these alterations might be an alternative to visualize potential biomarkers that could be used as interesting tools for further diagnostic upgrade, prioritizing simplicity, low cost and quickness. Methods: This contribution describes an accurate strategy to provide potential biomarkers related to cystic fibrosis, which may be understood as a potential tool for new diagnostic approaches and/or for monitoring disease evolution. Therefore, the present proposal consists of using skin imprints on silica plates as a way of sample collection, followed by direct-infusion high-resolution mass spectrometry and multivariate data analysis, intending to identify metabolic changes in skin composition of cystic fibrosis patients. Results: Metabolomics analysis allowed identifying chemical markers that can be traced back to cystic fibrosis in patients’ skin imprints, differently from control subjects. Seven chemical markers from several molecular classes were elected, represented by bile acids, a glutaric acid derivative, thyrotropin releasing hormone, an inflammatory mediator, a phosphatidic acid, and diacylglycerol isomers, all reflecting metabolic disturbances that occur due to of cystic fibrosis. Conclusion: The comfortable method of sample collection combined with the identified set of biomarkers represent potential tools that open the range of possibilities to manage cystic fibrosis and follow the disease evolution. This exploratory approach points to new perspectives about the development of diagnostic assay using biomarkers and the management cystic fibrosis.

Made available in DSpace on 2018-12-11T16:44:06Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-10-01 Recent outbreaks of Zika virus in Oceania and Latin America, accompanied by unexpected clinical complications, made this infection a global public health concern. This virus has tropism to neural tissue, leading to microcephaly in newborns in a significant proportion of infected mothers. The clinical relevance of this infection, the difficulty to perform accurate diagnosis and the small amount of data in literature indicate the necessity of studies on Zika infection in order to characterize new biomarkers of this infection and to establish new targets for viral control in vertebrates and invertebrate vectors. Thus, this study aims at establishing a lipidomics profile of infected mosquito cells compared to a control group to define potential targets for viral control in mosquitoes. Thirteen lipids were elected as specific markers for Zika virus infection (Brazilian strain), which were identified as putatively linked to the intracellular mechanism of viral replication and/or cell recognition. Our findings bring biochemical information that may translate into useful targets for breaking the transmission cycle. Innovare Biomarkers Laboratory School of Pharmaceutical Sciences University of Campinas Animal Viruses Laboratory Department of Genetics Evolution and Bioagents Institute of Biology University of Campinas Emerging Viruses Study Laboratory Department of Genetics Evolution and Bioagents Institute of Biology University of Campinas Vector Functional Genomics and Microbiology Laboratory UNESP Institute of Biotechnology São Paulo State University, Alameda das Tecomarias s/n Department of Bioprocesses and Biotechnology Faculty of Agronomical Sciences São Paulo State University, Rua José Barbosa 1780 Vector Functional Genomics and Microbiology Laboratory UNESP Institute of Biotechnology São Paulo State University, Alameda das Tecomarias s/n Department of Bioprocesses and Biotechnology Faculty of Agronomical Sciences São Paulo State University, Rua José Barbosa 1780

Ascaris lumbricoides is responsible for a highly disseminated helminth parasitic disease, ascariosis, a relevant parasitosis that responds for great financial burden on the public health system of developing countries. In this work, metabolic fingerprinting using high-resolution mass spectrometry (HRMS) was employed to identify marker molecules from A. lumbricoides in different development stages. We have identified nine biomarkers, such as pheromones and steroidal prohormones in early stages, among other molecules in late development stages, making up four molecules for fertilized eggs, four marker molecules for first larvae (L1) and one marker molecule for third larvae (L3). Therefore, our findings indicate that this approach is suitable for biochemical characterization of A. lumbricoides development stages. Moreover, the straightforward analytical method employed, with almost no sample preparation from a complex matrix (feces) using high-resolution mass spectrometry, suggests that it is possible to seek for an easier and faster way to study animal molding processes.

Leprosy is a chronic infectious disease caused by Mycobacterium leprae, which primarily infects macrophages and Schwann cells, affecting skin and peripheral nerves. Clinically, the most common form of identification is through the observation of anesthetic lesions on skin; however, up to 30% of infected patients may not present this clinical manifestation. Currently, the gold standard diagnostic test for leprosy is based on skin lesion biopsy, which is invasive and presents low sensibility for suspect cases. Therefore, the development of a fast, sensible and noninvasive method that identifies infected patients would be helpful for assertive diagnosis. The aim of this work was to identify lipid markers in leprosy patients directly from skin imprints, using a mass spectrometric analytical strategy. For skin imprint samples, a 1 cm(2) silica plate was gently pressed against the skin of patients or healthy volunteers. Imprinted silica lipids were extracted and submitted to direct-infusion electrospray ionization high-resolution mass spectrometry (ESI-HRMS). All samples were differentiated using a lipidomics-based data workup employing multivariate data analysis, which helped electing different lipid markers, for example, mycobacterial mycolic acids, inflammatory and apoptotic molecules were identified as leprosy patients' markers. Otherwise, phospholipids and gangliosides were pointed as healthy volunteers' skin lipid markers, according to normal skin composition. Results indicate that silica plate skin imprinting associated with ESI-HRMS is a promising fast and sensible leprosy diagnostic method. With a prompt leprosy diagnosis, an early and effective treatment could be feasible and thus the chain of leprosy transmission could be abbreviated.

Orientador: Marcelo Lancellotti Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia Resumo: Neisseria meningitidis, bactéria aeróbica Gram negativa, causa infecções meningocócicas que podem resultar em doenças severas incluindo meningite meningocócica, sepsis e morte. Uma vez que a administração precoce de antibióticos está relacionada às baixas taxas de morbidade e mortalidade, é de crucial importância iniciar o tratamento o mais rápido possível. A administração de antibióticos aos pacientes com suspeita de infecções meningocócicas antes da coleta de material biológico tem sido uma prática comum que pode prejudicar o diagnóstico microbiológico por cultura bacteriana. A abordagem nos últimos anos tem sido o desenvolvimento de técnicas alternativas que possam ser utilizadas para se obter o diagnóstico etiológico. Técnicas de biologia molecular, como a Reação em Cadeia da Polimerase (PCR) tem sido desenvolvidas para melhorar a sensibilidade e especificidade da detecção de microorganismo e aliadas a Nanotecnologia podem ser úteis para desenvolvimento de métodos alternativos de diagnóstico. Foi padronizada PCR para N. meningitidis ATCC 23247 Sorogrupo B com a utilização de primers do gene synD, através de gradiente de MgCl2. Também foi determinado o Limite de Detecção (LD) de DNA da PCR. Então a reação foi adjuvada com nanopartículas de sílica mesoposa SAB-15 e SBA-16. Não houve aumento do LD de DNA nas concentrações de nanopartículas testadas. Também foi realizado estudo entre a interação do DNA e SBA-15 e SBA-16 através de Gel Retardo e Dicroísmo Circular. Resultados sugerem interação fraca entre DNA e estas nanopartículas. Desta forma conclui-se que nanopartículas SBA-15 e SBA-16 não possuem interação com DNA a ponto de melhorar seu LD, não sendo úteis, nas concentrações testadas, para o desenvolvimento de métodos diagnósticos mais sensíveis Abstract: Neisseria meningitidis , Gram-negative aerobic bacteria , which cause meningococcal disease can result in severe diseases including meningococcal meningitis , sepsis and death. Since the early administration of antibiotics is related to lower rates of morbidity and mortality, it is crucial to start treatment as soon as possible . The administration of antibiotics to patients with suspected meningococcal disease before the collection of biological material has been a common practice that can harm the microbiological diagnosis of bacterial culture . The approach in recent years has been the development of alternative techniques that might be used to obtain the etiologic diagnosis. Molecular biology techniques such as the Polymerase Chain Reaction (PCR ) has been developed to improve the sensitivity and specificity of detection of a microorganism and allied Nanotechnology may be useful for development of alternative methods of diagnosis. PCR was standardized for N. ATCC 23247 meningitidis Serogroup B using primers gene SynD, through MgCl2 gradient . Also determined was the limit of detection (LD) of PCR DNA . Then the reaction was adjuvanted with silica nanoparticles mesoposa SAB- 15 and SBA - 16 . There was no increase in LD DNA concentrations of nanoparticles tested. Also it was studied the interaction between DNA and SBA - 15 and SBA - 16 by Gel Retardation and Circular Dichroism . Results suggest weak interaction between DNA and these nanoparticles . Thus it is concluded that nanoparticles SBA-15 and SBA-16 does not have interaction with DNA improve their point of LD and is not useful in concentrations, for the development of more sensitive diagnostic methods Mestrado Fármacos, Medicamentos e Insumos para Saúde Mestra em Biociências e Tecnologia de Produtos Bioativos